The modification of newly synthesized histones H3 and H4 by type B histone acetyltransferases has been proposed to play a role in the process of chromatin assembly. The type B histone acetyltransferase Hat1p and specific lysine residues in the histone H3 NH 2 -terminal tail (primarily lysine 14) are redundantly required for telomeric silencing. As many gene products, including other factors involved in chromatin assembly, have been found to participate in both telomeric silencing and DNA damage repair, we tested whether mutations in HAT1 and the histone H3 tail were also sensitive to DNA-damaging agents. Indeed, mutations both in specific lysine residues in the histone H3 tail and in HAT1 resulted in sensitivity to methyl methanesulfonate. The DNA damage sensitivity of the histone H3 and HAT1 mutants was specific for DNA double-strand breaks, as these mutants were sensitive to the induction of an exogenous restriction endonuclease, EcoRI, but not to UV irradiation. While histone H3 mutations had minor effects on nonhomologous end joining, the primary defect in the histone H3 and HAT1 mutants was in the recombinational repair of DNA double-strand breaks. Epistasis analysis indicates that the histone H3 and HAT1 mutants may influence DNA double-strand break repair through Asf1p-dependent chromatin assembly.

Hat1p and Hat2p are the two subunits of a type B histone acetyltransferase from Saccharomyces cerevisiae that acetylates free histone H4 on lysine 12 in vitro. However, the role for these gene products in chromatin function has been unclear, as deletions of the HAT1 and/or HAT2 gene displayed no obvious phenotype. We have now identified a role for Hat1p and Hat2p in telomeric silencing. Telomeric silencing is the transcriptional repression of telomere-proximal genes and is mediated by a special chromatin structure. While there was no change in the level of silencing on a telomeric gene when the HAT1 or HAT2 gene was deleted, a significant silencing defect was observed when hat1⌬ or hat2⌬ was combined with mutations of the histone H3 NH 2 -terminal tail. Specifically, when at least two lysine residues were changed to arginine in the histone H3 tail, a hat1⌬-dependent telomeric silencing defect was observed. The most dramatic effects were seen when one of the two changes was in lysine 14. In further analysis, we found that a single lysine out of the five in the histone H3 tail was sufficient to mediate silencing. However, K14 was the best at preserving silencing, followed by K23 and then K27; K9 and K18 alone were insufficient. Mutational analysis of the histone H4 tail indicated that the role of Hat1p in telomeric silencing was mediated solely through lysine 12. Thus, in contrast to other histone acetyltransferases, Hat1p activity was required for transcriptional repression rather than gene activation.

The Xeroderma Pigmentosum group C (XPC) protein is indispensable to global genomic repair (GGR), a subpathway of nucleotide excision repair (NER), and plays an important role in the initial damage recognition. XPC can be modified by both ubiquitin and SUMO in response to UV irradiation of cells. Here, we show that XPC undergoes degradation upon UV irradiation, and this is independent of protein ubiquitylation. The subunits of DDB-Cul4A E3 ligase differentially regulate UV-induced XPC degradation, e.g DDB2 is required and promotes, whereas DDB1 and Cul4A protect the protein degradation. Mutation of XPC K655 to alanine abolishes both UV-induced XPC modification and degradation. XPC degradation is necessary for recruiting XPG and efficient NER. The overall results provide crucial insights regarding the fate and role of XPC protein in the initiation of excision repair.

Type B histone acetyltransferases are thought to catalyze the acetylation of the NH 2 -terminal tails of newly synthesized histones. Although Hat1p has been implicated in cellular processes, such as telomeric silencing and DNA damage repair, the underlying molecular mechanisms by which it functions remain elusive. In an effort to understand how Hat1p is involved in the process of DNA double-strand break (DSB) repair, we examined whether Hat1p is directly recruited to sites of DNA damage. Following induction of the endonuclease HO, which generates a single DNA DSB at the MAT locus, we found that Hat1p becomes associated with chromatin near the site of DNA damage. The nuclear Hat1p-associated histone chaperone Hif1p is also recruited to an HO-induced DSB with a similar distribution. In addition, while the acetylation of all four histone H4 NH 2 -terminal tail domain lysine residues is increased following DSB formation, only the acetylation of H4 lysine 12, the primary target of Hat1p activity, is dependent on the presence of Hat1p. Kinetic analysis of Hat1p localization indicates that it is recruited after the phosphorylation of histone H2A S129 and concomitant with the recombinational-repair factor Rad52p. Surprisingly, Hat1p is still recruited to chromatin in strains that cannot repair an HO-induced double-strand break. These results indicate that Hat1p plays a direct role in DNA damage repair and is responsible for specific changes in histone modification that occur during the course of recombinational DNA repair.Newly synthesized histones H3 and H4 are rapidly acetylated in the cytoplasm on their NH 2 -terminal tail domains. This is an evolutionarily conserved modification that is removed once the histones are packaged into nucleosomes, suggesting a role for this modification in chromatin assembly (5). The acetylation of newly synthesized histones is thought to be catalyzed by type B histone acetyltransferases (9). The lone type B histone acetyltransferase identified to date is Hat1p (22, 30). When isolated from a variety of eukaryotic organisms, Hat1p is found in a complex with a WD40 repeat-containing regulatory subunit (Hat2p in Saccharomyces cerevisiae or Rbap46 in mammalian cells) (20,25,36,50). Consistent with its classification as a type B histone acetyltransferase, Hat1p is highly specific for histones that are free in solution with no demonstrable activity when chromatin-associated histones are used as a substrate. In addition, the enzyme is specific for histone H4 lysine residues 5 and 12, which are the sites of acetylation found on the newly synthesized pool of histone H4 in a wide variety of eukaryotic organisms (11,13,20,22,30,34,42,50). Hence, Hat1p has the requisite biochemical activities to function in the acetylation of newly synthesized histone H4. However, evidence directly linking Hat1p to the acetylation of histones in vivo is lacking (32).The acetylation of newly synthesized histones by type B histone acetyltransferases is presumed to play a role in histone deposition. Circumstantial evid...

The p38 MAPK is a family of serine/threonine protein kinases that play important roles in cellular responses to external stress signals, e.g. UV irradiation. To assess the role of p38 MAPK pathway in nucleotide excision repair (NER), the most versatile DNA repair pathway, we determined the efficiency of NER in cells treated with p38 MAPK inhibitor SB203580 and found that p38 MAPK is required for the prompt repair of UV-induced DNA damage CPD. We further investigated the possible mechanism through which p38 MAPK regulates NER and found that p38 MAPK mediates UV-induced histone H3 acetylation and chromatin relaxation. Moreover, p38 MAPK also regulates UVinduced DDB2 ubiquitylation and degradation via phosphorylation of the target protein. Finally, our results showed that p38 MAPK is required for the recruitment of NER factors XPC and TFIIH to UV-induced DNA damage sites. We conclude that p38 MAPK regulates chromatin remodeling as well as DDB2 degradation for facilitating NER factor assembly.

The new compounds, epoxydines A and B (1 and 2, resp.), along with the known and related metabolites, 3–6, were isolated from the fungal endophyte Phoma sp. The structures of the new compounds were elucidated by detailed spectroscopic analysis, and the relative configuration of 1 was confirmed by ROESY experiments. Preliminary studies indicated that compounds 2–5 possess good antibacterial, antifungal, and algicidal properties. Similarly, compound 1 showed antifungal and algicidal, and compound 6 antibacterial and algicidal properties.

The new bicyclic fusidilactones D (1) and E (2), along with the known and related fusidilactone B (3), have been isolated from the fungal endophyte Fusidium sp. The structures of the new compounds were elucidated by detailed spectroscopic analysis and the relative configurations were confirmed by an X-ray diffraction study of a single crystal of 3. The abso-

In this study, a cDNA encoding a novel acylCoA:diacylglycerol acyltransferase (DGAT)-like protein is identified and isolated from the diatom microalga Phaeodactylum tricornutum (PtDGAT3). Analysis of the sequence reveals that ptDGAT3 cDNA encodes a protein of 504 amino acids with a molecular mass of 64.5 KDa. The putative ptDGAT3 protein has two catalytic domains: a wax ester synthase-like acyl-CoA acyltransferase domain and a bacteria-specific acyltransferase domain, which shows higher similarity to the DGAT3 of Acinetobacter calcoaceticus than reported DGAT1 or DGAT2 from high plants or algae. Its activity was confirmed by heterologous expression of PtDGAT3 in a neutral lipid-deficient quadruple mutant yeast Saccharomyces cerevisiae H1246. The recombinant yeast restored the formation of a lipid body and displayed a preference to the incorporation of unsaturated C 18 fatty acids into triacyglycerol (TAG). This is the first characterized algal DGAT3 gene, giving further evidence to the occurrence of a DGAT3-mediated TAG biosynthesis pathway.