![]() ![]() It remains unclear what determines the different activities of human SETD2 and yeast Set2. The functional domains of SETD2 are highly conserved in yeast Set2 33, 34. By contrast, Set2, the yeast homolog of SETD2, is the sole enzyme responsible for the mono-, di-, and tri-methylations of H3K36 in yeast 32. SETD2 exclusively catalyzes H3K36 tri-methylation in mammalian cells, whereas mono- and di-methylations of H3K36 are implemented by the methyltransferases NSD1-3 and ASH1L 2, 31. However, it remains poorly understood how the H3.3K36M mutation is recognized by human SETD2 at the nucleosome level and how it affects the physiological functions of human SETD2 in the context of chromatin structure. A recent cryo-EM structure of Chaetomium thermophilum Set2, a fungal homolog of SETD2, bound to a ubiquitinated nucleosome showed that the catalytic domain of Set2 makes extensive interactions with the N-terminus of histone H3 and the C-terminal tail of histone H2A, and stabilizes nucleosome DNA in the unwrapped conformation 30. Recent cryo-electron microscopy (cryo-EM) studies of several histone modifiers, such as human MLL and PRC2 complexes, bound to their natural nucleosome substrates revealed that the modifications of histone tails require specific recognition of nucleosome and even higher-order chromatin structures by the enzymes 27, 28, 29. ![]() Structural and biochemical analysis of the catalytic domain of SETD2 toward an H3.3K36M peptide indicated that the oncohistone peptide occupies the substrate channel of SETD2 accompanied with the side chain of K36M inserted into the catalytic site, and inhibits the HMT activities of SETD2 against mononucleosomes 25, 26. This mutation results in loss of H3K36 methylation landscape and alteration of cancer-related gene expression 24. Moreover, the substitution of lysine 36 with methionine in the histone H3 variant H3.3 (abbreviated as H3.3K36M) has been identified in chondroblastoma and head and neck squamous cell carcinomas 22, 23. Frequent loss or mutations of the SETD2 gene have been observed in clear cell renal cell carcinoma, high-grade gliomas, esophageal squamous cell carcinoma, colorectal cancer, and acute leukemia 17, 18, 19, 20, 21. 8, 9, 10, 11, 12, 13, 14, 15, 16.ĭysfunction of the SETD2–H3K36me3 axis has been linked to a wide range of human malignancies. SETD2 directly associates with the hyperphosphorylated C-terminal domain (CTD) repeats of RNA polymerase II (pol II) through its Set2–Rpb1 interaction (SRI) domain to deposit the H3K36me3 marks co-transcriptionally 4, 5, 6, 7, and participates in the physiological regulation of chromatin condensation, histone exchange, pre-mRNA splicing, DNA damage repair, etc. Tri-methylation of histone H3 on lysine 36 (H3K36me3), primarily deposited by histone methyltransferase (HMT) SETD2 in mammalian cells, occurs at gene bodies of active chromatin and serves as one of the essential histone marks associated with active transcription 2, 3. Histone modifications play pivotal roles in a multitude of cellular processes, including transcription, DNA replication, and DNA damage repair 1. These findings have advanced our understanding of the structural basis for the tumorigenesis mechanism of the H3.3K36M mutation and highlight the effect of nucleosome conformation on the regulation of histone modification. Cryo-EM analysis of yeast H3K36 methyltransferase Set2 complexed with nucleosomes further revealed evolutionarily conserved structural features for nucleosome recognition in eukaryotes, and provides insights into the mechanism of activity regulation. ![]() The linker histone H1, which stabilizes the wrapping of nucleosome DNA at the entry/exit sites, exhibits an inhibitory effect on the activities of SETD2 and displays inversely correlated genome distributions with that of the H3K36me3 marks. By contrast with the stable association of SETD2 with the H3.3K36M nucleosome, the EM densities of SETD2 could not be observed on the wild-type nucleosome surface, suggesting that the association of SETD2 with wild-type nucleosome might be transient. These structural features were also observed in the previous cryo-EM structure of the fungal Set2–nucleosome complex. To investigate how the oncohistone mutation affects the function of SETD2 at the nucleosome level, we determined the cryo-EM structure of human SETD2 associated with an H3.3K36M nucleosome and cofactor S-adenosylmethionine (SAM), and revealed that SETD2 is attached to the N-terminal region of histone H3 and the nucleosome DNA at superhelix location 1, accompanied with the partial unwrapping of nucleosome DNA to expose the SETD2-binding site. Substitution of lysine 36 with methionine in histone H3.3 (H3.3K36M) is an oncogenic mutation that inhibits SETD2-mediated histone H3K36 tri-methylation in tumors. ![]()
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