October 20, 2017
Imagine that Jane is in trouble on the edge of the jungle. She needs to be saved soon or she will be sent back to Europe (which she does not want).
Tarzan knows he can’t get there in time by running along the jungle floor. But of course he has a trick up his sleeve—swinging from vine to vine!
He gets there in time, fends off her would-be kidnappers, and saves the day. All because he transferred from one strand of vine to another instead of “sliding” along the jungle floor.
A new study by Wollman and coworkers shows that at least two transcription factors in Saccharomyces cerevisiae, Mig1p and Msn2p, seem to share a lot in common with Tarzan. In a process termed intersegment transfer, they get to where they need to in the genome by “swinging” from DNA segment to DNA segment instead of just sliding along the DNA.
And transcription factors like these need to get to the right place in time to save the cell from outside threats. Just like Tarzan had to swing from vine to vine to save Jane.
This sort of approach would not necessarily work well with just a single transcription factor with a single DNA binding domain. It would sort of be like a one-armed Tarzan—it is hard to take advantage of swinging from vine to vine without at least two arms!
The authors argue that Mig1p gains its “extra arms” through joining together into a cluster. Now each Mig1p can bind DNA and drag the other transcription factors with them. A neat solution to the one arm problem.
Their basic approach is to use fluorescence microscopy to follow a GFP-Mig1p fusion protein in a single cell, something that has only become possible recently. What they found was that there were two populations of transcription factors—a diffuse set of smaller molecules and distinct, larger clusters made up of multiple Mig1p’s.
The clusters appeared to be the ones doing the work in the nucleus. Kinetic studies showed that they stayed in one place in the nucleus for over 100 seconds which is consistent with the clusters and not the monomers being bound to the DNA.
OK so this transcription factor tends to clump up into clusters and it looks like these clusters are the ones regulating gene expression. They also showed that a second transcription factor, Msn2p, did the same thing.
The authors next set out to see if this approach made sense for genetic regulation by running simulations of Mig1p finding its sites in the nucleus as either monomers or as clusters. It made sense to form clusters.
A lot of previous work has been done in S. cerevisiae in terms of the three dimensional map of the genome and where Mig1p DNA binding sites were located in this mesh of DNA. And in the course of their studies, Wollman and coworkers were able to estimate how many Mig1p molecules were in yeast cells and how many were in clusters.
They now had all the information they needed to run their simulations. When they crunched the numbers, they found that clusters fit their data much better than monomers (R2 = 0.75 vs. R2 < 0).
The final step was to work out what part of Mig1p was involved in forming the clusters. To do this, the authors compared Mig1p and Msn2p, the second transcription factor they studied that also formed into clusters, and looked for structural regions they might have in common.
What they found was both proteins had a highly disordered region. For Mig1p, it was at the C-terminus and for Msn2p it was at the N-terminus.
The hypothesis is that these disordered regions, which are both at the opposite end of the protein from the DNA binding domain, interact and form ordered structures that enable clusters to form. Wollman and coworkers used circular dichroism to show that when Mig1p was put in conditions that favor cluster formation, there was a transition consistent with unstructured protein becoming structured.
What we seem to have is a cluster of transcription factors connected in the middle with their DNA binding domains pointing out. This rolling cluster can more easily hop from DNA strand to DNA strand to find the right spots to bind.
Without this mechanism, Mig1p couldn’t get to where it needs to in time. It is as if Tarzan had 6-9 arms circling his body so he could get to Jane even more quickly.
With the wide range of tools available and our deep understanding of how yeast works and how a yeast cell is organized, our trusted ally S. cerevisiae again teaches us something fundamental about how our biology works.
Mig1p may be able to swing like Tarzan but it can’t yell like him. Or like Carol Burnett!
by Barry Starr, Ph.D., Director of Outreach Activities, Stanford Department of Genetics
Categories: Research Spotlight