The Structural And Functional Research Of Transcription Factor Cdc73from Saccharomyces Cerevisiae

Pafl complex which contains Pafl, Cdc73, Ctr9, Leol and Rtfl (recently discovered hSki8is one subunit in human Pafl complex) subunits is a conserved complex in eukaryotes. The main function of Pafl complex is taking part in transcription including initiation, elongation and termination together with RNA pol II. As one of the first identified subunits of PaflC, yeast Cdc73(yCdc73) takes part in many transcription-related processes, including binding to RNA polymerase II, recruitment and activation of histonemodification factors and communication with other transcriptional activators. The human homologue of yCdc73, parafibromin, has been identified as a tumour suppressor linked to breast, renal and gastric cancers.In this work, we targeted yeast Cdc73for structural studies and determined the C-terminal structure of yCdc73. The C-terminal of Cdc73is very conserved and a DALI server search indicates the C-terminal structure of Cdc73is a canonical a6/p6GTPase-like fold, although the primary sequence of it has very low identity to GTPases and is not conservative at the key region. No significant GTP-binding ability and hydrolysis activity were detected by ITC and enzyme activity assay. So Cdc73, with a GTPase-like fold, may not be a GTPase. The determination of the structure of Cdc73has provided further insights into the assembly way of Pafl complex and the structural basis for study on Cdc73-RNA PolⅡ interaction and human related diseases.DNA, the carrier of genetic information, is always induced a wide range of lesions by exogenous or endogenous DNA damaging agents. The genome will be instabile or re-arrange if the lesions of DNA are not repaired, and then some related diseases will occur. There are many DNA repair pathways to different DNA lesions, including base excision repair, nucleotide excision repair, mismatch repair and homologous recombination repair. In addition to these accurate repair pathways, when presence of replication-blocking lesions, there is another DNA damage tolerance (DDT) mechanism which does not remove DNA damages in the template strand, but bypass the DNA lesions during replication to prevent replication fork collapse, which is also named post-replication repair (PRR). An error-free pathway within PRR is very similar to the Rad51dependent homologous recombination repair (HRR). The yeast Shu complex is comprised of four subunits, Csm2, Psy3, Shu land Shu2, which promotes Rad51dependent homologous recombination via inhibit the activity of Srs2. The human orthologous of yeast Psy3, Shul and Shu2are Rad51D, Xrcc2and Sws1respectively. Sws1, the Shu2orthologous in fission yeast, was reported to have the ability to interact with Srs2. A newly identified human Swsl associate protein, Swsapl can form a stable complex with Swsl and takes part in homologous recombination repair. We are trying to solve the structure of Swsl-Swsapl complex and explain its function mechanism.