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Frankly I am a programmer, I've been looking at sites like NCBI and others, but couldn't find what I was looking for so if someone can help:
Can you give me the probabilities of
- mutations in Saccharomyces Cerevisiae's telomeres.
- Probability of having an insertion
- Probability of having a deletion.
- Probability of having a substitution.
The reason I need this is that I want to generate true-telomeric data based on, the telomere structure consensus.
The TERT gene provides instructions for making one component of an enzyme called telomerase. Telomerase maintains structures called telomeres, which are composed of repeated segments of DNA found at the ends of chromosomes. Telomeres protect chromosomes from abnormally sticking together or breaking down (degrading). In most cells, telomeres become progressively shorter as the cell divides. After a certain number of cell divisions, the telomeres become so short that they trigger the cell to stop dividing or to self-destruct (undergo apoptosis). Telomerase counteracts the shortening of telomeres by adding small repeated segments of DNA to the ends of chromosomes each time the cell divides.
In most types of cells, telomerase is either undetectable or active at very low levels. However, telomerase is highly active in cells that divide rapidly, such as cells that line the lungs and gastrointestinal tract, cells in bone marrow, and cells of the developing fetus. Telomerase allows these cells to divide many times without becoming damaged or undergoing apoptosis. Telomerase is also abnormally active in most cancer cells, which grow and divide without control or order.
The telomerase enzyme consists of two major components that work together. The component produced from the TERT gene is known as hTERT. The other component is produced from a gene called TERC and is known as hTR. The hTR component provides a template for creating the repeated sequence of DNA that telomerase adds to the ends of chromosomes. The hTERT component then adds the new DNA segment to chromosome ends.
Unlike gray hair, one of the most significant signs of aging is invisible to the naked eye. Deep inside cells, at the tips of thread-like chromosomes, structures known as telomeres protect chromosomes from deterioration—a bit like the way caps at the ends of shoelaces prevent fraying. Telomeres naturally shorten as people age.
But sometimes, an inherited gene mutation causes telomeres to shorten at a faster rate. Abnormal telomere shortening results in accelerated-aging syndromes that affect many parts of the body and can occur in children or adults. The severity of short telomere syndromes varies, but they increase cancer risk and can lead to organ failure and death.
With help from the Center for Individualized Medicine, Mayo Clinic uses a precision-medicine approach to manage short telomere syndromes. Mayo's Premyeloid & Bone Failure Disorder Clinic provides diagnostic testing, multidisciplinary treatment and genetic counseling for short telomere syndromes.
"Short telomere syndromes have been recognized for several decades. But diagnosis has been very difficult because it requires highly specialized testing. With the advent of precision genomics, we have the opportunity to identify and manage these disorders, for the benefit of patients," says Mrinal Patnaik, M.B.B.S., a hematologist who directs the premyeloid disorder clinic.
Although short telomere syndromes are considered rare, Mayo Clinic sees five to seven people with the disorders a month. "We think short telomeres are much more common than has been reported, and anticipate that these new precision-medicine tools will bring a fair number of cases to light," Dr. Patnaik says.
'The mysterious telomere'
The 2009 Nobel Prize in medicine was awarded for discoveries about what the Nobel committee called "the mysterious telomere." Short telomeres affect parts of the body where stem cells actively divide, including bone marrow, skin and the tissues lining the lungs and digestive tract.
"These stem cells rely on telomeres to keep their integrity. Short telomeres cause the stem cells to prematurely die," Dr. Patnaik says.
Short telomeres can lead to scarring in the lungs and liver, narrowing of the digestive tract, bone marrow failure and immune-system deficiency. The severity of the inherited condition increases with each generation—a phenomenon known as genetic anticipation.
"Children inherit from their parents not only the genetic mutation causing short telomere syndrome but also shorter and shorter telomeres," Dr. Patnaik says. "As a result, the syndromes tend to occur at a younger age and with more severe manifestations in each generation. Eventually the life span is highly limited."
Initial diagnosis is a challenge because the signs and systems of short telomere syndromes are diverse. "There is a lack of awareness," Dr. Patnaik says. "A person with lung fibrosis and failing bone marrow might see a lung specialist or a blood specialist who isn't familiar with these multispecialty syndromes and doesn't put the clues together."
Mayo Clinic looks for certain signs and symptoms with unexplained causes, including:
- Personal or family history of premature graying of hair
- Low red blood cell, white blood cell or platelet counts
- Thickened, stiff or scarred lung or liver tissue
If short telomere syndrome is suspected, Mayo Clinic can arrange for sophisticated testing that measures the length of telomeres in an individual's blood cells. Once short telomeres are identified, Mayo Clinic has a genetic sequencing panel to help find the mutation causing short telomeres. If genetic sequencing doesn't uncover a mutation, whole exome sequencing—which looks at all disease-causing genes in an individual's DNA blueprint—can be performed.
Certain genetic mutations are known to be associated with short telomeres. But only 40 to 50 percent of people with short telomeres have one of these known mutations.
"The fact that more than half our patients with short telomeres do not have detectable gene mutations on sequencing panels indicates that we haven't yet discovered all the mutations that affect telomere length," Dr. Patnaik says. "There also may be nongenetic mechanisms involved—which is a Pandora's box we haven't even opened yet."
Seeking new treatment options
Recent research indicates that treatment with danazol, an anabolic steroid, may slow the rate of telomere shortening, as well as improve blood counts and stabilize lung and liver disease in people with short telomeres. Laboratory experiments with gene therapies are also underway. Mayo Clinic is involved in research efforts in both these areas.
"Unfortunately, while we increasingly understand the genetics and consequences of short telomeres, much work remains to be done with regards to effective treatment modalities," Dr. Patnaik says.
Transplantation can be an option for people who experience organ failure. However, individuals with short telomere syndrome often need multiorgan transplants: bone marrow transplantation as well as a liver or lung transplant, which makes the process extremely challenging.
"The vast majority of people with short telomere syndrome are turned down for transplant because not many centers are equipped to perform multiorgan transplantation," Dr. Patnaik says. "Our precision genomics clinic is working with the various transplant groups within Mayo Clinic to pursue safe multiorgan transplantation for these patients. At Mayo Clinic, we have a great opportunity to use precision medicine to benefit people with short telomere syndromes."
This post originally appeared on the Center for Individualized Medicine blog on Feb. 14, 2019.
