COEXPRESSION AND HISTONE ACETYLTRANSFERASE ACTIVITY OF HMOF/HMSL1V1 COMPLEX

Open Access
Author:
Aurori, Kieran James
Area of Honors:
Biochemistry and Molecular Biology
Degree:
Bachelor of Science
Document Type:
Thesis
Thesis Supervisors:
  • Song Tan, Thesis Supervisor
  • Ming Tien, Honors Advisor
  • Wendy Hanna Rose, Faculty Reader
Keywords:
  • MOF
  • MSL1v1
  • histone
  • nucleosome
  • histone acetyltransferase
  • protein
  • chromatin
  • dosage compensation
  • epigenetics
Abstract:
Nuclear DNA contains all of the information required for eukaryotic life despite having to fit within the very confined space afforded by the nuclear envelope. In order to accomplish this task, DNA wraps around histone proteins creating nucleosomes, which, with connecting DNA, subsequently pack into chromatin and higher order structures. Regulation of this chromatin packing is governed by a series of epigenetic modifiers including histone acetyltransferases (HATs). A notable HAT complex is the MSL complex, conserved from Drosophila to humans. The Drosophila MSL complex is involved in dosage regulation and contains the catalytic males absent on the first (MOF) subunit as well as MSL1, MSL2 and MSL3. Both Drosophila and human complexes acetylate a lysine residue on the N-terminal tail of histone H4 protein, decreasing DNA interaction with histone proteins and increasing transcription. The catalytic subunit of the human MSL complex, hMOF, also interacts with a distant relative of the hMSL1 subunit, hMSL1v1. The hMSL1v1/hMOF complex not only acts catalytically to acetylate the histone H4 protein, but also has implications in human tumorigenesis. In order to provide a recombinant source of the hMSL1v1/hMOF complex for biochemical and biophysical studies, I have coexpressed and purified the hMSL1v1/hMOF complex from E. coli using a polycistronic expression system. I was able to observe expression of the individual subunits through two separate tagged versions of the complex and to positively identify the hMOF subunit in each tagged version through Western Blot analysis. The complex also showed catalytic activity on histone proteins and on the nucleosome in a histone acetyltransferase assay.