Tiffanie Lee Quantitative Biology Dr Brian Strahl Department of Biochemistry and Biophysics How does ASH1L read chromatin What histone reader domains are involved in binding ASH1L and MLL cooperate to maintain hematopoiesis and transcription in immune cells in the blood ID: 1009303
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1. Determining the Histone Post-translational Modification Binding Specificities of ASH1L Histone Reader DomainsTiffanie LeeQuantitative BiologyDr. Brian Strahl, Department of Biochemistry and Biophysics
2. How does ASH1L read chromatin? What histone reader domains are involved in binding?ASH1L and MLL cooperate to maintain hematopoiesis and transcription in immune cells in the bloodMixed-lineage leukemias result from chromosomal translocations. These cases are frequently acute and may be resistant to chemotherapy, resulting in poor outcomes, esp. for childrenInhibiting ASH1L binding can potentially treat MLL-rearranged leukemiasHypothesis: The BRD, BAH, and PHD domains are involved in binding to two post-translational modifications
3. Results1. Successfully made mutant constructs of GST-ASH1L mutants in histone reader domains using site directed mutagenesis2. Affinity purified single mutants and triple mutant3. Confirmed binding of ASH1L TR on H3 peptides with peptide pulldown assaysFuture directions:Reconfirm sequencing and troubleshoot issues with creating single point mutant constructsDetermine binding specificities of single- and double-point mutantsTest on nucleosomes, human embryonic kidney cellsBRDpm PHDpm BAHpmBRD PHD BAH TSDM1.2.3.