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Older Dads Give Good Telomeres, But Longevity? Not So Much
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Human chromosomes (grey) capped by telomeres (white) U.S. Department of Energy Human Genome Program
, “Telomeres just grabbed me and kept leading me on.” And lead her on they did---all the way to the Nobel Prize in Medicine in 2009
. Telomeres are DNA sequences that continue to fascinate researchers and the public, partially because people with longer telomeres
tend to live longer. So the recent finding that older men father offspring with unusually lengthy telomeres sounds like great news. Men of advanced age will give their children the gift of longer lives---right? But as is so often the case in biology, things aren't that simple, and having an old father may not be an easy route to a long and healthy life. Every time a piece of DNA gets copied, it can end up with errors in its sequence, or mutations. One of the most frequent changes is losing scraps of information from each end of the strand. Luckily, these strands are capped with telomeres, repeating sequences that do not code for any proteins and serve only to protect the rest of the DNA. Each time the DNA makes a copy, its telomeres get shorter, until these protective ends wear away to nothing. Without telomeres, the DNA cannot make any more copies, and the cell containing it will die. But sperm are not subject to this telomere-shortening effect. In fact, the telomeres in sperm-producing stem cells not only resist degrading, they actually grow. This may be thanks to a high concentration of the telomere-repairing enzyme telomerase in the testicles researchers are still uncertain. All they know is that the older the man, the longer the telomeres in his sperm will be. And a recent study
in the Proceedings of the National Academy of Sciences found that these long telomeres can be passed down to offspring: The children of older fathers are born with longer telomeres in all their cells. In fact, the long telomeres can carry over for two generations, with older paternal grandfathers passing their long telomeres to both their children and their sons’ children. The longest telomeres of all went to children whose fathers and fathers' fathers had reproduced at advanced ages. (But daughters of older fathers didn’t pass on the extra-long telomeres at all.) Getting longer telomeres is exciting because telomere length is associated with longevity. A telomere that shrinks with each cell division is like a ticking clock counting down the cell’s life when a father passes longer telomeres to his offspring, he’s essentially giving their cellular clocks more ticks. So children of older fathers should live longer! There’s just one problem: they don’t. Some [ pdf
] of historical data on longevity have found that older fathers, particularly those who reproduce beyond the age of 65, are actually associated with a shorter lifespan for their children, especially their daughters. Middle-aged offspring of men in the oldest age group were 60 percent more likely to die than the progeny of younger men. The children of older dads are also more likely to have lower IQs
. Why aren’t the longer telomeres granting them longer, better lives?
The issue is that sperm cells aren’t only gaining telomere length---they’re also picking up more mutations. Unlike women, whose eggs are generally created all at once, while a female is still in utero
, men produce sperm throughout their lives. And every time a sperm stem cell begins the process of spawning new sperm, it has to divide, creating the opportunity for mutations to form. The sperm produced by older sperm stem cells will have longer telomeres, but they will also have a greater load of mutations, which tends to shorten offspring lifespan. So the question is: Between telomere length and mutation load, which one dominates the offspring’s chances of a long life? Because studies of historical data found that offspring of older fathers die earlier, you might think that the mutation load wins. But this may not be entirely correct. First of all, fathers’ ages are far from the only contributors to their offspring's health. For example, data for these longevity studies was collected in the 1800s and 1900s, before reliable birth control. Fathers often had many children before middle age, which means the offspring of older fathers were likely to have many siblings. The tenth child of an older father would receive less attention, fewer resources, and perhaps less of an opportunity for long life than, say, an only child born today whose parents chose to wait a while before reproducing. Social factors and available resources play a big part in a child’s lifespan. In contrast with the studies that used historical data, a 2009 study of modern elderly Canadians
found no connection between the seniors’ likelihood of dying and their parents’ ages. However, this study only looked at the offspring of men between the ages of 25 and 45. It may be that the adverse effect of genetic mutations on offspring longevity only becomes important for men over a certain age. After all, one of the historical studies found the effect to be much more significant in the oldest age group: men who reproduced after the age of 65. This suggests there may be a sweet spot, a paternal age that maximizes telomere length while minimizing mutations, that would be an ideal time for reproduction---at least, in terms of genetic quality. But even if there is a slight advantage to having middle-aged but not old fathers, it's not likely to be a big advantage, or we would see some positive effect in longevity data. Ultimately, telomeres and mutations don’t determine exactly how or when we die. These genetic factors shape our lives, but they don't control our destinies.
The small GTPase protein KRAS is a signal-transducing protein, which binds GDP in its inactive state and GTP in its active state . The gene KRAS is frequently mutated in various human cancers. The mutation is most often, in about 86% of the cases , found at G12. In fact, every missense mutation at G12 (G12X) is oncogenic. The oncogenic properties associated with KRAS G12X mutation are characterized by the deficiency of the intrinsic GTPase activity and the insensitivity for GTPase-activating proteins (GAPs) [3,4]. These alterations lead to increased KRAS signaling, as there is more active GTP-bound protein present. Still, the mutant KRAS undergoes GDP–GTP cycling . The basis of the specific G12X mutation frequencies has remained unclear, except for the G12C transversion mutation (c.34G>T) associated with smoking in lung cancer [6,7].
An interesting discrepancy among KRAS G12X mutants is observed in their intrinsic GTPase activity . The G12A mutation exhibits the most hindered intrinsic hydrolysis (
1% compared to the wild-type), whereas the G12C mutation displays the least hindered activity (
72%). All G12X mutants, however, show insensitivity to GAPs that accelerate hydrolysis . Importantly, not only RAS G12X mutants exhibit a discrepancy in GTP hydrolysis, but they also give rise to differences in the preferred signaling pathway (in terms of effector protein binding) [9,10]. This behavior was first observed in NSCLC cell lines , where KRAS G12D showed activation of PI3K and MEK signaling, while G12C and G12V mutants exhibited activated RalGDS-pathway and diminished growth factor-dependent Akt activation. Furthermore, an NMR study revealed different binding preferences for mutant HRAS G12V compared to wild-type HRAS, with various effector proteins . Here, HRAS G12V showed reduced interactions with Raf and enhanced binding with RalGDS. However, given that the non-hydrolysable GTP-analog GNP was used in the study, the difference is not due to impaired hydrolysis. Similarly with HRAS, KRAS G12X mutants exhibit reduced affinity to Raf compared to wild-type . The G12D, G12R, and G12V mutants display highly reduced affinity to Raf, while the affinity of G12A is only moderately reduced. Interestingly, the affinity of the G12C mutant is similar to that of wild-type.
To bind RAS, the effector proteins use a ubiquitin (UB)-like fold: a RAS-binding domain (RBD) or a RAS-association domain (RA) [11,12]. While KRAS has not been co-crystallized with any of its effector proteins, distinct effector proteins have been resolved in complex with HRAS: RalGDS (PDB ID: 4G0N) , Raf-1 (PDB ID: 1LFD) , PI3Kγ (PDB ID: 1HE8) , PLCε (PDB ID: 2CL5) , RASSF5 (PDB ID: 3DDC)  and AF-6 (PDB ID: 6AMB) . These effector proteins bind to HRAS on top of its switch regions: switch-I (residues 30–40) and switch-II (residues 58–72), and the binding conformation of HRAS is almost identical in all of the complexes (S1A Fig). Given this, and since the G12X mutation is far from the binding interface (S1B Fig), Smith and Ikura  proposed that the discrepancies in the effector protein binding profiles of the mutants are due to altered switch dynamics. Overall, switch-I displays highly dynamic characteristics manifested as two different states when GTP is bound to RAS, and the distribution between these states is altered in mutants [19–22]. Given that the switch regions in HRAS and KRAS are identical (S1C Fig), their expected binding mode to their effectors is alike. A model of KRAS in complex with A-Raf-RBD tethered to a lipid-bilayer nanodisc suggested by NMR data agrees with this binding mode . At the cellular level, the isoform specificity to effector proteins is primarily determined via membrane interactions , but the differences among RAS isoforms’ absolute effector protein binding affinities rise from allosteric effects . It was observed that even a single point mutation in RAS (Q61L) has long-range effects on dynamics and alters effector protein interactions .
