Histone variants: new versions of ancient genes

From the Malik Lab, Basic Sciences Division

There is nothing permanent except change. -Heraclitus

There is perhaps no more fundamental property of life than change, both with respect to personal experience and, as the basis of evolution, to the nature of biology itself. The constant genetic permutations that fuel the evolutionary process have given rise to an incredible complexity and diversity of life, but even in the face of this unstoppable force, some aspects of life are so foundational that they remain essentially immutable. The inevitability of death. The fact of reproduction. The chemical nature of the genetic code. Unsurprisingly, some of the genes associated with these phenomena are themselves quite resistant to evolutionary change. Take, for instance, the histones. The most ancient of these proteins, which form a scaffold around which our DNA is wrapped, arose billions of years ago and have changed little since. But some histone genes, in an apparent bucking of this trend, have evolved much more recently and, in some cases, are still evolving rapidly. In a new article in Molecular Biology and Evolution, postdoctoral fellow Dr. Pravrutha Raman and Dr. Harmit Malik, a Professor in Fred Hutch’s Basic Sciences division and connoisseur of genes that are evolving more rapidly than they seem to have any business to, in collaboration with Fred Hutch’s Dr. Toshi Tsukiyama and Dr. Antoine Molaro of the Genetics, Reproduction and Development Institute, identified several new and rapidly evolving histone genes in mammals.

Histone genes can be broadly categorized into core and variant genes. The ancient and extremely conserved core histones (H2A, H2B, H3, and H4) “package genomes after DNA replication”, write the authors. “In contrast, variant histones promote specialized chromatin functions, including DNA repair, genome stability, and epigenetic inheritance.” While a number of variant histone proteins have been previously identified, the group noted that very few variants of the H2B protein have been identified in animals. This suggested that our understanding of the histone repertoire in animals may be incomplete. They closely examined 18 mammalian genomes to identify regions with homology to known histone variants. “We were able to obtain a near-comprehensive list of all variant H2B open reading frames”, they explained, which contained considerable diversity in sequence and length. By performing phylogenetic analysis to understand the evolutionary relationships between these sequences, the found seven distinct classes of H2B variants (Figure 1), including five that were broadly present in mammals.

The authors next compared the H2B variants to the core H2B protein in order to understand what distinguishes these proteins. They found a range of differences among the proteins that could affect histone function, including those with potential impacts on DNA binding or packing, protein posttranslational modifications, protein-protein interactions, or even those that could promote function outside of the nucleus. Thus, it appears that each variant has evolved a distinct function within the body.

Next, the group examined the evolutionary dynamics of each variant to determine the extent to which they have diversified or remained stable over evolutionary time. They observed high rates of purifying selection in all genes, suggesting that they likely have important biological roles and are therefore somewhat constrained in what sequence changes can be tolerated by the animal, but they also found clear evidence of evolutionary change. All H2B variants, for example, appeared to be evolving more rapidly than the core H2B protein, some of them much more rapidly. Likewise, they observed that many of the variants had been lost or duplicated in particular animal lineages. “Overall, our findings of strong purifying selection suggests that H2B variants perform vital functions leading to their overall retention, whereas our findings of positive selection suggest that they have been subject to recurrent genetic innovation”, the authors summarized.

Finally, as a step towards understanding the function of these variants, the authors examined tissue-specific RNA sequencing data to determine when and where in the body they are expressed. They found that most of the variants are expressed in the testes, which is known to be a common site of histone variant expression. However, two of the variants were primarily expressed in ovaries and early embryos, an “unusual expression site for histone variants”, according to Dr. Raman, suggestive of the tantalizing probability that they have evolved new functions in accordance with their new expression patterns.

Dr. Raman, reflecting on the significance of this work, says “Although histones are typically highly conserved and slowly evolving, we discover mammalian H2B variants that have highly divergent sequences and display high evolutionary turnover with gains and losses across species…our work reveals that even repertoires of fundamental proteins like histones can be continually altered by biological forces in the germline.” Looking forward, she notes that the major missing piece is an understanding of what these histone variants are doing. “Most mammalian H2B variants, and especially our newly discovered H2B variants have not been functionally characterized. We plan to use human cells to decipher the molecular properties of these histones. Furthermore, germline-histone variants including some H2B variants can be aberrantly expressed in cancer cells. We are interested in the causes and consequences of this aberrant expression on human cancers.” Dr. Raman was also keen to point out that two additional members of the Malik Lab, undergraduate Callie Rominger and staff scientist Dr. Janet Young, made significant contributions to this work.

histone phylogeny
Phylogeny of the mammalian core and variant histone H2B proteins. Image provided by Dr. Pravrutha Raman

This work was supported by the Foundation pour la Recherche Médicale, the National Institutes of Health, and the Howard Hughes Medical Institute.

Fred Hutch/UW Cancer Consortium members Harmit Malik and Toshio Tsukiyama contributed to this work.

Raman P, Rominger MC, Young JM, Molaro A, Tsukiyama T, Malik HS. 2022. Novel Classes and Evolutionary Turnover of Histone H2B Variants in the Mammalian Germline. Mol Biol Evol. 39(2):msac019.