Human diseases of telomerase dysfunction: insights into tissue aging

There are at least three human diseases that are associated with germ-line mutations of the genes encoding the two essential components of telomerase, TERT and TERC. Heterozygous mutations of these genes have been described for patients with dyskeratosis congenita, bone marrow failure and idiopathic pulmonary fibrosis. In this review, we will detail the clinical similarities and difference of these diseases and review the molecular phenotypes observed. The spectrum of mutations in TERT and TERC varies for these diseases and may in part explain the clinical differences observed. Environmental insults and genetic modifiers that accelerate telomere shortening and increase cell turnover may exaggerate the effects of telomerase haploinsufficiency, contributing to the variability of age of onset as well as tissue-specific organ pathology. A central still unanswered question is whether telomerase dysfunction and short telomeres are a much more prominent factor than previously suspected in other adult-onset, age-related diseases. Understanding the biological effects of these mutations may ultimately lead to novel treatments for these patients

Facioscapulohumeral muscular dystrophy: Are telomeres the end of the story

Facioscapulohumeral muscular dystrophy (FSHD) is a progressive myopathy with a relatively late age of onset (usually in the late teens) compared with Duchenne and many other muscular dystrophies. The current FSHD disease model postulates that contraction of the D4Z4 array at chromosome 4q35 leads to a more open chromatin conformation in that region and allows transcription of the DUX4 gene. DUX4 mRNA is stable only when transcribed from certain haplotypes that contain a polyadenylation signal. DUX4 protein is hypothesized to cause FSHD by mediating cytotoxicity and impairing skeletal muscle differentiation. We recently showed in a cell culture model that DUX4 expression is regulated by telomere length, suggesting that telomere shortening during aging may be partially responsible for the delayed onset and progressive nature of FSHD. We here put our data in the context of other recent findings arguing that progressive telomere shortening may play a critical role in FSHD but is not the whole story and that the current disease model needs additional refinement

Decreasing initial telomere length in humans intergenerationally understates age-associated telomere shortening

Telomere length shortens with aging, and short telomeres have been linked to a wide variety of pathologies. Previous studies suggested a discrepancy in age-associated telomere shortening rate estimated by cross-sectional studies versus the rate measured in longitudinal studies, indicating a potential bias in cross-sectional estimates. Intergenerational changes in initial telomere length, such as that predicted by the previously described effect of a father's age at birth of his offspring (FAB), could explain the discrepancy in shortening rate measurements. We evaluated whether changes occur in initial telomere length over multiple generations in three large datasets and identified paternal birth year (PBY) as a variable that reconciles the difference between longitudinal and cross-sectional measurements. We also clarify the association between FAB and offspring telomere length, demonstrating that this effect is substantially larger than reported in the past. These results indicate the presence of a downward secular trend in telomere length at birth over generational time with potential public health implications

The Roles of Telomerase in the Generation of Polyploidy during Neoplastic Cell Growth

AbstractPolyploidy contributes to extensive intratumor genomic heterogeneity that characterizes advanced malignancies and is thought to limit the efficiency of current cancer therapies. It has been shown that telomere deprotection in p53-deficient mouse embryonic fibroblasts leads to high rates of polyploidization. We now show that tumor genome evolution through whole-genome duplication occurs in ∼15% of the karyotyped human neoplasms and correlates with disease progression. In a panel of human cancer and transformed cell lines representing the two known types of genomic instability (chromosomal and microsatellite), as well as the two known pathways of telomere maintenance in cancer (telomerase activity and alternative lengthening of telomeres), telomere dysfunction-driven polyploidization occurred independently of the mutational status of p53. Depending on the preexisting context of telomere maintenance, telomerase activity and its major components, human telomerase reverse transcriptase (hTERT) and human telomerase RNA component (hTERC), exert both reverse transcriptase-related (canonical) and noncanonical functions to affect tumor genome evolution through suppression or induction of polyploidization. These new findings provide a more complete mechanistic understanding of cancer progression that may, in the future, lead to novel therapeutic interventions

Establishment of clonal myogenic cell lines from severely affected dystrophic muscles - CDK4 maintains the myogenic population

