Abstract
In S. cerevisiae, the lysine methyltransferase Set1 is a member of the multiprotein complex COMPASS. Set1 catalyzes mono-, di- and trimethylation of the fourth residue, lysine 4, of histone H3 using methyl groups from S-adenosylmethionine, and requires a subset of COMPASS proteins for this activity. The methylation activity of COMPASS regulates gene expression and chromosome segregation in vivo. To improve understanding of the catalytic mechanism of Set1, single amino acid substitutions were made within the SET domain. These Set1 mutants were evaluated in vivo by determining the levels of K4-methylated H3, assaying the strength of gene silencing at the rDNA and using a genetic assessment of kinetochore function as a proxy for defects in Dam1 methylation. The findings indicate that no single conserved active site base is required for H3K4 methylation by Set1. Instead, our data suggest that a number of aromatic residues in the SET domain contribute to the formation of an active site that facilitates substrate binding and dictates product specificity. Further, the results suggest that the attributes of Set1 required for trimethylation of histone H3 are those required for Pol II gene silencing at the rDNA and kinetochore function.
Highlights
Eukaryotic DNA is assembled into higher-order chromatin structures that promote compaction and protection of DNA
The Set1 sequence was aligned with other KMTases to identify the four conserved SET domain motifs and the locations of conserved amino acids that may play an important role in protein methylation
The results show that the steady-state level of Set1 protein was similar in protein extracts from wild-type cells (SET1+ and SET1+::ura3-52) and each of the sixteen amino acid substitution mutants
Summary
Eukaryotic DNA is assembled into higher-order chromatin structures that promote compaction and protection of DNA. The structure of chromatin is dynamic to provide access to the underlying DNA template for nuclear processes, such as transcription and replication, and is controlled by several mechanisms [1]. The mechanisms of chromatin regulation by methylated histones are not as well understood as those governed by acetylated histones, a large body of work supports roles for methylated histones in the regulation of euchromatin and heterochromatin [2,3,4,5]. Histone methylation can be more complex than other covalent modifications because multiple methyl groups can be present at the same lysine residue that may alter the function of chromatin in different ways [2,7]. Regulatory proteins can discriminate the different methylated forms of a histone, providing means to increase the types of signals presented by chromatin [8]
Sign-in/Register to view highlights & summary Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
