ANALYSIS OF A FUNCTIONAL ROLE FOR CONSERVED LYSINE RESIDUES OF THE H4 N-TERMINUS IN PREMATURE mRNA SPLICING EFFICIENCY

Open Access
Author:
Joy, Jaimy
Area of Honors:
Biochemistry and Molecular Biology
Degree:
Bachelor of Science
Document Type:
Thesis
Thesis Supervisors:
  • Joseph C. Reese, Thesis Supervisor
  • David Scott Gilmour, Honors Advisor
  • Wendy Hanna Rose, Faculty Reader
Keywords:
  • Histone
  • H4
  • splicing
Abstract:
Chromatin, repeating units of tightly coiled DNA wrapped around histones, is the most important level of DNA organization. The N-termini of histones are subject to a variety of post-translation modifications that can either promote or discourage vital cell processes, such as DNA replication, transcription, and splicing. Recent studies have established a vital role for histone H3 in splicing, but little investigation concerning H4 has been done. Therefore, this project sought to determine a functional role for lysine residues of the histone H4 in splicing efficiency in S. cerevisiae. Reporter assays revealed that H4 mutants with all native lysine residues of H4 modified to glutamine, a splicing defect was observed. In an H4 mutant with lysine to glutamine point mutations at positions 5, 8, and 12 with a tripeptide insertion, GKG, at position 3, the splicing defect was moderately rescued. Interestingly, when pre-mRNA leakage of each of the aforementioned strains was analyzed, the results were opposite of the expected outcome. The wildtype strain showed the greatest pre-mRNA leakage while the H4 all K to Q strain showed the least. The low beta-galactosidase activities, however, may partially explain this anomaly. Preliminary qPCR analyses using ECM33 as the gene of interest do not show splicing defects in the H4 mutants compared to wildtype, but this may be attributed to the abundance of the gene. Future investigations and analyses using qPCR are of high priority.