<p>Abstract</p> <p>Background</p> <p>A hallmark of muscular dystrophies is the replacement of muscle by connective tissue. Muscle biopsies from patients severely affected with facioscapulohumeral muscular dystrophy (FSHD) may contain few myogenic cells. Because the chromosomal contraction at 4q35 linked to FSHD is thought to cause a defect within myogenic cells, it is important to study this particular cell type, rather than the fibroblasts and adipocytes of the endomysial fibrosis, to understand the mechanism leading to myopathy.</p> <p>Results</p> <p>We present a protocol to establish clonal myogenic cell lines from even severely dystrophic muscle that has been replaced mostly by fat, using overexpression of CDK4 and the catalytic component of telomerase (human telomerase reverse transcriptase; hTERT), and a subsequent cloning step. hTERT is necessary to compensate for telomere loss during <it>in vitro </it>cultivation, while CDK4 prevents a telomere-independent growth arrest affecting CD56+ myogenic cells, but not their CD56- counterpart, <it>in vitro</it>.</p> <p>Conclusions</p> <p>These immortal cell lines are valuable tools to reproducibly study the effect of the FSHD mutation within myoblasts isolated from muscles that have been severely affected by the disease, without the confounding influence of variable amounts of contaminating connective-tissue cells.</p

Sorbitol dehydrogenase overexpression and other aspects of dysregulated protein expression in human precancerous colorectal neoplasms: a quantitative proteomics study

Colorectal adenomas are cancer precursor lesions of the large bowel. A multitude of genomic and epigenomic changes have been documented in these preinvasive lesions, but their impact on the protein effectors of biological function has not been comprehensively explored. Using shotgun quantitative MS, we exhaustively investigated the proteome of 30 colorectal adenomas and paired samples of normal mucosa. Total protein extracts were prepared from these tissues (prospectively collected during colonoscopy) and from normal (HCEC) and cancerous (SW480, SW620, Caco2, HT29, CX1) colon epithelial cell lines. Peptides were labeled with isobaric tags (iTRAQ 8-plex), separated via OFFGEL electrophoresis, and analyzed by means of LC-MS/MS. Nonredundant protein families (4325 in tissues, 2017 in cell lines) were identified and quantified. Principal component analysis of the results clearly distinguished adenomas from normal mucosal samples and cancer cell lines from HCEC cells. Two hundred and twelve proteins displayed significant adenoma-related expression changes (q-value < 0.02, mean fold change versus normal mucosa ±1.4), which correlated (r = 0.74) with similar changes previously identified by our group at the transcriptome level. Fifty-one (∼25%) proteins displayed directionally similar expression changes in colorectal cancer cells (versus HCEC cells) and were therefore attributed to the epithelial component of adenomas. Although benign, adenomas already exhibited cancer-associated proteomic changes: 69 (91%) of the 76 protein up-regulations identified in these lesions have already been reported in cancers. One of the most striking changes involved sorbitol dehydrogenase, a key enzyme in the polyol pathway. Validation studies revealed dramatically increased sorbitol dehydrogenase concentrations and activity in adenomas and cancer cell lines, along with important changes in the expression of other enzymes in the same (AKR1B1) and related (KHK) pathways. Dysregulated polyol metabolism might represent a novel facet of metabolome remodeling associated with tumorigenesis

Human telomerase RNA and telomerase activity in immortal cell lines and tumor tissues

Telomerase activity has been detected in many human immortal cells lines and in tumor tissues, whereas it is generally absent from primary cell strains and from many tumor adjacent tissue samples. With the recently cloned human telomerase RNA (hTR), we used Northern analysis to follow the levels of hTR in primary, precrisis, and immortalized cells. It was surprising that the amount of hTR was high in cell strains that lacked telomerase activity, and the levels did not parallel the increases in telomerase activity, which accompanies immortalization. In addition, although the hTR levels were somewhat higher in tumor samples compared to nontumor tissues, the level of hTR in a variety of different human tumors did not predict the level of telomerase activity in the tumor. Thus, whereas hTR was detected in all samples that have telomerase activity, the presence of the RNA was not a good predictor of the presence or amount of telomerase activity