Previous molecular dynamics (MD) simulation studies of KRAS at microsecond timescales have mainly focused on the dynamical differences between the three wild-type RAS isoforms (HRAS, KRAS, NRAS) , differences among selected KRAS and HRAS mutants [27,28], the role of the hypervariable region (HVR) , KRAS’s membrane association or orientation [30–32], and KRAS oligomerization on the membrane . The total simulation times of these studies were in the range of 1–8 μs, which is reasonable but likely not sufficient to unravel long-time dynamics associated with slow conformational changes. More importantly, there is a lack of comprehensive atomistic MD simulations of all KRAS G12X mutants with extensive simulation times, allowing a reliable analysis for the differences in structure and dynamical behavior between the wild-type and the mutants, especially in the effector protein binding interface.
What is the underlying cause for the broad range of different G12X mutations? How do these distinctly different mutations manifest themselves in the structure, dynamics, and function of KRAS? This knowledge is crucial to understand KRAS oncogenesis and to develop future therapies targeting mutant KRAS harboring tumors. Therefore, in the present study we first assessed to what extent G12X mutation frequencies are explained by mutation probability. Intriguingly, an outstanding mutational bias emerged from the data. We next employed state-of-the-art atomistic MD simulations (total simulation time 170 μs) to study the dynamical behavior of KRAS with its natural ligands (GDP, GTP) bound, both in the wild-type KRAS and with all existing oncogenic G12X mutations. The results provided compelling evidence that mutations alter the dynamics of KRAS, that the alteration is mutation specific, displays allosteric characteristics, and that the alteration is manifested especially in the effector protein-binding interface. Furthermore, our data suggest that the observed mutational bias and the oncogenic properties of the individual KRAS G12X mutants are caused, at least in part, by mutation-specific altered dynamics.
Dyskeratosis congenita (DC) is an inherited bone marrow failure syndrome known as the prototype of telomere diseases. In addition to the clinical triad (nail dystrophy, hyperpigmentation, and leukoplakia), very short telomeres (below the 1 st percentile) is a marker for the diagnosis of DC (Calado & Young, NEJM 2009). Telomere dysfunction was associated with DC after discovery of DKC1 mutations in patients presenting the X-linked form of the syndrome. Novel mutations in telomere biology genes (TERC, TERT, NOP10, NHP2, TINF2, TCAB1, CTC1, RTEL1, ACD, and PARN) have been described in patients with DC. Variations in all these genes can affect telomere protection and maintenance, leading to telomere shortening and development of telomeropathies. In this study, we mapped the TERT, TERC, DKC1, and TINF2 genes for mutations in 15 patients (median age = 10 years M/F = 11/4) with DC and very short telomeres, in order to classify these cases in a molecular level and determine the frequency of these mutations in our cohort. Survival for twelve patients who underwent allogeneic hematopoietic stem cell transplant (HSCT) was assessed and correlated with mutational status and telomere length. Diagnosis of DC was made according to the definition of Calado & Young (2009). Telomere length was measured in nucleated blood cells by flow-FISH and mutational screening was performed on genomic DNA extracted from peripheral blood cells by direct sequencing. Seven non-synonymous mutations were identified in TINF2 (40%), two in DKC1 (13%), one in TERT (6%), and one in TERC (6%). The TINF2 variants R282H and R282C had been already described as pathogenic, as well T66A and A353V DKC1 variants (Knight et al, 1999 Savage et al, 2008 Walne et al, 2008). The heterozygous variant R282H (c. 845 G>A) in TINF2 were found in 4 unrelated patients. One of them also harbor the variants Q120R and Q157H in the same gene. The heterozygous mutation in TINF2 R282C (c. 844 C>T) was found in one patient, that also presented the common polymorphism A279T in TERT. The pathogenic variants T66A (c.196A>G) and A353V (c.1058C>T) in DKC1 were found in two different male patients. Moreover, three novel mutations were identified in our cohort, r.94 C>T in TERC, F290C in TINF2, and R696Cin TERT. The heterozygous mutation r.94 (C>T) found in TERC was located at the pseudoknot P2b region of the gene and the patient who carries that presented a severe aplastic anemia and all DC clinical triad. The novel heterozygous F290C (c.859 T>G) variant is located at the "hot spot" in exon 6 of TINF2 andwas found in one patient that presented a severe phenotype of DC. In silico analysis with SIFT and Polyphen-2 predicted that this variant is not tolerated and probably damaging, which is consistent with the pathogenicity of the mutation. The homozygous mutation R696C (c.2086 C>T) in TERT was found in one patient and also in his two brothers. All of them presented reduced blood cell count, clinical features of DC, and severe aplastic anemia. The family screening identified the father and sister as heterozygous for the same mutation, but both asymptomatic. DNA sample from the mother were not available for this study. In silico analysis by SIFT and Polyphen 2.0, predicted that the R696C mutation is not tolerant and possible damaging to telomerase activity, respectively. To validate in silico analysis, TRAP assay with cell lysates obtained from telomerase-negative VA13 cell line transfected with wild type or R696C mutated TERT vector and TERC vector is under evaluation. Consistently with previously studies, telomere length in patients with TINF2 mutations were the shortest compared with the other telomeres genes mapped in this study. Although, the phenotype and severity of the disease does not appear to change according to the mutated gene. Also, the mutational status (p=0.28) or telomere length (p=0.21) did not influence the survival rates of patients after HSCT.
Flow-FISH was able to identify patients with very short telomeres and validated telomere length measurement as a diagnostic tool for DC. Direct sequencing of the most commonly mutated genes in DC in a cohort of patients with telomeres below 1 st percentile was able to characterize the genetic cause of this disease in more than 70% of the cases. The identification of genetic defect in DC can manage clinical decisions and is essential to genetic counseling prior to bone marrow transplantation.
