| Telomeres are specialized structures at the ends of eukaryotic chromosomes and are composed of conserved sequences of DNA repeats. Telomeric DNA is not completely synthesized by conventional DNA polymerase, but progressively shorten with each cellular division. Cellular aging is characterized by a decrease in telomere length, and this has been implicated as a mitotic clock that signals cells to stop division when telomeres reach a critically short length. Telomerase is a ribonucleoprotein that synthesizes telomere repeats onto chromosome ends and is involved in maintaining telomere length in germline and cancer cells( > 90 % ).The catalytic subunit (TERT) appears to be the limiting component in most telomerase-negative cells. As TERT is expressed constitutively, the presence of telomerase activity is correlated with TERT mRNA expression in extracts from tissue culture cells and normal and cancer tissues.In order to investigate the expression pattern of TERT in reproductive system cells of mouse and address the biological significance of TERT, both immunohistochemistry and in situ hybridization experiments were applied to explore the pattern of TERT expression in different differentiated reproductive cells in the testis and ovaries of mouse. Furthermore, in order to investigate the relationship between TRET and cell cycle related genes and the regulative functions of TERT in proliferation of mouse cancer cells, techniques of RNA interference and gene chip were used to analyze the expression of cell cycle related genes after RNA interference targeted TERT in EL-4 lymphoma cells of mouse. The main results are as following:1. Spermatogonia and sertoli cells are found in the testis of 10-day mice;but there is no positive expression of TERT protein and mRNA. In testis of 20-day mice, active meiosis and mitosis are found, and there are large numbers of primary spermatozoas, in which there are highly positive TERT expression. In testis of 30-day mice, seminiferous epithelium has differentiated into secondary spermatocytes and spermatids cells, and some sperms have been formed. Meanwhile, TERT positive expression can be found in the primary spermatocytes and spermatids cells. In primary spermatocytes, especially inpachytene spermatocytes, the strongest TERT expression level was detected. The seminiferous tubules cavity in testis of 60-day and 90-day mice are expanded and TERT positive expressions can only be detected in the spermatids cells and primary spermatocytes. TERT expression pattern during the process of sperm generation indicates a weak-strong-weak trend of the dynamic character, which may be closely related with the meiosis in spermatogenesis and have important regulative effect on the proliferation and differentiation of different levels of sperm-generation cells. And the role of TERT during the spermatogenesis also ensures the transmission of full-length chromosomes to progeny.2. During the oogenesis, no positive TERT expression has been found in the oocytes of 10-day, 20-day, 30-day , 60-day and 90-day mice. Strongly TERT positive expression is found in the follicle cells of primordial follicles. This indicates that the follicle cells are main source of telomerase activity in ovaries. During the development of ovary the expression TERT becomes weaker and weaker, which is closely related with the growth of the follicles. The relation between TERT and follicle cells possibly influences not only proliferative capacity of follicle cells, but also communion between follicle cells and oocytes.3. The results of RT-PCR and Western blot show that, after transfection to the EL-4 lymphoma cells of mouse, the TERT specialized shRNA effectively repressed the expressions of endogenous TERT mRNA and protein, which made the targeted genes partially silent. The MTT experiment demonstrates that the amount of EL-4 cells dose not show significant difference between the sample treated with TERT specialized shRNA and the contrast without treatment. This result indicates that the repression of TERT expressions does not essentially lead to apparent death of cells at the time of detection, which may due to the phenotypic lag phenomenon caused by the inhibition of telomerase.4. According to the result of gene chip analysis, among the 96 cell cycle genes of mice, 43 genes show significant expression difference between the sample treated with TERT specialized shRNA and the contrast, and all of 43 genes belong to the down-regulated gene. According to the biological functions of the genes in the cell cycles of mice ,they can be divided into five groups: (1) Genes encoding cyclin, such as CyclinAl, CyclinB2, CyclinE2, CyclinF, CyclinG1, CyclinH etc.. ( 2 ) Genes encoding cyclin-dependent kinase, such as CDK4, CDK5, CDK7 etc.. (3) CDC25B...
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