Histone modifications: Now summoning sumoylation.

H istones can be modified in many ways to affect gene expression, including acetylation deacetylation (1), phosphorylation (2), methylation (3), and ubiquitylation (4). Now, in this issue of PNAS, Shiio and Eisenman (5) report that sumoylation is yet another histone modification, and, interestingly, it may regulate transcriptional repression. Although there may appear to be a bewildering array of histone modifications involved with gene regulation, there could be a fairly simple paradigm underlying them. Eukaryotic DNA is packaged within the nucleus through its association with histone proteins (H2A, H2B, H3, and H4), forming the fundamental repeating unit of chromatin, the nucleosome. The precise architecture of chromatin dictates whether it is permissive or resistant to transcription and other DNA-templated processes, such as replication, DNA repair, and recombination. Hence, it is not surprising that mechanisms that promote changes in chromatin structure are central to transcriptional regulation. One key pathway to alter chromatin structure involves covalent modifications of the histone tails. Small ubiquitin-related modifier (SUMO) shares 18% identity with ubiquitin and adopts a similar 3D structure (6). Ubiquitylation has a role in protein degradation, whereas SUMO does not. The size of SUMO and ubiquitin (11 and 9 kDa, respectively) clearly distinguishes them from the other known posttranslation modifications of histones, which are all small chemical groups. SUMO and its ATP-dependent pathway of conjugation to substrates are conserved in all eukaryotes investigated, yeast through humans. One important consideration regarding sumoylation of histones is that its target residue, lysine, is a putative substrate for multiple modifying enzymes (Table 1). Acetylation of lysines is dynamically opposed by deacetylating enzymes. Typically, histone acetylation is activating and histone deacetylation is repressing, and, in a symmetrical fashion, DNA-bound activators and repressors recruit these enzymes to target genes (1). By contrast, lysine methylation on histones appears to be quite stable (no demethylases have been reported despite considerable effort to identify them) and may, in fact, provide a ‘‘memory’’ mark (3). Consistent with its increased stability, lysine methylation can, depending on context, contribute a ‘‘permanent’’ mark of either open, active euchromatin or closed, repressed heterochromatin (7). Perhaps, then, it is not surprising that acetylation or methylation can occur on the same lysine residues to allow dynamic regulation by acetylation or more stable regulation by methylation. The current study by Shiio and Eisenman (5) on histone sumoylation and recent reports on histone ubiquitylation (4, 8–10) bring up many questions of how these much larger polypeptides function within chromatin; i.e., are they activating or repressing, are they dynamic or static, do they occupy the same lysines, and, finally, do they oppose one another? Shiio and Eisenman provide some provocative observations and some initial answers to these questions. SUMO has been identified bound to many proteins, but to understand histone sumoylation, it may be most informative to examine the known effects of sumoylation on DNA-binding transcription factors. Sumoylation has been shown to stimulate activity; notable examples include heat shock factor HSF1 and tumor suppressor p53 (11–13). However, sumoylation most frequently correlates with decreased transcriptional activity (e.g., Elk1, Sp3, c-Myb, and cJun) (11) and, thus, repression of target genes. Interestingly, Shiio and Eisenman’s study indicates that sumoylation of histone H4 also correlates with transcriptional repression, at least within an artificial transfected reporter model. First, they establish that mammalian histone H4 is sumoylated both in vivo and in vitro, and this modification appears to be more efficient than sumolylation of the other core histones. They show that expression of a reporter is decreased by targeting UBC9, which conjugates SUMO to its substrate, to the reporter. Moreover, the targeted UBC9 results in reduced levels of promoter-associated acetylated histone H3 and marked elevation in heterochromatic protein 1 (HP1). Notably, HP1 is a key structural protein of heterochromatin and binds to methylated Lys-9 on histone H3. HP1 can also contribute to repression of individual euchromatic genes (14). The amino-terminal tail of histone H4 contains five lysines, all of which may be candidates for sumoylation. However, none of the histone proteins, including histone H4, contain the putative consensus sequence for sumoylation ( -Lys-XGlu, where is a large hydrophic residue and X is any amino acid). In addition, as described above, many of these lysine residues may undergo other modifications (Table 1). Consequently, it is exceedingly difficult to study the physiological significance of sumoylation. One solution to this problem, used by Shiio and Eisenman as well as by others studying sumoylation and ubiquitylation of nonhistone substrates, is to genetically fuse SUMO to the putative target protein. Thus, Shiio and Eisenman observe that SUMO-H4 associates with chromatin and can be coimmunoprecipitated with endogenous histone deacetylase 1 (HDAC1) and HP1. These data provide strong, but indirect, evidence that sumoylation of H4 associates with agents mediating gene repression. Possibly analogous to this is a recent study proposing that p300 CBP can repress transcription by recruiting HDAC6 by way of SUMO (15).