Telomeres form the ends of chromosomes and help maintain genomic structural integrity [Moon and Jarstfer, 2007 ]. They consist of tandem hexameric (TTAGGG)n nucleotide repeats with a single-stranded overhang and protein complex. The overhang folds back to form a t-loop [Griffith et al., 1999 ], which prevents the telomere ends from being recognized as break points by the DNA damage repair machinery [Palm and de Lange, 2008 ]. Many proteins bind to or interact with the telomere to maintain telomeric integrity. Shelterin is an ordered associated protein complex that consists of TERF1, TERF2, TINF2, TERF2IP, ACD, and POT1. This complex helps form the t-loop, and protects the telomeres from degradation and inappropriate DNA repair, thereby avoiding end-to-end fusion, atypical recombination, and premature senescence [Palm and de Lange, 2008 ]. The telomerase reverse transcriptase (TERT) and its telomere template-containing RNA component (TERC) are telomere-associated proteins that add telomeric repeats to elongate telomeres [Collins and Mitchell, 2002 ]. Telomerase activity is usually absent in differentiated cells. There are numerous other important telomere-associated proteins and complexes that transiently associates with telomeric DNA, including proteins involved in DNA repair (e.g., ATM and MRE11A) and helicases (e.g., BLM and RECQL) [Aubert and Lansdorp, 2008 ]. A large number of additional proteins interact either directly or indirectly at telomeric ends, and regulate protein–protein and protein–DNA interactions, cellular protein trafficking, and additional telomere-specific functions.
Telomeric repeats range in size from 0.15 to 50 kilobases (kb), and progressively shorten with each cell division [Aubert and Lansdorp, 2008 ]. Telomere length is dependent on many factors including age, replicative history of the cell, chromosome arm, and tissue type [Aubert and Lansdorp, 2008 Wise et al., 2009 ]. There is considerable interindividual variation in telomere length [Aviv et al., 2009 Nordfjäll et al., 2009 ], and a genetic influence on length variation. Twin and family studies have estimated the heritability of mean leukocyte telomere length to range from 44 to 84% [Jeanclos et al., 2000 Njajou et al., 2007 Slagboom et al., 1994 Vasa-Nicotera et al., 2004 ]. Quantitative-trait linkage analyses have mapped loci influencing telomere length to chromosomes 12q12.22 and 14q23.2 [Andrew et al., 2006 Vasa-Nicotera et al., 2004 ]. Fine mapping of the 12q12.22 locus identified a polymorphism in the BICD1 gene that was significantly associated with shorter telomere length [Mangino et al., 2008 ]. A genome-wide association study (GWAS) identified two single nucleotide polymoerphisms (SNPs) on chromosome 18q12.2 associated with leukocyte telomere length in the region of the gene VPS34/PIKC3C [Mangino et al., 2009 ], which has been suggested to be involved in controlling telomere length variation in yeast [Rog et al., 2005 ]. Other recent GWASs have identified associations between leukocyte telomere length and the telomere-associated protein TERC [Codd et al., 2010 Levy et al., 2010 ] and a gene suggested to be involved with telomere length regulation, OBFC1 (oligonucleotide/oligosaccharide-binding folds containing one) [Levy et al., 2010 ].
The inverse relationships between telomere length and aging, and its role in age-related and premature aging diseases have been well documented [Aubert and Lansdorp, 2008 Garcia et al., 2007 ]. Telomere attrition has also been associated with inflammatory processes, oxidative stress, and an unhealthy lifestyle [Mirabello et al., 2009 Morlá et al., 2006 von Zglinicki, 2002 ]. There is growing evidence that short telomeres are associated with the initiation and progression of cancer [Blasco et al., 1997 Hackett and Greider, 2002 ].
Cancer GWAS have shown that SNPs in genes encoding telomere-associated proteins at 5p15.33 (TERT-CLPTM1L locus) and RTEL1 were associated with risk of glioma [Shete et al., 2009 Wrensch et al., 2009 ], pancreatic [Petersen et al., 2010 ], and/or lung cancer [Jin et al., 2009 Landi et al., 2009 McKay et al., 2008 ]. In addition, an association study of multiple tumor types suggests that this TERT-CLPTM1L region may contain important markers of overall cancer risk [Rafnar et al., 2009 ]. It is possible that these cancer-associated sequence variants in telomere-associated genes may be associated with shorter telomeres.
Studies of dyskeratosis congenita, an inherited bone marrow failure and cancer predisposition syndrome have also been important in understanding the consequences of telomere dysfunction [Savage and Alter, 2009 ]. Patients with dyskeratosis congenita have extremely short telomeres for their age, very high risk of several cancers, and germline mutations in genes important in the maintenance of telomeres (DKC1, TERC, TERT, NOLA3, TINF2, or NOLA2) [Armanios, 2009 Savage and Alter, 2009 ]. The phenotypic spectrum of telomere biology disorders also includes patients with isolated aplastic anemia [Yamaguchi et al., 2005 ], acute myelogenous leukemia [Calado et al., 2009 ], and idiopathic pulmonary fibrosis [Armanios et al., 2007 ] who may have mutations in TERC or TERT.
The majority of the associated and telomeric complex proteins are highly evolutionarily conserved [de Lange, 2004 Kanoh and Ishikawa, 2003 Li et al., 2000 Mirabello et al., 2008 Nakamura and Cech, 1998 Savage et al., 2005 ]. A recent population genetics study targeting 37 telomere maintenance genes in 53 worldwide populations found that these genes have limited genetic variation [Mirabello et al., 2008 ]. The majority of telomere genes had low diversity, high ancestral allele frequencies, and low population differentiation [Mirabello et al., 2008 ].
There is little known about how common genetic variation relates to telomere length. We hypothesize that common genetic variation in genes encoding telomere-associated proteins could affect telomere length. We evaluated the association between genetic variation in 43 candidate telomere biology genes and leukocyte telomere length using SNP markers from GWAS of breast and prostate cancers [Hunter et al., 2007 Yeager et al., 2007 ]. These genes encode proteins that are thought to be involved either in telomere length maintenance or with telomere binding proteins necessary for telomere stability and structure. Telomere length data was obtained from prospectively collected blood samples and measured using quantitative-polymerase chain reaction (Q-PCR) by the same laboratory [De Vivo et al., 2009 Mirabello et al., 2009 ].
Telomeres and age-related disease: how telomere biology informs clinical paradigms
Department of Oncology, Sidney Kimmel Comprehensive Cancer Center, and McKusick-Nathans Institute of Genetic Medicine, Johns Hopkins University School of Medicine, Baltimore, Maryland, USA.
Address correspondence to: Mary Armanios, Department of Oncology, Johns Hopkins University School of Medicine, 1650 Orleans St., Cancer Research Building I Room 186, Baltimore, Maryland 21287, USA. Phone: 410.502.3817 Fax: 410.955.0125 E-mail: [email protected]
Find articles by Armanios, M. in: JCI | PubMed | Google Scholar
Telomere length shortens with age and predicts the onset of replicative senescence. Recently, short telomeres have been linked to the etiology of degenerative diseases such as idiopathic pulmonary fibrosis, bone marrow failure, and cryptogenic liver cirrhosis. These disorders have recognizable clinical manifestations, and the telomere defect explains their genetics and informs the approach to their treatment. Here, I review how telomere biology has become intimately connected to clinical paradigms both for understanding pathophysiology and for individualizing therapy decisions. I also critically examine nuances of interpreting telomere length measurement in clinical studies.
