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Transcript
Werner Syndrome Marc Ialenti and Rushi Parikh Werner Syndrome (WS) is an autosomal recessive disease that leads to the premature manifestation of clinical symptoms associated with normal aging. Clinical symptoms include: short stature, graying/loss of hair, osteoporosis, cataracts, atherosclerosis, type II diabetes, hypogonadism, skin ulcers, reduced fertility, and high incidence of malignant neoplasms. They appear in one’s 20’s and 30’s, and myocardial infarction is the chief cause of death at approximately 48 of age. Individuals with WS display significant genomic instability in the form of DNA repair and replication defects, problems involving transcription and telomere maintenance, chromosomal rearrangements, attenuated apoptosis, and recombinational defects. The Werner protein (WRN) is involved in many crucial DNA metabolic pathways, particularly DNA repair processes, and so it serves as a key link between faulty DNA repair and the mechanisms of aging and cancer (1). WRN Biochemistry and Genetics WRN is a member of the RecQ family of DNA helicases and possesses ATPdependent 3’-5’ helicase and 3’-5’ exonuclease activity. The N-terminal region of WRN has exonuclease domains, the central region contains the RecQ helicase domains, and the C-terminal region possesses the nuclear localization signal (NLS) that controls the nuclear targeting of WRN. The preferred substrates of WRN helicase include DNA structures such as Holliday junctions, tetraplex DNA, forked duplexes, and mismatch “bubble” duplexes, as well as DNA-RNA heteroduplexes. WRN exonuclease targets DNA duplexes containing mismatch “bubbles”, a Holliday junction, or forks (2). The NLS directs WRN to localize at nucleoli (transcription centers), replication forks (DNA replication structures), DNA foci (DNA repair sites), and AA-PML bodies (telomere metabolism sites) (1). 35 WRN mutations have been identified to date with each resulting in absent or truncated protein products; it should be noted that WS is the only phenotype derived from WRN mutations. Mutations types include: insertions, deletions, and splicing donor/acceptor site mutations that result in frame shifts (no protein product), and premature stop codons that result in truncated proteins. These mutations predominantly occur in large introns and regulatory regions (2). The most prevalent mutation is 1336 C>T that accounts for nearly 25% of WS cases. The IVS 25-1G>C founder effect mutation accounts for 60% of WS cases amongst the Japanese population (3). All WRN mutations are characterized by the absence of the C-terminal region, and so the resulting loss of NLS and improper transport of WRN to specific nuclear regions seems key for WS pathogenesis (1). WRN Molecular Biology WRN is a critical participant in several DNA metabolic pathways—DNA repair, replication, and recombination as well as transcription. WRN not only unwinds/digests abnormal DNA structures during DNA metabolism, but it also helps regulate DNA repair and recombination through unwinding/digestion of intermediate DNA structures. Consequently, WRN clearly plays a key role in maintaining genomic integrity (1). WRN’s role in DNA replication is supported by the fact that individuals with WS have cells that undergo premature replicative senescence, exhibit longer S-phase, and show a reduction of replication initiation sites in comparison to cells of normal individuals (4). Recent studies have elucidated WRN’s interaction with DNA replication components DNA polymerase δ, Replication Protein A, and FEN-1, which is involved in the processing of Okazaki fragments during lagging strand synthesis (1). The integral role WRN plays in DNA repair is supported by WS cells having nonhomologous chromosomal rearrangements that are consistent with defective DNA repair mechanisms such as base excision repair and homologous recombinational repair (1). For instance, recent studies have shown WRN’s key association with the Ku heterodimer and DNA-dependent protein kinase, two major components involved in repairing double strand DNA breaks (5). WRN’s role in DNA damage response pathways (DNA repair) is further supported by its interaction with p53, a tumor suppressor protein that is critical for apoptosis, cell cycle arrest, and senescence. Recent studies indicate that the overexpression of WRN causes a increase in p53-dependent transcriptional activity and that p53-mediated apoptosis is attenuated by WS cells (6). As stated earlier, WRN is also involved in telomere metabolism. Telomere dysfunction and shortening leads to replicative senescence and genomic instability. Recent studies report that the expression of exogenous telomerase (telomere extender) in WS fibroblast lengthens the cellular life span. These findings suggest that premature