Identification of Uhp1, a ubiquitinated histone-like protein, as a target/mediator of Rhp6 in mating-type silencing in fission yeast

J Biol Chem. 2003 Mar 14;278(11):9185-94. doi: 10.1074/jbc.M212732200. Epub 2003 Jan 2.

Abstract

Mating-type silencing in Schizosaccharomyces pombe is brought about by cooperative interactions between cis-acting DNA sequences flanking mat2P and mat3M and the trans-acting factors, namely Swi6, Clr1-Clr4, Clr6, and Rik1. In addition, DNA repair gene rhp6, which plays a role in post-replication DNA repair and ubiquitination of proteins including histones, is also involved in silencing, albeit in a unique way; its effect on silencing and chromatin structure of the donor loci is dependent on their switching competence. Earlier, we hypothesized the existence of a mediator of Rhp6 that plays a role in reestablishment of the chromatin structure coincidentally with DNA replication associated with mating-type switching. Here we report the identification of a 22-kDa protein as an in vivo target and mediator of Rhp6 in mating-type silencing. The level of this protein is greatly elevated in sng1-1/rhp6(-) mutant and rhp6Delta as compared with wild type strain. Both the deletion and overexpression of the gene encoding this protein elicit switching-dependent loss of silencing. Furthermore, the 22-kDa protein undergoes Rhp6-dependent multiubiquitination and associates with mat2 locus during S phase in wild type cells. Interestingly, it contains a histone-fold motif similar to that of histone H2A, and like histone H2A, it interacts strongly with histone H2B in vitro. These results indicate that the 22-kDa protein, renamed as the ubiquitinated histone-like protein Uhp1, is an in vivo target/mediator of Rhp6 in silencing. Thus, regulation of association of Uhp1 with chromatin and ubiquitination followed by degradation may play a role in reestablishment of inactive chromatin structure at the silent mating-type loci.

MeSH terms

  • Amino Acid Motifs
  • Amino Acid Sequence
  • Blotting, Northern
  • Blotting, Western
  • Chromatin / metabolism
  • Chromatography
  • Cross-Linking Reagents / pharmacology
  • DNA / metabolism
  • Dimerization
  • Escherichia coli / metabolism
  • Flavodoxin / chemistry*
  • Flavodoxin / metabolism*
  • Gene Deletion
  • Gene Silencing
  • Glutaral / pharmacology
  • Glutathione Transferase / metabolism
  • Histones / metabolism
  • Iodine / pharmacology
  • Ligases / chemistry*
  • Ligases / metabolism*
  • Lysine / chemistry
  • Meiosis
  • Molecular Sequence Data
  • Mutation
  • Precipitin Tests
  • Protein Binding
  • Protein Folding
  • Recombinant Fusion Proteins / metabolism
  • Recombinant Proteins / metabolism
  • Reverse Transcriptase Polymerase Chain Reaction
  • S Phase
  • Saccharomyces cerevisiae / metabolism
  • Schizosaccharomyces / metabolism*
  • Schizosaccharomyces pombe Proteins / chemistry*
  • Schizosaccharomyces pombe Proteins / metabolism*
  • Sequence Homology, Amino Acid
  • Time Factors
  • Ubiquitin / metabolism*
  • Ubiquitin-Conjugating Enzymes*

Substances

  • Chromatin
  • Cross-Linking Reagents
  • Flavodoxin
  • Histones
  • OBR1 protein, S pombe
  • Recombinant Fusion Proteins
  • Recombinant Proteins
  • Schizosaccharomyces pombe Proteins
  • Ubiquitin
  • DNA
  • Iodine
  • Ubiquitin-Conjugating Enzymes
  • rhp6 protein, S pombe
  • Glutathione Transferase
  • Ligases
  • Lysine
  • Glutaral