Understanding basic biological mechanisms holds the potential to advance clinical paradigms. The emerging impact of telomerase and telomere biology in medicine provides a clear example of this promise. Research in this area was initially sparked by fundamental questions about how genomes are protected at chromosome ends, and focused on curiosity-driven questions in maize, yeast, and protozoa ( 1 ). These highly conserved molecular mechanisms have now led to unforeseen benefits for understanding idiopathic disease and have opened a new area of translational research. Here I review the trajectory of the evolving role of telomere biology in clinical paradigms and highlight how it has become central to understanding the pathophysiology of age-dependent disorders as well as for informing new approaches to their treatment.
Telomeres define the ends of linear chromosomes. They are made up of repetitive DNA sequences that are bound by specialized proteins. The human telomeric DNA sequence is a tandem repeat of TTAGGG that extends several kilobases (a mean of 10 kilobases in umbilical cord blood) ( 2 – 4 ). The telomere-binding complex of proteins, known as shelterin, together with telomere DNA, functions as a dynamic unit that protects chromosome ends from being recognized as broken DNA, thus preventing their degradation and participation in fusion events ( 5 ). Telomeres are therefore essential for the maintenance of genomic integrity.
Telomerase is the specialized polymerase that synthesizes new telomere repeats ( 6 , 7 ). It offsets the shortening that normally occurs with cell division since the replication machinery does not copy fully to the ends. Telomerase has two essential core components, the telomerase reverse transcriptase (TERT) and the telomerase RNA (TR), the latter of which provides the template for telomere repeat addition ( 8 – 10 ). In human cells, telomerase is the primary mechanism by which telomeric DNA is synthesized de novo. As will be discussed, mutations in the TERT and TR genes are considered the most common cause of inherited human telomere-mediated disease ( 11 ). Even with mild perturbations in telomerase activity, telomere length homeostasis is disturbed and manifests in what has become recognized as a discrete syndrome complex, which recapitulates age-dependent disease processes ( 12 , 13 ). As such, these mutations and their clinical consequences are the primary focus of this Review.
Telomeres have long been linked to processes of cellular aging. Since the 1990s it has been known that telomere length predicts the onset of replicative senescence ( 14 , 15 ), a permanent state of cell cycle arrest that primary cells reach after they undergo a finite number of cell divisions ( 16 ). The fact that telomeres also shorten in vivo in humans with advancing age made a further compelling case for the idea that telomeres play a role in age-related processes. The evidence reviewed here shows that telomere shortening is sufficient to provoke age-related pathology. Several factors ensure that telomere shortening is a default state in somatic cells. Although telomerase offsets the end-replication problem, its levels are tightly regulated and only a few telomeres are elongated in each cell cycle ( 17 ). Therefore, even cells that may be relatively enriched for telomerase activity, such as hematopoietic stem cells, undergo telomere shortening with age ( 4 ). The incremental elongation of telomeres by telomerase can also be seen across generations ( 18 ). For both humans and mice, the telomere length of parentes determines the telomere length of their offspring ( 19 – 21 ). These observations have further established telomere length as a unique genotype (at times referred to as “the telotype”) and as a source of genetic variation across human populations ( 22 ).
When telomeres become critically short, they become dysfunctional and activate a DNA damage response that resembles double-strand breaks ( 23 ). The resulting signaling cascade provokes apoptosis and/or a permanent cell cycle arrest that, until recently, has been considered the primary functional consequence of senescence. Cell type–dependent factors determine whether apoptosis, senescence, or a combined phenotype predominates in response to dysfunctional telomeres ( 14 , 24 , 25 ), although the molecular effectors that discriminate between these pathways are not entirely understood.
Recently, a more complex understanding of the senescence phenotype has been emerging and suggests a closer link to disease mechanisms than was previously appreciated. For example, although senescent cells are quiescent in the cell cycle, for reasons that are not entirely clear, their gene expression profile is altered ( 26 ). One consequence of this altered gene expression is that senescent cells secrete a predictable profile of cytokines, chemokines, and proteases into culture media, a phenotype known as the senescence-associated secretory phenotype (SASP) ( 27 , 28 ). In vivo, the SASP has been hypothesized to play a role in the clearance of damaged cells ( 29 ). Telomere dysfunction is furthermore associated with a state of decreased cellular metabolic activity ( 25 , 30 ). In mice with short telomeres, defective cellular metabolism in the setting of senescence manifests as mitochondrial dysfunction and aberrant Ca 2+ signaling that cause insulin secretory defects by pancreatic β cells ( 25 ). These defects disturb glucose homeostasis in vivo. The fact that cellular senescence is associated with defective signaling and metabolism provides new contexts for understanding mechanisms of degenerative disease with age, particularly because these defects might occur in the absence of overt histopathology ( 25 ).
The most compelling evidence that telomeres contribute to aging comes from the fact that mutant telomerase and telomere genes cause telomere shortening that manifests in age-related phenotypes (see Telomere syndrome manifestations that overlap with human age-related phenotypes). Because telomere shortening is acquired universally with age, these disorders have a particular relevance for understanding mechanisms of age-related disease. Telomere-mediated disorders show two hallmarks of age-related disease: degenerative organ failure and a cancer-prone state ( 31 ). Age-related disease is additionally marked by atherosclerosis however, premature vascular disease has not been reported and does not, in our experience, seem to be accelerated in individuals with telomere disorders.
Eight genes have been implicated in monogenic telomere disorders (reviewed in ref. 22 ). The most prevalent are heterozygous mutations in TERT and TR, which cause autosomal dominant disease. The dominant mode of inheritance occurs as a result of the sensitivity of telomere maintenance to telomerase levels, even when only one allele is perturbed ( 12 , 18 , 32 – 35 ). Mutations in TERT and TR usually cause significant morbidity after the reproductive age is reached, and a greater number of offspring are affected as a result of their dominant mode of inheritance. They are thus estimated to be the most prevalent cause of inherited telomere disorders, comprising at least 90% of cases ( 11 ). Mutations in genes encoding the X-linked telomerase accessory component, DKC1, which is essential for human TR stability, and the autosomal shelterin gene, TINF2, explain a significant subset of pediatric telomere syndrome cases, especially in the setting of dyskeratosis congenita, which was the first genetic disorder to be linked to telomere biology ( 36 – 38 ). Biallelic mutations in the conserved telomere component 1 gene, CTC1, which plays a putative role in telomere lagging strand synthesis, have also been recently implicated in rare autosomal recessive cases that also have predominantly pediatric presentations ( 39 – 41 ). There remains a subset of cases with inherited telomere phenotypes for which the mutant genes are unknown their identification is the focus of ongoing research.
Telomere-mediated disease has diverse presentations that span the age spectrum. Their type, age of onset, and severity depend on the extent of the telomere length defect. In infancy, severe telomere shortening manifests as developmental delay, cerebellar hypoplasia, and immunodeficiency, features that are recognized in the rare Hoyeraal-Hreidarsson syndrome ( 42 ). In children and young adults, telomere-mediated disease causes bone marrow failure and at times may be recognized in the mucocutaneous syndrome dyskeratosis congenita, which is defined by a triad of mucocutaneous features — skin hyperpigmentation, dystrophic nails, and oral leukoplakia ( 33 , 43 , 44 ). Telomere-mediated disease manifests in adults as isolated or syndromic clustering of idiopathic pulmonary fibrosis (IPF), liver cirrhosis, and bone marrow failure ( 31 ). Mutant TERT and TR genes account for 8%–15% of familial and 1%–3% of sporadic pulmonary fibrosis cases ( 45 – 47 ). Because IPF affects at least 100,000 individuals in the United States, it is considered the most prevalent manifestation of the telomere disorders ( 11 ). An individual who carries a telomerase mutation will therefore most frequently be clinically recognized as an adult with familial pulmonary fibrosis. Adult-onset telomere disease may rarely also manifest as sporadic or familial myelodysplastic syndrome or acute myeloid leukemia ( 48 – 50 ). The co-occurrence of IPF and bone marrow failure within a single family is highly predictive for the presence of a germline telomerase defect ( 51 ).
Although the manifestations of telomere-mediated disease occur in multiple organs and may appear clinically different, it has been proposed that their shared short telomere length defect unifies them under the umbrella of a single syndrome continuum ( 12 , 22 , 31 , 45 , 46 , 51 ). This molecular classification is significant because the telomere defect is present in the germline of these patients and thus, even when a single presentation predominates, complications that are relevant to managing symptoms and averting complications may arise in other organs. The regrouping of what have historically been considered unrelated disorders provides new clinical insights as these conditions significantly overlap. The consideration of the telomere syndromes as a single spectrum exemplifies how a molecular classification of disease may help explain previously mysterious complications of treatment and refine clinical approaches.
The clinical manifestations of telomere shortening can be divided into two broad categories: those affecting high-turnover tissues and those affecting low-turnover tissues. This distinction is important for understanding disease patterns because the high-turnover phenotypes tend to appear first in pediatric populations and represent more severe disease (ref. 51 and Figure 1). For example, telomere syndromes in infancy manifest as severe immunodeficiency, which affects B cells, T cells, and NK cells, coincident with the extraordinary replicative demands on the adaptive immune system during this period of development ( 42 , 52 , 53 ). Bone marrow phenotypes tend to appear later in children and young adults as isolated cytopenias or aplastic anemia ( 43 , 45 , 51 , 54 ). The hematopoietic defects have been studied in animal models and represent a stem cell failure state whereby short telomere length limits both stem cell number and function ( 33 , 51 , 55 – 57 ). The telomere-mediated bone marrow failure phenotype is stem cell autonomous because allogeneic stem cell transplantation can reverse this state. The gastrointestinal epithelium, another high-turnover compartment, is also affected in a subset of patients who develop an enteropathy marked by villous blunting that resembles celiac disease ( 53 ). These intestinal phenotypes are similarly thought to be caused by stem cell failure that appears as villous atrophy in mice with short telomeres ( 18 , 58 ).
Clinical manifestations of telomere disorders and their onset relative to tissue turnover rate. Shown are representative images of diagnostic histopathology and radiographic studies in patients with telomere-mediated disease (A–D) and 5-ethynyl-2′-deoxyuridine (EdU) incorporation detected in corresponding mouse tissues (E–H). The estimated turnover rate of more than 90% of cells is indicated for each pair of images. (A) Photomicrograph of a bone marrow biopsy showing an acellular marrow replaced by adipose tissue with only remnants of hematopoiesis, taken from an individual with aplastic anemia. Image reproduced with permission from Annual Reviews of Genomics and Human Genetics ( 31 ). (B) Histopathology of a duodenal biopsy from a patient with telomere-mediated enteropathy shows profound villous atrophy. Image reproduced with permission from Aging Cell ( 53 ). (C) Abdominal CT scan image from a patient with liver cirrhosis, as evidenced by the nodular liver surface, the caudate lobe hypertrophy, and splenomegaly. (D) Lung windows of a chest CT scan from a carrier of the telomerase mutation show classic basilar honeycombing changes pathognomonic for IPF. (E) Flow cytometry plot of EdU incorporation in the bone marrow after a short (2-hour) pulse, showing that nearly one-third of the cells have undergone division. (F) Immunohistochemistry of intestinal section after a EdU pulse (5 days) shows that nearly all enteric epithelial cells are positively labeled (brown). (G) Brown staining shows EdU-labeled hepatocytes after EdU labeling (14 days). (H) Image of terminal bronchiole shows EdU-positive lung epithelial cells (red) identified by the Clara cell antigen (green) after 14 day label.
More commonly, telomere-mediated disease manifests in slow-turnover tissues, such as the lung and the liver (Figure 1). These phenotypes frequently appear as de novo adult-onset disease, in contrast to the pediatric presentations of dyskeratosis congenita and related disorders. IPF presents at a mean age between 50 and 60 years (range 31–87) ( 35 , 45 , 47 , 51 , 59 , 60 ), and telomere-related cryptogenic liver fibrosis, based on reported cases, presents at a mean of 37 years (range 20–57) ( 12 , 59 , 61 ). The mechanisms of these adult-onset disorders can also be distinguished in animal models. In contrast to the high-turnover phenotypes that are readily evident in the telomerase knockout mouse, telomere dysfunction in slow-turnover organs serves as the first of multiple acquired “hits” that contribute ultimately to organ failure (Figure 2). For example, mice with short telomeres do not develop de novo lung phenotypes, but acquire them only after chronic injury such as with cigarette smoke ( 62 ). Similarly, liver damage is only detected when mice with short telomeres are challenged with carbon tetrachloride ( 63 ). In the endocrine pancreas, telomere dysfunction cooperates with genetically induced endoplasmic reticulum stress to cause β cell apoptosis and manifest in worsening diabetes severity ( 25 ). Therefore, in tissues in which adult cell turnover is minimal, telomere dysfunction disturbs organ homeostasis because of cumulative hits in long-lived cells and eventually culminates in what appears as irreversible adult-onset disease (Figure 2). The cell types responsible for the telomere-induced fibrotic disorders are not known, but it has been hypothesized that these disorders, similar to the telomere-dependent bone marrow and intestinal defects, represent stem cell failure states ( 45 ). This framework has important implications for treatment strategies, as discussed below.
Model for understanding the mechanisms of telomere-mediated disease in high- and low-turnover tissues. In high-turnover tissues (left), cell replication is the primary determinant of disease onset. In contrast, in low-turnover tissues (right), other genetic and acquired hits contribute to disease onset. In both cases, telomere dysfunction induces apoptosis and/or senescence. The senescence phenotype may be associated with gene expression changes, mitochondrial dysfunction, aberrant Ca 2+ signaling, and the SASP.