senescence in WS cells is associated with telomeres, and more specifically, that WRN mutations lead to accelerated telomere shortening and disruption of telomere structure that the protective effects of telomerase may counteract against (7). Future WRN Considerations There are still many key features of WS that have yet to been explained. A better understanding of the regulation of WRN cellular localization is crucial for elucidating its cellular roles. Alterations such as post-translational modifications and specific protein interactions may prove pivotal in the nuclear targeting of WRN. Additionally, how does the lack of WRN helicase and exonuclease activities as well as the absence of WRN protein interactions lead to the premature aging symptoms that define WS? Tackling these and other issues regarding WS will lead to a better understanding of the aging process and cancer predisposition (1). WS Clinical Presentation and Diagnostic Testing The age of onset of many of the symptoms caused by a WRN mutation is approximately 10 – 20 years of age. Various methods of diagnosis have been established based on the diseases’ cardinal and secondary signs and symptoms. Since this disease is extremely rare, it is important to follow the established standards for diagnosis of WS. Two diagnostic criteria have been established in order to standardize diagnosis based on phenotype presentation. The International Registry of Werner Syndrome has developed a list of cardinal and secondary symptoms that may be used to diagnose WS. The cardinal signs and symptoms, based on an onset after ten years of age, include bilateral cataracts, characteristic skin, “bird like” faces (i.e. a pinched nasal bridge and loss of subcutaneous tissue), short stature, premature graying and/or thinning of scalp hair, consanguinity, and positive 24-hour urinary hyaluronic acid test (8). The secondary symptoms that have been described are type 2 diabetes mellitus, hypogonadism, osteoporosis, evidence of ostoesclerosis of the distal phalanges or toes, soft tissue calcificiation, atherosclerosis, neoplasms, flat feet, and abnormally high pitched voice (8,9). A definite diagnosis is defined as one in which all of the cardinal symptoms plus two secondary are identified. A probable diagnosis is one in which three cardinal signs plus two secondary are identified. Finally, a possible diagnosis is one in which either cataracts or dermatologic alterations and any four of the secondary symptoms are identified. A second method of diagnosing WS was established in 1997. This proposed method is defined as one in which four of the five following symptoms are identified: consanguinity, characteristic facial appearance, premature senescence, scleroderma-like skin changes, and endocrine-metabolic disorders (10). Despite the fact that these phenotype based standards have been set, it is important to use diagnostic and genetic testing in order to diagnose WS. One of the most reliant and widely used diagnostic tests for WS is a test for hyperhyaluronic aciduria. WS patients show significantly higher levels of hyaluronic acid in serum and urine than same aged controlled levels. The serum and urine hyaluronic acid levels of WS patients are almost equal to those of normal controls over 80 years old (12). Although this diagnostic test is normally helpful in determining the diagnosis of WS, WS patients normally require genetic analysis to confirm the diagnosis. WS Genetic Testing Since the WRN gene is the only known gene to be associated with Werner syndrome, defining the parameters for genetic testing is well determined. A genetic sequence analysis of the WRN coding region detects mutations in both alleles for approximately 90% of all individuals affected by WS (10). A false negative may be determined if the genetic mutation is located in the intron, which is normally not sequenced for genetic mutation. If a mutation is identified, a Western Blot is performed in order to determine the amount of protein (leucocytes) production that is affected by the mutation. Patient Counseling and Family Planning Following the diagnosis of WS, a physician must counsel his or her patient in regards to the management of the disorder and family planning. At the time of initial diagnosis, the physician should suggest that the patient undergo further medical testing in order to determine the extent of significant findings and possible treatments. Type 2 diabetes mellitus is favorably controlled in many patients who are diagnosed with WS (11). Surgical treatment is used to treat ocular cataracts in patients with WS. Most other malignancies are treated with standard treatments. The large amount of secondary complications that are caused by WS must also be considered during patient counseling. Patients should be directed to continue surveillance