Characterizing the Structure of Tos4 as a Suppressor of a Budding Yeast Oncohistone Model Restricted; Files Only
Zhang, Allen (Spring 2026)
Abstract
In eukaryotes, histone proteins are essential for many homeostatic processes. Histones not only wrap and condense DNA into chromosomes for cell division but also undergo post-translational modifications to regulate gene expression. However, missense mutations that alter a single amino acid residue can convert histones into “oncohistones” that drive carcinogenesis through altered gene expression and unregulated cell division. To study oncohistones, budding yeast can be employed as a powerful system because of the simple yet highly conserved nature of the histone proteins. We leveraged budding yeast to model pathogenic missense mutations that convert histones into oncohistones to explore how these changes alter histone function. Our studies that model a histone H3 oncohistone H3K36M revealed that these oncohistone model mutant cells are sensitive to diverse cellular stressors. We exploited this growth sensitivity to perform a high copy suppressor screen with the goal of eventually uncovering carcinogenic mechanisms. One of the suppressor genes identified is TOS4, which suppresses the caffeine-sensitive growth of H3K36R/M mutant cells. The Tos4 protein interacts with the histone deacetylase (HDAC) complex Rpd3L via the Tos4 Forkhead-associated (FHA) domain to regulate gene expression under replication stress, but other aspects of Tos4 function are poorly characterized. To identify functionally important domains of Tos4, our study employed AlphaFold 3 to define additional folded domains in Tos4. We then used these structural predictions to design several truncation mutants to define domains that are necessary for the suppression of the H3K36 oncohistone model. In addition, we further explored the function of Tos4 in replication stress by taking advantage of hydroxyurea-sensitive yeast cells that lack Tos4 and Dun1, a checkpoint kinase that is critical for maintaining nucleotide levels under replication stress. Using this assay, we identified domains of Tos4 that are required for Tos4 function in promoting cell growth under replication stress. We found that Domain B, but not Domain A, contributes to both the DNA damage response (DDR) and suppression of H3K36R/M caffeine-sensitive growth. The results of this study may provide greater insight into how H3K36 mutants and other oncohistones drive carcinogenesis and identify key pathways that could be targeted for novel therapies.
Table of Contents
INTRODUCTION..............................................................................................................................1
MATERIALS AND METHODS............................................................................................................ 9
RESULTS....................................................................................................................................... 12
DISCUSSION................................................................................................................................. 19
REFERENCES................................................................................................................................ 22
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File download under embargo until 28 May 2028 | 2026-04-06 21:00:11 -0400 | File download under embargo until 28 May 2028 |
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