Telomere length is the primary determinant of disease onset and predominant presentation in telomere disorders. This observation is supported by the fact that in families that carry mutant telomerase genes and display autosomal dominant inheritance, the disease worsens and appears earlier with each successive generation as the telomere length shortens ( 12 , 64 ). Genetic anticipation due to telomere shortening was first recognized in telomerase-null mice, which develop worsening phenotypes with successive breeding ( 18 , 58 , 65 ). In very late generations, mice die at pre-reproductive ages, which eventually limits the genetic lineage ( 19 ). The severity of the genetic anticipation in human families correlates in part with the extent of telomerase loss of function — families with functionally null telomerase alleles show more evident changes in onset across consecutive generations, in contrast to families that carry hypomorphic mutations ( 12 , 35 ). Telomere phenotypes also evolve in autosomal dominant telomere syndromes. In older generations, slow-turnover disease tends to predominate, with IPF being the primary first complication. In later generations, a bone marrow failure–predominant phenotype often comes to attention first ( 51 ). Therefore, a single telomerase gene mutation can have heterogeneous manifestations within a given family ( 51 ). This evolving pattern is unique to these Mendelian disorders and distinguishes the telomere syndromes from other conditions that show genetic anticipation, such as the trinucleotide repeat expansion syndromes ( 66 ). Clinically, this pattern of inheritance poses particular challenges to genetic counseling discussions with at-risk individuals, as the type and onset of disease may be heterogeneous and difficult to predict.
Telomere-mediated organ failure typically has a protracted course, especially in adults who may have subclinical disease for many years before becoming symptomatic ( 67 ). In some cases, for example with an offending insult such as an infection or an exposure to drug toxicities, acute declines can be sustained. In the past, this progressive course has led to a view of telomere disorders (such as aplastic anemia and IPF) as autoimmune processes, and to their empiric treatment with immunosuppression ( 33 ). With clear causal links to telomere defects, and with a growing appreciation for the full spectrum of telomere phenotypes, it is now possible to identify affected patients and thus to refine the treatment approach. Patients with telomere-related syndromes are known to have a higher incidence of adverse events with cytotoxic therapies ( 44 ) which makes the diagnostic considerations particularly important. The current treatment for telomere-mediated organ failure is primarily supportive, and its complete reversal is feasible only with organ transplantation. Below I highlight some examples in which telomere biology has affected clinical paradigms.
Patients with telomere syndromes may have subtle cosmetic features of aging (e.g., premature hair graying), but dysmorphic features are not sensitive and, in our experience, not sufficiently robust to make the diagnosis, even with training. In the setting of bone marrow transplantation, such diagnostic decisions are particularly imperative because patients with telomere syndrome have historically had poor outcomes with conventional bone marrow transplantation (reviewed in ref. 44 ). Morbidity and mortality occur primarily because of pulmonary and liver toxicity related to chemotherapy used in standard conditioning transplant regimens. With appreciation for the broad telomere-related clinical spectrum, and with the availability of DNA sequencing and telomere length measurement, improved selection has allowed for the testing of reduced-intensity regimens in dedicated studies for patients with telomere disorders. This approach has shown promising short-term outcomes ( 68 ).
IPF treatment is another evolving area in which telomere biology challenges current treatment approaches. IPF is a progressive disorder with a mean survival of 3 years from diagnosis ( 69 ). No approved treatments for IPF are currently available, and lung transplantation is accessible to only a small subset of patients who develop end-stage lung disease (less than 5%) ( 70 , 71 ). Although telomerase mutations are the most commonly identifiable genetic cause for familial pulmonary fibrosis, short telomere length in pathological ranges is a common feature even in IPF patients without mutations ( 11 ). The telomere length defect is likely in the germline, as it is concurrently seen in multiple leukocyte subsets as well as lung epithelial cells ( 46 ). This observation has led to the idea that short telomere length may be a risk factor for this disease ( 46 ). In support of the idea that telomere length might play a role in driving apparently sporadic IPF is the observation that a subset of IPF patients concomitantly develops cryptogenic liver cirrhosis, another telomere-mediated phenotype ( 46 ). In the past two decades, the idea that IPF may be an immune-mediated disease has led to the use of immunosuppressive therapy outside and within clinical trials (ref. 72 and references therein). A recent phase III trial that randomized patients with IPF to the immunosuppressive regimen of N-acetyl cysteine alone, a combination of N-acetyl cysteine, prednisone, and azathioprine, or placebo alone was stopped early because the mortality rate in the group receiving combination treatment was 8-fold higher than that in the placebo group ( 73 ). The majority of deaths were reported as respiratory in nature, but it remains unclear whether they were indirectly related to systemic toxicity. Although the role of telomere defects in sporadic forms of IPF is not yet fully understood, the lack of efficacy combined with the increased toxicity seen in recent immunosuppression trials suggests that future clinical approaches to IPF treatment should account for the fact that patients with this form of idiopathic interstitial pneumonia may be exquisitely sensitive to cytotoxic drugs. IPF patients also fare poorly with cancer treatment, an observation that is not commonly noted in patients with other lung disorders ( 74 ). It has been suggested that the apparently irreversible scarring pattern of IPF may represent a stem cell failure state that will not amenable to reversal with immunosuppression similar to telomere-mediated aplastic anemia ( 45 ). Ultimately, fundamental research in lung biology following the telomere genetic clues has the potential to open paths to new treatment paradigms for age-dependent fibrotic lung disease.
One important breakthrough that has emerged from the study of human monogenic disorders is the delineation of clinically meaningful thresholds for telomere shortening. Through the use of the telomere length method of flow cytometry and fluorescence in situ hybridization ( 75 ), the lymphocyte telomere length in patients with telomere syndrome can be stratified relative to age-matched controls in the population. Early studies that have examined this tool in the monogenic telomere disorders suggest that a threshold below the tenth percentile is sensitive, and below the first percentile is fairly specific, for distinguishing individuals who carry mutant telomere genes from their relatives who are noncarriers ( 45 , 46 , 76 ). These ranges have allowed for the use of this validated method for testing telomere length in the diagnostic work-up of suspected telomere disorders.
The fact that certain age-adjusted thresholds of telomere length have predictive value in clinical settings is significant because short leukocyte telomere length has been associated with numerous disease states and environmental factors, including chronic inflammatory states such as cancer (reviewed in ref. 77 ), cardiovascular disease (reviewed in ref. 20 ), and acquired states such as emotional stress, poor socioeconomic status, and education levels (reviewed in refs. 78 , 79 ). Although some of these variables have shown statistically significant telomere shortening consistently across studies, the biological consequences of this relative shortening cannot be equated with the severe telomere length defects seen in the monogenic telomere disorders (Figure 3). While the differences may be statistically significant, the absolute telomere length change in some cases may be small and might therefore reflect acquired replicative stress states rather than telomere-driven degenerative changes such as with the monogenic telomere syndromes. This important caveat should be considered in the interpretation of telomere epidemiology studies.
