Identification of Uhp1, a ubiquitinated histone-like protein, as a target/mediator of Rhp6 in mating-type silencing in fission yeast.
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.
Naresh
Alpana
A
Institute of Microbial Technology, Sector 39A, Chandigarh-160 036, India.
Saini
Sharanjot
S
Singh
Jagmohan
J
D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D015152:Blotting, Northern; D015153:Blotting, Western; D002843:Chromatin / Q000378:metabolism; D002845:Chromatography; D003432:Cross-Linking Reagents / Q000494:pharmacology; D004247:DNA / Q000378:metabolism; D019281:Dimerization; D004926:Escherichia coli / Q000378:metabolism; D005418:Flavodoxin / Q000737:chemistry* / Q000378:metabolism*; D017353:Gene Deletion; D020868:Gene Silencing; D005976:Glutaral / Q000494:pharmacology; D005982:Glutathione Transferase / Q000378:metabolism; D006657:Histones / Q000378:metabolism; D007455:Iodine / Q000494:pharmacology; D008025:Ligases / Q000737:chemistry* / Q000378:metabolism*; D008239:Lysine / Q000737:chemistry; D008540:Meiosis; D008969:Molecular Sequence Data; D009154:Mutation; D011233:Precipitin Tests; D011485:Protein Binding; D017510:Protein Folding; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D011994:Recombinant Proteins / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D016196:S Phase; D012441:Saccharomyces cerevisiae / Q000378:metabolism; D012568:Schizosaccharomyces / Q000378:metabolism*; D029702:Schizosaccharomyces pombe Proteins / Q000737:chemistry* / Q000378:metabolism*; D017386:Sequence Homology, Amino Acid; D013997:Time Factors; D025801:Ubiquitin / Q000378:metabolism*; D044763:Ubiquitin-Conjugating Enzymes
D002843:Chromatin; D003432:Cross-Linking Reagents; D005418:Flavodoxin; D006657:Histones; C093707:OBR1 protein, S pombe; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; D029702:Schizosaccharomyces pombe Proteins; D025801:Ubiquitin; D004247:DNA; D007455:Iodine; D044763:Ubiquitin-Conjugating Enzymes; C063863:rhp6 protein, S pombe; D005982:Glutathione Transferase; D008025:Ligases; D008239:Lysine; D005976:Glutaral