Abstract 1152 Quantitative Assessment of Antisense mRNA Expression in a Budding Yeast Oncohistone Model

Histone proteins are essential to package chromatin within the cell nucleus as well as to regulate gene expression. Post-translational modifications (PTMs) on histones, such as methylation of lysine residues, regulates histone functions. Critically, missense mutations in histone genes can convert histones into oncohistones, such as H3K36M, that alter histone PTMs, dyregulate histone function, and drive cancer phenotypes. One potential way that oncohistones can dysregulate gene expression is through altered chromatin structure. Previous studies provide evidence that altered histones, such as H3K36 mutants, can lead to cryptic transcription and increased production of antisense RNAs. Based on our prior observations and the established relationship between chromatin structure and gene expression, we tested the hypothesis that missense mutations in the histone H3 gene, particularly H3K36R and H3K36M, could cause an increase in antisense RNAs. To test this hypothesis, we employed a budding yeast oncohistone model. Budding yeast contains two genes that encode histone H3, HHT1 and HHT2. In this oncohistone model, the HHT2 gene is edited and the HHT1 gene is deleted to produce cells that express the histone variant as the sole histone H3 protein in the cells. Total RNA from these yeast cells was isolated, converted to cDNA via reverse transcriptase, and quantified using qPCR. Specific antisense RNAs previously identified to be altered in yeast H3K36 mutant cells were measured by qPCR. Preliminary results revealed an increase in antisense RNA in H3K36R and H3K36M mutant cells relative to wildtype control, in support of our hypothesis. This study enhances our molecular understanding of oncohistone mutations and offers novel insights into how histone changes may contribute to dysregulation of gene expression. These observations can be extended to human cells to explore the implications of these results in oncohistone-driven cancers. This research is supported and funded by 1)Department of Biology, Emory College of Arts and Sciences, and 2) Department of Radiation Oncology, Emory University School of Medicine, Atlanta, GA 30322.