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Regulatory Role of ALU RNA Transcripted by RNA Polymerase Ii on PKR Phosphorylation

Author: YangMei
Tutor: ShenZuo
School: Chongqing Medical University
Course: Internal Medicine
Keywords: ALU sequence PKR Apoptosis RNA polymerase II Interferon
CLC: Q255
Type: Master's thesis
Year: 2010
Downloads: 22
Quote: 0
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Abstract


Objective: To study the ALU own RNA polymerase III promoter RNA polymerase II transcription , to explore the physiological function of the RNA polymerase II transcription ALU . Methods : ( 1 ) full- Alu gene sequence into pcDNA3.1 vector downstream of the CMV promoter , to construct the recombinant plasmid with pcDNA3.1- ALU ; ( 2 ) transfected 293 cells , extracted total cellular RNA by RT- PCR screening stably expressing ALU cloning ; ( 3 ) Western- blot assay stable expression the PKR phosphorylation level PCDNA3.1-ALU293 cells and interferon . ( 4 ) MTT and Cellular DNA Fragmentation ELISA assay stably expressing PCDNA3.1-ALU293 cell proliferation inhibition of apoptosis . (5) MTT Cellular DNA Fragmentation ELISA assay transient expression PCDNA3.1-ALU293 cell proliferation inhibition of apoptosis . (6) MTT Cellular DNA Fragmentation ELISA assay transient expression of the cell group of PCDNA3.1-ALUHepG2 proliferation inhibition and apoptosis . Results : (1) Alu gene sequence inserted into the pcDNA3.1 vector directly downstream of the CMV promoter could be efficient transcription of RNA polymerase II described ALU own RNA polymerase III promoter , does not affect the RNA polymerase II transcription . (2) western-blot results show that : ALU transcribed by RNA polymerase II is able to activate PKR activity . (3) Apoptosis : The results show that the RNA polymerase II transcription of the ALU and the interferon - treated cells cells proliferation was suppressed, and as compared with the control group , and more susceptible to apoptosis . Conclusion : ALU own RNA polymerase III promoter had no significant effect on the RNA polymerase II transcription , but by the formation of double-stranded RNA activated PKR activity , induce apoptosis .

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