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Molecular and Cellular Biology, December 2002, p. 8366-8374, Vol. 22, No. 23
0270-7306/02/$04.00+0     DOI: 10.1128/MCB.22.23.8366-8374.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.

Two Ubiquitin-Conjugating Enzymes, Rhp6 and UbcX, Regulate Heterochromatin Silencing in Schizosaccharomyces pombe

Eun Shik Choi,1 Hyun Soo Kim,1 Yeun Kyu Jang,1,2* Seung Hwan Hong,1 and Sang Dai Park1*

School of Biological Sciences, Seoul National University, Seoul 151-742,1 Branch of Lung Cancer Research, Division of Common Cancer Research, National Cancer Center, Goyang 411-764, Republic of Korea2

Received 5 June 2002/ Returned for modification 23 July 2002/ Accepted 28 August 2002

Methylation of histone H3 has been linked to the assembly of higher-order chromatin structures. Very recently, several examples, including the Schizosaccharomyces pombe mating-type region, chicken ß-globin locus, and inactive X-chromosome, revealed that H3-Lys9-methyl (Me) is associated with silent chromatin while H3-Lys4-Me is prominent in active chromatin. Surprisingly, it was shown that homologs of Drosophila Su(var)3-9 specifically methylate the Lys9 residue of histone H3. Here, to identify putative enzymes responsible for destabilization of heterochromatin, we screened genes whose overexpressions disrupt silencing at the silent mat3 locus in fission yeast. Interestingly, we identified two genes, rhp6+ and ubcX+ (ubiquitin-conjugating enzyme participating in silencing), both of which encode ubiquitin-conjugating enzymes. Their overexpression disrupted silencing at centromeres and telomeres as well as at mat3. Additionally, the overexpression interfered with centromeric function, as confirmed by elevated minichromosome loss and antimicrotubule drug sensitivity. On the contrary, deletion of rhp6+ or ubcX+ enhanced silencing at all heterochromatic regions tested, indicating that they are negative regulators of silencing. More importantly, chromatin immunoprecipitation showed that their overexpression alleviated the level of H3-Lys9-Me while enhancing the level of H3-Lys4-Me at the silent regions. On the contrary, their deletions enhanced the level of H3-Lys9-Me while alleviating that of H3-Lys4-Me. Taken together, the data suggest that two ubiquitin-conjugating enzymes, Rhp6 and UbcX, affect methylation of histone H3 at silent chromatin, which then reconfigures silencing.

* Corresponding author. Mailing address for Yeun Kyu Jang: Branch of Lung Cancer Research, Division of Common Cancer Research, National Cancer Center, Goyang 411-764, Republic of Korea. Phone: (82-31) 920-2001. Fax: (82-31) 920-2139. E-mail: ykjang{at}ncc.re.kr. Mailing address for Sang Dai Park: School of Biological Sciences, Seoul National University, Seoul 151-742, Korea. Fax: (82-2) 887-6279. E-mail: sdpark{at}plaza.snu.ac.kr.

Molecular and Cellular Biology, December 2002, p. 8366-8374, Vol. 22, No. 23
0022-538X/02/$04.00+0     DOI: 10.1128/MCB.22.23.8366-8374.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.



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