Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School
Member, Harvard Center for Cancer Research
The Buratowski Lab studies eukaryotic gene transcription, chromatin regulation, and RNA processing using a wide range of biochemical, molecular, and genetic techniques.
The Buratowski Lab studies eukaryotic gene expression, concentrating on RNA polymerase II transcription and its coupling to RNA processing and chromatin modification. We are concentrating on three areas: (A) the functions and interactions of the RNA polymerase II (RNApII) transcription factors, (B) communication between chromatin and transcription complexes, particularly through non-coding transcription, and (C) mRNA processing enzymes and their interactions with RNApII. Using the yeast Saccharomyces cerevisiae, a combination of biochemical and genetic techniques are being brought to bear on these questions. Several dozen proteins are required simply to initiate transcription, and many more are required for processes linked to transcription. Therefore, it is now necessary to decipher the functions of each of the individual factors. Some of our recent projects:
1. The RNApII C-terminal domain (CTD) and mRNA processing enzymes. We discovered that the phosphorylated CTD acts as a binding site for mRNA processing enzymes to link transcription and mRNA processing. The CTD phosphorylation changes at various points of transcription initiation and elongation, binding different sets of factors involved in regulation of elongation, termination, capping, splicing, and polyadenylation. We are comparing this process on coding and non-coding transcription units. In collaboration with Jarrod Marto, we are using mass spectrometry to analyze the factors associated with transcription initiation and elongation complexes.
2. Factors that modulate transcription RNApII elongation and termination. We showed that different mechanisms are used for termination at different classes of genes. Genes that encode polyadenylated mRNAs use an exonuclease-dependent pathway, while genes for the non-polyadenylated sn/snoRNAs use a pathway that includes the exosome and the Nrd1 and Nab3 RNA binding proteins. We are working to further understand the two pathways and how the choice is made between them.
3. Connections between transcription and chromatin structure. Transcription causes major changes in the nucleosomes that package the gene. For example, the histone methyltransferases Set1 and Set2 are targeted to promoter and coding regions, respectively, via binding to the phosphorylated RNApII CTD. These transcription-coupled histone methylation patterns have been linked to human cancers, but yeast provides a perfect model system for getting at their basic functions. Recently, we found that the Set3 histone deacetylase mediates the effects of non-coding transcription on overlapping coding genes.
Set3 HDAC mediates effects of overlapping noncoding transcription on gene induction kinetics. Cell 150(6):1158-69 (2012)
Direct analysis of phosphorylation sites on the Rpb1 C-terminal domain of RNA polymerase II. Mol. Cell 61, 297-304 (2016)
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