Regulation of donor preference during yeast mating-type switching
Abstract (Summary)iii Mating-type switching in Saccharomyces cerevisiae is directional. MATa cells choose HML? for recombination and MAT? cells choose HMRa. This is called “donor preference” and controlled by the recombination enhancer (RE). Donor preference during mating-type switching has been attributed to differences in chromatin structure for the left arm of chromosome III. I mapped the structure of ~45 kbp of the left arm of chromosome III in a and ? cells in logarithmically growing cultures and in a cells during switching. Other than the RE, chromatin structure was identical in the two cell types. Changes in chromatin structure during switching were confined to RE and HML. My analysis indicates that primary chromatin structure does not cause the documented differences in recombinational frequency of the left arm of chromosome III in a and ? cells. In the process of addressing the role of chromatin in mating type switching, I carried out the first study of chromatin structure spanning an extensive region in an organism whose genomic sequence is known. This region includes thirty ORFs, six potential replication origins, two known silencers and a recombination enhancer, thus may serve as a representative of overall genome organization. Significant features of organized chromatin exist for the entire region. DNase I hypersensitive sites reside at the promoter region of nearly every gene, suggesting that a basic chromatin structural feature exists at every promoter. ~25% of the ORFs possess extended regions of positioned nucleosomes. My study on the chromatin structure of an extended region contributes to understanding how yeast organizes its primary chromatin structure. A number of a-specific non-coding RNAs are transcribed from the RE locus, iv downstream of the two ?2/Mcm1 operators. This transcription is cell cycle regulated. Additionally, proper mating-type switching and donor preference require a cell-cycle regulated factor. Mcm1 and Fkh1 regulate RE activity in a cells. Mcm1 binding is required for both RE transcription and Fkh1 binding. This requirement can be bypassed by inserting another promoter into the RE, which increases donor preference and opens the chromatin structure of RE. The data presented in this thesis suggest that the role of Mcm1 in a-cell donor preference is to activate RE by providing a promoter-related function such as recruiting the chromatin remodeling proteins and/or other transcription factors. Transcription activation and open chromatin enhances Fkh1 binding and the level of this binding determines the level of donor preference. My study on RE activation helps understanding the mechanism of RE function in increasing the rate of recombination between HML and MAT.
School Location:USA - Pennsylvania
Source Type:Master's Thesis
Date of Publication: