New & Noteworthy

Acetylation Regulates the Nuclear Pore Complex

July 15, 2022

Several recent studies have done an excellent job characterizing the architecture of the yeast nuclear pore complex (NPC). With so much new information, researchers are now able to ask probing questions about how NPCs mediate communication between the nucleus and the rest of the cell. Considering that signals perceived from the environment need to reach the transcriptional machinery in the nucleus, and that mRNA transcripts made in response to these signals need to get back out to get translated, the NPC has a lot of communicating to do. A study in a recent issue of the EMBO Journal by Gomar-Alba et al. makes strong strides toward understanding how this communication is accomplished.

On the nuclear side of the NPC resides a substructure called the nuclear basket that has previously been shown to play roles in regulating gene expression and mRNA export. The nuclear basket also interacts with lysine acetyltransferases (KATs) and deacetylases (KDACs) that are best known for modulating transcription via reversible acetylation of histones in chromatin. These enzymes, however, can also act on non-histone proteins and have been linked to numerous cell processes, including DNA damage repair, cell division, and signal transduction.

From Gomar-Alba et al, 2022

Promotion of mRNA export is another function linked to acetylation, specifically by the NuA4 histone acetyltransferase complex, for which the catalytic subunit is Esa1p. Gomar-Alba et al. show in this recent study that Esa1p is the primary lysine acetyltransferase that promotes cell cycle entry—and also that it acetylates the nuclear pore protein Nup60p.

Acetylation of Nup60p promotes mRNA export, which in turn triggers fast entry into the Start phase of the cell cycle, thereby promoting cell division. Nup60p accomplishes this increased export by recruiting the TREX-2 transcription-export complex to the nuclear basket once Nup60p becomes acetylated. The deacetylated form of Nup60p has lower affinity for TREX-2 and thus mRNA export decreases. Deacetylation of Nup60p is performed by Hos3p, which acts in opposition to Esa1p in removing Esa1p-transferred acetyl residues.

Perhaps the most intriguing finding in this study is that Hos3p localizes primarily to daughter cells after cell division, causing displacement of the mRNA export complex and thus slowing G1/S phase transition. This action prevents premature division in the smaller daughter cells, as they require additional growth to meet the size control threshold for entry into a new cell cycle. Accomplishing this level of control with a single enzyme acting on a single nuclear pore protein is a simple, elegant solution.

As usual, studies in yeast make enormous impact on understanding cell division in other organisms.

Categories: Research Spotlight

Tags: cell cycle control, nuclear export, mRNA export, Saccharomyces cerevisiae, nuclear pore, acetylation

Strung Out Chromosomes and Coyotes

January 29, 2018


As Wile E. Coyote knows, when setting a bear trap, you need to make sure the tension is just right. As you can see in this video, if there isn’t enough, then the Road Runner gets away:

And of course, Wile E. ends up getting caught in the trap!

Like our super-genius coyote friend, Saccharomyces cerevisiae cells need to make sure there is the right amount of tension sometimes, too. One of those times, for example, is tension on their chromosomes at the metaphase stage of cell division.

If the chromosomes aren’t attached to the spindle in just the right way, the cell will not let mitosis move from metaphase to anaphase. The cell won’t put down the birdseed until it “knows” the trap is set properly.

While Wile E. Coyote probes the tension by eyeballing it, yeast does it in a much more thorough way. Two key pieces of the sensing machinery in yeast are the tension sensing motif (TSM) from histone H3, 42KPGT, and the Shugoshin 1 protein, Sgo1p.

The current model is that the TSM of histone H3 acts as an anchoring point or a landing pad for Sgo1p, the protein responsible for sensing that there is the right amount of tension between chromosomes and the spindle/kinetochore. When the TSM is mutated with, for example, a G44S allele, the idea is that Sgo1p can no longer bind causing chromosomes to sometimes segregate incorrectly.

Previous work had shown that mutating Gcn5p can partially rescue the effects of the G44S allele. In a new study in GENETICS, Buehl and coworkers provide evidence that the reason it can has to do with the tail of histone H3. Basically, this tail can act as a secondary anchor if the TSM, the first anchor point, is mutated. But the tail can only work if it is not acetylated by Gcn5p.

CoyoteRoadrunner

Bad things can happen when plans go astray for Wile E. Coyote and for chromosome segregation in yeast. (Wikimedia)

It is as if Wile E. Coyote’s bear trap is broken so he has to fashion a new trigger to get it to snap. When the TSM of H3 is gone, yeast can fashion a new sensor by eliminating the acetylation on the tail of the H3 histone.

Buehl and coworkers focused on the tail because it is a known target of Gcn5p acetylation. In particular, there are four lysine residues in the tail—K9, K14, K18, and K23—that are acetylated by Gcn5p to varying degrees. The most acetylated is K14 followed by K23 and K18. K9 is hardly acetylated at all.

When they deleted the histone tail, they found that the effects of the TSM mutation, G44S, were more severe. But when they mutated the four lysines all at once, the effects of G44S weren’t as bad. This is consistent with the unacetylated tail serving as a landing pad for Sgo1p in the absence of the TSM. No tail, no binding, leading to chromosome mis-segregation.

As might be expected if this was linked to Gcn5p, each individual lysine mutant had effects that corresponded to their level of acetylation by Gcn5p. So K14A did the best job of partially rescuing the G44S allele, K18A and K23A did an okay job and K9A had very little effect.

Buehl and coworkers provide three additional sets of experiments to show that G44S causes chromosome mis-segregation and that K14A can partially rescue it. There is only time to discuss one of these, but you can read the paper for the other two.

This strategy uses the idea that diploids cannot mate because of a repression function on each copy of chromosome III (chrIII). If a diploid loses one of its chrIII’s, then it will be able to mate.

As expected, the authors found that both wild type and the K14A mutant had very few diploids that could mate. But it was a different story for the G44S mutant—it had many more diploids able to mate, presumably due to the loss of one chrIII. The double mutant K14A/G44S had more diploids mate than wild type but less that G44S on its own.

This is what we would expect if K14A can only affect tension sensing in a TSM mutant. It would do little on its own but could partially rescue the TSM mutant G44S.

ROADrunner

If Wile E. Coyote had a backup plan like yeast does for chromosome segregation, he might get to dine on the Road Runner. (Rosenfeld Media)

The authors next used chromatin immunoprecipitation (ChIP) assays to directly show that the K14A mutation in histone H3 allowed Sgo1p to bind when the TSM was mutated. There was one hitch though—Sgo1p bound most everywhere and not just at the pericentromeres where it usually does.

In a final set of experiments, the authors used GST-pulldowns to show a direct interaction between the H3 tail and Sgo1p. To their surprise, they found that acetylating the tail did not affect Sgo1p binding. They suggest that perhaps the acetylated tail interacts with something that blocks Sgo1p from binding.

Yeast has a backup just in case its main way of sensing tension between the spindle/kinetochore and chromosome, the TSM of histone H3, breaks down. In that case, it can use the tail of histone H3, assuming it is not acetylated.

If only Wile E. Coyote included backups like this in his crazy plans! Then he could finally dine on that tasty Road Runner he is always chasing.

by Barry Starr, Ph.D., Director of Outreach Activities, Stanford Genetics

Categories: Research Spotlight

Tags: Histone H3, chromosome segregation, metaphase, acetylation

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