of the various complications that may be caused by WS (2). The patient should be advised to maintain a healthy lifestyle in order to decrease the risk of developing atherosclerosis at a young age (2). Depending on the age of the patient, a discussion of the possibility of family planning may be necessary. Many patients diagnosed with WS will experience a decline in fertility soon after sexual maturity. Testicular atrophy and loss of primordial follicles have been hypothesized as reasons why fertility declines following sexual maturity(2). This high rate of infertility may cause patients to feel depressed or isolated. Those patients who are determined to be fertile may request additional genetic planning in order to determine risks involved with offspring. One of the most important features of the WRN mutation is its high concentration among the Japanese population. In Japan, the frequency of heterozygosity is between 1/20,000 – 1/40,000 (3). Compared to the US population, which is predicted to be approximately 1/200,000, the Japanese have a much greater risk of carrying a mutant allele. Worldwide, 1200 patients were reported from 1904 to 1996, and 845 of these patients were from Japan (3). The WRN mutation is inherited in an autosomal recessive manner. The rarity of this allele in the United States population makes the risk of a carrier’s child developing this disease to be extremely low. This risk, however, is greatly increased if the couple is in a consanguineous relationship. The high allele frequency within the Japanese population must also be taken into consideration when family planning is being discussed. Finally, since no prenatal testing for WS is available, parental genetic testing should be recommended for those who are planning a family. One of the most difficult aspects of WS for many patients is dealing with the emotional and psychological aspects of knowing that the age of mortality for this disease is approximately 45 years of age. Mortality is most commonly due to atherosclerotic heart failure or malignant tumor development (2). Discussion of these issues with WS some patients may develop during the treatment process. It is important that the physician remembers these facts during his or her counseling of the patient. Despite the fact that WS is a rare genetic disorder, it may manifest as a physically, emotionally, and psychologically damaging disorder. Population genetics serves as a valuable resource when determining a patient’s risk of developing WS. Following an appropriate clinical workup, a diagnosis of WS may be established. The physician must recognize the various complications that are associated with this disorder, and must be prepared to treat each or refer the patient to someone who can treat them. References 1. Opresko, P. L., Cheng, W., Kobbe, ,C., Harrigan, J.A., and Bohr, V.A. (2003) Werner syndrome and the function of the Werner protein; what they can teach us about the molecular aging process. Carcinogenesis, Vol. 24, No. 5, 791-802 2. Hanson, N., Martin, G.M., Oshima, J. (2005) Werner Syndrome. Gene Review 3. Satoh M, Imai M, Sugimoto M, Goto M, Furuichi Y (1999) Prevalence of Werner's syndrome heterozygotes in Japan. Lancet 353:1766 4. Salk,D., Bryant,E., Hoehn,H., Johnston,P. and Martin,G.M. (1985) Growth characteristics of Werner syndrome cells in vitro. Adv. Exp. Med. Biol., 190, 305– 311. 5. Cooper,M.P., Machwe,A., Orren,D.K., Brosh,R.M., Ramsden,D. and Bohr,V.A. (2000) Ku complex interacts with and stimulates the Werner protein. Genes Dev., 14, 907–912. 6. Blander,G., Kipnis,J., Leal,J.F., Yu,C.E., Schellenberg,G.D. and Oren,M. (1999) Physical and functional interaction between p53 and the Werner's syndrome protein. J. Biol. Chem., 274, 29463–29469. 7. Wyllie,F.S., Jones,C.J., Skinner,J.W., Haughton,M.F., Wallis,C., WynfordThomas,D., Faragher,R.G. and Kipling,D. (2000) Telomerase prevents the accelerated cell ageing of Werner syndrome fibroblasts. Nature Genet., 24, 16– 17. 8. Nakura J, Wijsman EM, Miki T, Kamino K, Yu CE, Oshima J, Fukuchi K, Weber JL, Piussan C, Melaragno MI, et al. (1994) Homozygosity mapping of the Werner syndrome locus (WRN). Genomics 23:600-8 9. Tsunoda K, Takanosawa M, Kurikawa Y, Nosaka K, Niimi S (2000) Hoarse voice resulting from premature ageing in Werner's syndrome. J Laryngol Otol 114:61-3 10. Goto M (1997) Hierarchical deterioration of body systems in Werner's syndrome: Implications for normal ageing. [Clinical review of Japanese Werner syndrome cases] Mech Ageing Devel 98:239-254 11. Yokote K, Honjo S, Kobayashi K, Fujimoto M, Kawamura H, Mori S, Saito Y (2004) Metabolic improvement and abdominal fat redistribution in Werner syndrome by pioglitazone. J Am Geriatr Soc 52:1582-3 12. Tanabe M and Goto M (2001) Elevation of serum hyaluronan level in Werner's syndrome. Gerontology 47:77-81