Telomere syndromes have defined pathological ranges of telomere shortening. Although short telomere length (TL) has been associated with numerous conditions, in some cases, the shortening reflects acquired replicative stress states rather than telomere-driven degenerative changes. (A) Putative dataset showing large effect size and short telomere length outside of the normal age-adjusted range. (B) Small and statistically significant change in telomere length in hypothetical dataset is less likely to reflect a telomere-mediated process.
Clinical observations in patients with telomere syndromes also shed light on the role of telomeres in cancer, which until recently had been primarily studied in cell culture and animal models. Like other DNA repair disorders, telomere disorders are cancer prone however, the overall incidence is relatively low ( 80 ). The cancer-related mortality in patients with telomere syndrome is not known, but it has been estimated that 10% of patients with dyskeratosis congenita are diagnosed with cancer ( 54 , 80 ). However, that estimate likely includes skin squamous cell cancers, which are prevalent in this group of patients and are usually not lethal ( 54 ). Cancers in dyskeratosis congenita have a predilection for high-turnover tissues, with squamous cell carcinomas of the skin and upper aerodigestive tract, myelodysplasia, and acute myeloid leukemia being the most common ( 80 ). In a cohort of adults with IPF with TERT mutations, 10% self-reported a history of cancer, although this rate was not adjusted for age or other exposures ( 60 ). These clinical observations make it clear that although telomere syndrome patients are at significantly increased risk for developing cancer, degenerative disease accounts for the majority of the morbidity and mortality in at least 90% of cases.
The relatively low overall incidence of cancers in patients with telomere disorders underscores the fact that in the presence of an intact DNA damage response, short telomere length predominantly causes cell loss in humans. These observations are in line with the long-hypothesized role of telomere shortening as a powerful tumor-suppressive mechanism ( 81 ). Studies in animal models have shown that short telomeres suppress tumorigenesis by mediating p53-dependent apoptosis and senescence ( 82 ). In mice with short telomeres that also lack p53, genomic instability fuels carcinogenesis ( 83 – 85 ). Whether short telomere length in human cancers may contribute to genomic instability at a low level remains a question of ongoing study. Other explanations have been hypothesized to underlie the tumor-prone nature of telomere syndromes, such as compromised immunosurveillance due to the associated immunodeficiency phenotype ( 22 ). The stem cell exhaustion state itself has also been proposed to contribute to tumorigenesis, and this would explain the tumor-prone nature of stem cell failure states, such as occurs with non–telomere-mediated aplastic anemia. The clinical study of disease driven by telomere defects provides a unique opportunity to refine current ideas about the role of telomere dysfunction in human cancer development and progression.
Hypothesized molecular mechanisms for aging in modern biology have abounded. These have included stem cell failure, mitochondrial dysfunction, genotoxic stress, and epigenetic changes. Recent cumulative evidence points to telomere shortening as sufficient to provoke all these mechanisms. The manifestations of telomere-mediated disease, especially in adults, can be subtle and are often indistinguishable from the slow, gradual functional decline that is a hallmark of aging. The compelling clinical evidence therefore points to telomere shortening itself as being sufficient, or perhaps more broadly representing forms of genotoxic stress that contribute to age-related changes.
In the past decade, telomere biology has provided a molecular rationale for unifying a group of historically considered unrelated disorders under the umbrella of telomere syndromes. The rich, context-dependent clinical presentations of these single-gene disorders and their now appreciated overlap highlight how a molecularly based understanding of disease can refine clinical care at the bedside. This new understanding underscores how the interpretation of increasingly available genetic information might require clinical contextualization before it can be readily applied. Beyond these conceptual considerations, telomere biology has of late brought new tools for diagnosis as well as for understanding disease mechanisms in areas that have long been perplexing to clinicians. Such novel paradigms are particularly needed when it comes to approaching difficult problems such as IPF. The coming years will undoubtedly point to new examples of how the biology of these DNA ends may advance clinical care.
I am particularly indebted to Jonathan Alder for helpful discussions and for assistance with the figures, and to Carol Greider for critical comments on the manuscript. I acknowledge funding support from the NIH (grants R21 HL104345 and RO1 CA160433), the Maryland Stem Cell and Commonwealth Foundations, and the Flight Attendants Medical Research Institute.
Conflict of interest: The author has declared that no conflict of interest exists.
Reference information: J Clin Invest. 2013123(3):996–1002. doi:10.1172/JCI66370.
The immortal DNA strand hypothesis, originally proposed by Cairns in 1975, poses that adult mammalian stem cells do not segregate DNA strands randomly after proliferation . Instead, stem cells might preferentially retain the parental ancestral strand, whereas the duplicated strand is passed onto differentiated cells with limited life span (Fig 1). In principle, such hierarchical tissues could produce differentiated progeny indefinitely without accumulating any proliferation-induced mutations in the stem cell compartment [2,3]. Experimental evidence supporting this hypothesis comes from BrdU stain tracing experiments both in vitro and in vivo [4–7]. Evidence from spindle orientation bias in mouse models of normal and precancerous intestinal tissue corroborated these findings, suggesting that strand segregation is then lost during tumourigenesis . However, many of the experiments suffer from uncertainties in stem cell identity and a definite mechanism of strand recognition remains unknown . Hence why Cairns hypothesis remains controversial .
a) During replication of the ancestral DNA strand, errors (dashed line) might occur. If these errors are not corrected by intrinsic DNA repair mechanisms, they become permanently fixed in daughter cells after the next cell division. However, the original ancestral strand is still present and can provide the blue print for additional non-mutated copies of DNA. b) In principle, a stem cell driven tissue allows for non-random DNA strand segregation. Preferentially segregating ancestral DNA strands into stem cells and duplicated strands into differentiated cells with limited life span can drastically reduce the accumulation of somatic mutations in tissues.
Orthogonal studies based on the expected accumulation of somatic mutations in healthy human tissues have argued against the immortal strand hypothesis [11,12]. However, the mere accumulation of somatic mutations in healthy tissue neither supports nor negates the immortal strand hypothesis in vivo. Here, we show that measuring the change of the mutational burden and, most crucially, the change of the variance of the mutational burden with age allows determining the probability of DNA strand segregation and the per cell mutation rate in healthy human tissues. First, we outline the approach and then apply it to genomic data from healthy human colon, small intestine, liver, skin and brain tissue. The data comes from four recent independent studies on mutational burden in healthy tissues [13–16], which contain information on in total 39 individuals at different ages and analysed genomes of 341 single cells. We find evidence for non-random strand segregation in all adult tissues and significant differences in somatic mutation rates between tissues, but less prominent strand-segregation in brain tissue during early development.
One contribution of 15 to a Discussion Meeting Issue ‘The new science of ageing’.
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