Study On The Expression And Mutation Of Telomere Binding Proteins In Acute Leukemia Cells And Their Effects On Telomerase

Many studies have reported telomerase is tight-related with malignant hematological diseases.Telomere shortens and telomerase activation was found both in malignant hematological cell line cells and leukemia patients' samples. However, the mechanism of telomerase activation is unclear. The regulation of telomerase activity could be explained by many aspects, telomere binding protein is one of the new one. Since the first telomere binding protein Telomere Repeat Binding Factor (TRF1) was reported in 1995, about 10 proteins have been reported. They located around telomere, participate in many important cell activities as maintaining telomere structure, mitosis regulation, cell cycle regulation and DNA damage repair etc. Among them, 4 proteins were reported regulate telomerase activity, they are TRF1, Pinxl, tankyrase and tankyrase-2. Here, TRF1 and Pinxl are inhibitors of telomerase activity, while tankyrase and tankyrase-2 are enhancers of telomerase activity. Their expressions and relation with telomerase in acute leukemia cells is unclear.In this study, mRNA expressions of these 4 telomere binding proteins in AL cells were detected and their relations with the mRNA expression of telomerase reverse transcriptase gene (hTERT) were analyzed at first. Then the expression and mutation of TRF1 in malignant hematology cell line cells was analyzed. After that a further study on the telomerase activity and hTERT mRNA expression along with 4 telomere binding proteins' expression during the differentiation of NB4 cells induced by ATRA was performed to analyze the relation of telomere binding proteins with telomerase during differentiation, so as to indicate the possible mechanism of telomere binding proteins' effects on telomerase. Expression of 4 telomere binding proteins in leukemia cells have not been reported, nor have the studies on the mutation analysis of TRF1 in malignant hematological cell line cells and the studies of the expression of telomere binding proteins and telomerase activity during differentiation of NB4 cells were reported.This paper was divided into three parts.Part I : Real-time quantitative RT-PCR with fluorescence probe hybridization was used todetect the expression of 4 telomere binding proteins in 30 samples of AL patients, analyze their relation with hTERTmRNA expressions.The 30 AL cases are all enrolled at their first diagnose, 18 were male and 12 female, average age is 32, 17 samples were of acute non-lymphocytic leukemia (ANLL) while 13 of acute lymphoblastic leukemia (ALL), 10 patients' peripheral WBC was above lOO 10~9 /L while 20 were below 100 109 /L at diagnose, 14 were with abnormal karyotype and 16 with normal karyotype.The expression of hTERTmRNA in AL samples (0.00244, 0-0.0320) is significantly higher than in normal mononuclear cells (2.79X10-4, 0-0.0078)( P<0.001). Its expression has no significant difference between ANLL and ALL cells (P=0.742) . TRF1 mRNA expression in AL samples(0.0171, 0.0127-0.193)is lower than that in normal mononuclear cells(0.0457, 0.00839-0.262) (P =0.019). Its expression in ANLL (0.0126, 0.0127-0.0546) is lower than that in normal mononuclear cells (P<0.001) , but its expression in ALL cells (0.0745, 1.92 l0-6-0.193) has no significant difference with that in normal mononuclear cells(P=0.761 ).TRF1 expression in ANLL is significantly lower than that in ALL cells( P=0.001). The expression of TRFI mRNA in AL samples and normal mononuclear cell samples has no significant correlation with hTERTmRNA expression(r=-0.173, P=0.207) . Pinxl mRNA expression in AL samples (0.00312, 5.42 l0-4~0.024) is significantly higher than that in normal mononuclear cells (7.89 l0-4, 0-0.00863) (P=0.002). Its expression in ANLL and ALL cells has no significantly differences (P=0.869) . The expression of Pinxl mRNA in AL samples and normal mononuclear cell samples has significant positive correlation with hTERTmRNA expression (r=0.296, P=0.028) . tankyrase mRNA expression in AL(0.0209, 0.0106-0.0997) is significantly higher than that in normal mon...