histone tales
by Leslie H. Lang

epetition breeds familiarity. In only a few short years, images of the spiraling and twisted DNA double helix have become firmly established as the iconic darling for the genome sciences, much like the candy-striped red and white pole had been to men's barbershops everywhere.

Yes, one might say the double helix does contain an almost kitschy purity, evocative of a landmark scientific achievement few of us fully comprehend. Still, the image of the double helix remains only a representation of the molecule. "It would surprise a lot of people to know that DNA is not just two linear strands. It's wrapped around histone proteins to form a highly folded complex called chromatin," says Brian Strahl, assistant professor of biochemistry and biophysics. This complex of nucleic acids and proteins binds DNA into higher-order structures, ultimately forming a chromosome.

The core histones appear in all organisms that have nucleated cells, including yeast and mammals. Four core histone proteins (H2A, H2B, H3, H4) each contain a "head," or globular domain, and an amino "tail." Of interest to Strahl is that these histones, specifically processes that modify them, are thought to play a major role in controlling gene expression and cell division.

nother image. Think of chromatin's structure as a telephone cord with a bead between each coil. Each bead represents a nucleosome — chromatin's fundamental repeating unit consisting of DNA wrapped twice around the four histone "core" proteins, their tails wagging and sometimes touching outside the nucleosome. Meanwhile, a fifth histone (H1) serves as a "linker histone" between nucleosomes. Now fold the cord on itself again and again.

This image approximates the scientifically known. Stretches of nucleosomes are folded upon themselves to create higher-order chromatin structures, albeit still not well defined. Although the chromatin packaging allows efficient storage of genetic information (the length of the entire complement of 46 chromosomes in a human cell is about one meter), it also impedes a wide range of cell processes, including access to DNA by transcription factors — the proteins that regulate gene expression. In other words, DNA must become unblocked to allow its information to be read and to produce messenger RNA (mRNA), which in turn must exit the nucleus and become translated into a protein product.

How that might occur — how DNA becomes more accessible to transcription factors — is currently an area of intense research scrutiny, including at UNC-Chapel Hill.

.: Yi Zhang and Brian Strahl: Imagine chromatin's structure as a telephone cord with a bead between each coil. Each bead represents a nucleosome  —  chromatin's fundamental repeating unit. Photo by Steve Exum; click to enlarge. :.

Strahl, Yi Zhang, assistant professor of biochemistry at the Lineberger Com-prehensive Cancer Center, and others have modified the older view of many scientists that histones play a passive role in chromosomal architecture, a view of histones as primarily structural, packaging DNA into chromatin fibers while having little to do with gene regulation.

Independently — Strahl working with yeast cells and Zhang with mammalian cells — the two are discovering that histones play a more dynamic role in chromatin, namely, its loosening or tightening.

The researchers' attention is focused on histone methylation, the addition of a methyl group to lysine, one of the amino acids that comprise the tail region of histone molecules.

"We've known for three decades that histones can be methylated, but nobody knew the identity of any of the enzymes responsible for this methylation until two years ago," Zhang says. That was when the first such enzyme was identified which specifically methylates histone H3 at lysine 9. Its presence there was linked to chromosome areas of gene silencing or inactivation.

hang's lab has since identified the enzyme SET7, which specifically modifies lysine 4 on the histone H3 tail. This modification makes the chromatin structure more open so other proteins can access particular genes, Zhang says. Moreover, methylation of the same histone at lysine 4 and lysine 9 have opposite effects. Thus, according to Zhang, methylation at either site could determine either gene activation or gene silencing. Still, the situation is probably more complex than that. Among the possibilities, SET7 could have functioning partners yet unidentified, Zhang says. He recently reported discovering another two enzymes, and his lab is intensely studying their functions.

For his recent entry into histone modification, Strahl and former colleagues at the University of Virginia identified and characterized Set2, a novel histone that is responsible for methylating lysine 36 on the H3 tail. However, this modification helps to repress or silence gene transcription. Thus, Set2 might be "a coregulator of transcription" in the sense that it turns genes "off" instead of "on," as in the case of SET7.

"During development, you have different sets of genes that are important for, say, limb formation, and when the limbs are completed, the genes responsible for them must be turned off," Strahl says.

It may well be that methylation and other modifications are part of an emerging "histone code" of modifications that ultimately regulate gene expression. Strahl and his former mentor at the University of Virginia, David Allis, postulated such a code in a 2000 paper in the journal Nature. This code would be in addition to the now familiar genetic code of repeating As, Cs, Gs, and Ts of DNA nucleotide sequences. Through this histone code, differentially modified histone proteins could organize the genome into stretches of active and silent regions. Moreover, these regions would be inherited during cell division.

"We believe that methylation and other modifications that affect histone proteins, including acetylation and phosphorylation, are all dynamically involved and play critical roles in gene activation and deactivation at the appropriate times," Strahl says.

This process, he explained, possibly could work by the ability of these modifications to bring in additional proteins that result in opening or closing of the chromatin molecule.

"Yi and I, as well as other labs, are at the frontier of understanding about the enzymes that are so important for the dynamic regulation of chromatin," Strahl says. "Our findings add to our knowledge of a basic and very important process in human biology. They could offer new insight as to why certain genes in cancer are inappropriately expressed and how that might be corrected."

eanwhile, in Bill Marzluff's sprawling and bustling Fordham Hall laboratory, the histone focus is even more basic than trying to tease out chromatin dynamics. "We work on the histone messenger RNA level — DNA's blueprint for histone proteins — its regulation, processing, transport, and degradation," graduate student Judy Erkmann says. Her work explores how histone mRNA is transported from the cell nucleus to the cytoplasm.

.: Bill Marzluff's lab discovered the stem-loop binding protein of histone mRNA. Photo by Steve Exum; click to enlarge. :.

The focus on histone mRNA stems largely from the Marzluff team's 1996 discovery and cloning of the stem-loop binding protein, SLBP. This unique protein is a major regulatory player in histone mRNA. It latches onto the looped tail of histone mRNA and signals the synthesis of histone proteins crucial to cell functioning during embryogenesis and throughout the organism's life.

But SLBP does more than simply hitch a ride on a loop of nucleotides. It also doggedly performs a string of important duties after it takes the mRNA into a specific region of the nucleus. It interacts with other proteins to make sure the mRNA is properly processed into its final form. And then after helping get it out to the cytoplasm — the cell's factory floor — SLBP remains bound to histone mRNA, making sure that its instructions are properly translated.

"A lot of people work on histones, but just a few groups in the world actually work on understanding how histone mRNA is synthesized and regulated," says research assistant professor Zbigniew Dominski, whose own work focuses on how mRNA is processed and matures into a translatable message. "In this lab we cover all the different steps of histone mRNA message metabolism."

"And that's very important because the synthesis of histone proteins depends strictly on metabolism of the mRNA," says Ricardo Sanchez, the lab's newest Ph.D. recipient. Histone mRNA translation and regulation remain his major research interest.

And what if the metabolic regulatory process goes awry? Recent findings in Drosophila — fruit flies — by Robert Duronio, associate professor of biology, in collaboration with Marzluff, highlight the possibilities at the edge of life's beginnings. Mutated or non-functioning SLBP is associated with failure to develop beyond the early embryo. A similar outcome may well apply to other multicellular creatures, including us.

Les Lang is assistant director of public affairs for science communications in the UNC-Chapel Hill school of medicine.