SiRAD6 Increase UV Sensitivity In Caenorhabditis Elegans

Objective:ultraviolet(uv)is a important source of DNA damage.Living cells will start DNA damage repair mechanism after DNA damage.The repair pathways including nucleotide excision repair(NER)and the homologous end connection(HR).RAD6(UBC-1)is a E2 ubiquitin conjecting enzymes.DNA damage repair defects will lead to the body's increase sensitivity to uv damage,induce mutation and tumorigenesis.This study by explore the funcito of RAD6(UBC-1)in DNA repair by RNA interference in c.elegans.Methods:Obtain the total RNA of wild type c.elegans by RNA extraction,obtained cDNA library by reverse transcription PCR.With specific primers amplification(UBC-1)gene.By agarose electrophoresis gene purification purposes.Clone the UBC-1 into L4440 plasmid and thansform to oliHT15(DE3),We performed SiUBC-1 by feeding the elegans with L4440-UBC-1 transformed DE3.DE3 with L4440 as control.DNA damage was induced by different doses of UV irradiation to the elegans.The lifecycle,egg number and related mRNA expression were detected.Results:UBC-1 gene cloning and L4440 RNA interfere plasmids constructed.UBC-1 mRNA level detection by TR-PCR.The UBC-1 gene mRNA level was down regulated 78.1%conpared with wild type elegans.Determination mortality rate of the elegans under different dose of UVC irradiation.The number of eggs,the hatching rate and the head oscillating frequency counts were calculated.eggs and egg hatching are significantly reduced in SiUBC-1 elegans.RNAi of UBC-1 induce more sensitive to radiation in elsganse,.Analysis of related mRNA level were determinged by real-time RT-PCR.,Total RNA were extracted 0,4,15,20 h after 100 mj UVC irradiation.RT-PCR results show that the SiUBC-1 elegans,MRE-11 and CEP-1 expression were significantly higher after UV irradiation.Conclusions:RAD6 as ubiquitin cross-linking enzyme E2,a key enzyme in the growth of c.elegans,maintain the normal form of c.elegans,germ cell development and play an important role in the process of such as eggs hatch;SiUBC-1 induce the life span decreased in the c.elegans,the spawning and hatching rate decreased obviously.SiUBC-1 results in DNA damage repare deficiat and induced more appoptosis in germ cells.UV induced germ cell appoptosis by MRE-11,CEP-1 pathway in SiUBC-1 elegans.RAD6 repair the DNA damage by XPC-1,XPA-1and XPG-lpathway in C.elegans.