title: A systematic mutational analysis of a histone H3 residue in budding yeast provides insights into chromatin dynamics.
abstract: In previous work using the Saccharomyces cerevisiae model system, a mutant version of histone H3-H3-L61W-was found to confer a variety of abnormal growth phenotypes and defects in specific aspects of the transcription process, including a pronounced alteration in the distribution pattern of the transcription elongation factor Spt16 across transcribed genes and promotion of cryptic transcription initiation within the FLO8 gene. To gain insights into the contribution of the H3-L61 residue to chromatin function, we have generated yeast strains expressing versions of histone H3 harboring all possible natural amino acid substitutions at position 61 (H3-L61X mutants) and tested them in a series of assays. We found that whereas 16 of the 19 H3-L61X mutants support viability when expressed as the sole source of histone H3 in cells, all 19 confer abnormal phenotypes ranging from very mild to severe, a finding that might in part explain the high degree of conservation of the H3-L61 residue among eukaryotes. An examination of the strength of the defects conferred by each H3-L61X mutant and the nature of the corresponding substituted residue provides insights into structural features of the nucleosome required for proper Spt16-gene interactions and for prevention of cryptic transcription initiation events. Finally, we provide evidence that the defects imparted by H3-L61X mutants on Spt16-gene interactions and on repression of intragenic transcription initiation are mechanistically related to each other.
authors.affiliation: Biology Department, Hendrix College, Conway, Arkansas 72032.
authors.forename: Paige
authors.initials: P
authors.lastname: Johnson
authors.affiliation: Biology Department, Hendrix College, Conway, Arkansas 72032.
authors.forename: Virginia
authors.initials: V
authors.lastname: Mitchell
authors.affiliation: Biology Department, Hendrix College, Conway, Arkansas 72032.
authors.forename: Kelsi
authors.initials: K
authors.lastname: McClure
authors.affiliation: Biology Department, Hendrix College, Conway, Arkansas 72032.
authors.forename: Martha
authors.initials: M
authors.lastname: Kellems
authors.affiliation: Biology Department, Hendrix College, Conway, Arkansas 72032.
authors.forename: Sarah
authors.initials: S
authors.lastname: Marshall
authors.affiliation: Biology Department, Hendrix College, Conway, Arkansas 72032.
authors.forename: Mary K
authors.initials: MK
authors.lastname: Allison
authors.affiliation: Biology Department, Hendrix College, Conway, Arkansas 72032.
authors.forename: Harrison
authors.initials: H
authors.lastname: Lindley
authors.affiliation: Biology Department, Hendrix College, Conway, Arkansas 72032.
authors.forename: Hoai-Trang T
authors.initials: HT
authors.lastname: Nguyen
authors.affiliation: Biology Department, Hendrix College, Conway, Arkansas 72032.
authors.forename: Jessalyn E
authors.initials: JE
authors.lastname: Tackett
authors.affiliation: Biology Department, Hendrix College, Conway, Arkansas 72032 duina@hendrix.edu.
authors.forename: Andrea A
authors.identifier: http://orcid.org/0000-0002-6040-6825
authors.initials: AA
authors.lastname: Duina
mesh_terms: D019943:Amino Acid Substitution; D002843:Chromatin / Q000378:metabolism*; D006657:Histones / Q000235:genetics*; D009154:Mutation; D009707:Nucleosomes / Q000737:chemistry; D010641:Phenotype; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000254:growth & development; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics; D061785:Transcription Initiation, Genetic; D035602:Transcriptional Elongation Factors / Q000235:genetics
chemical_list: D002843:Chromatin; D006657:Histones; D009707:Nucleosomes; C071238:SPT16 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D035602:Transcriptional Elongation Factors
keywords: Spt16; chromatin; histone H3; intragenic transcription initiation; transcription elongation