New & Noteworthy
Polygamous DNA Replication
June 26, 2014
Regulation ensures that most replication origins fire only once during a cell cycle, just as social mores ensure that most people have only one spouse at a time. But the exceptions can be interesting. Portrait of a Young Married Couple by Jacob Jordaens, image from Wikimedia Commons
Once someone is married, there are lots of things that keep them from starting a second marriage at the same time. Laws, fear of losing the first spouse, social mores and so on all create a situation where the vast majority of people have only a single spouse at any one time.
As each of these inhibitions is lifted, people will be more or less inclined towards polygamy, depending on who they are and the culture they live in. For example, if having multiple spouses becomes acceptable socially, then some people might dive right in while others might hold off.
It turns out that origins of replication are similar. There are many layers of control that keep an origin from firing more than once during any cell cycle. But just like people and polygamy, when a few inhibitory layers are removed, some origins are more likely to fire more than once in a cell cycle than are others.
In a new study out in PLOS Genetics, Richardson and Li have identified a DNA sequence that makes nearby origins of replication fire more than once during a cell cycle when certain regulatory mechanisms have been disabled. The authors hypothesize that these reinitiation promoters (RIPs) may be important for promoting genetic diversity by causing genomic duplication of specific regions under certain circumstances.
This lab had previously shown that the origin ARS317 reinitiates more frequently when global regulation is removed from some key players in initiation: Cdc6p, the Mcm2-7 complex, and the origin recognition complex (ORC). They disabled the regulation of all three of these by mutating each to prevent their recognition by the master regulator cyclin-dependent kinase (CDK, whose catalytic subunit is Cdc28p). In this study, they identified a second origin, ARS1238, that also reinitiated more often under these conditions. The authors next set out to identify why these origins reinitiated under these conditions.
The first thing they found was that chromosomal context didn’t matter a whole lot. Both origins reinitiated at around the same rate when they were in their natural context or when moved to other chromosomes. The ability to reinitiate must be contained in the sequence of the DNA that was moved.
They next showed through deletion and linker scanning analysis that the two origins both required an AT-rich, ~60 base pair sequence to reinitiate. This sequence needed to be within around 35-75 base pairs of the origin to promote reinitiation. Not any old stretch of AT-rich DNA would do; a specific DNA sequence was necessary, suggesting that this DNA is not required for reinitiation just because it is more easily unwound.
These authors have shed light on a key process in the life of a cell—the firing of an origin of replication once and only once during any cell cycle. It is critical for a cell that origins do not routinely reinitiate to prevent widespread genomic duplications that left unchecked would be very dangerous to the cell.
Richardson and Li have shown that not all origins are created equally, in that some are more likely to reinitiate under certain conditions than are others. If similar regions in mammalian cells turn out to be hotspots for genetic changes in cancers, then scientists may be able to target them to prevent the cancer’s genetic progression. We may be able to reintroduce laws to keep polygamy at bay.
by D. Barry Starr, Ph.D., Director of Outreach Activities, Stanford Genetics
Shared Domains and Phosphorylation Sites on Protein Pages
June 24, 2014
We have redesigned the Protein page to include a new tabular display of protein domains. This table provides the identifier for each domain and illustrates the respective locations of the domains within the protein. In addition to this new table, the domains are displayed in an interactive network diagram that presents the proteins that share these domains with your protein of interest (see figure below, left).
Another new feature on the Protein page is the display of phosphorylation sites within the protein’s sequence (as curated by BioGRID). This feature is available for both the reference strain S288C and other commonly used S. cerevisae strains, using the pull-down to select the desired strain view (see figure below, right) .
Left: Proteins (gray circles) that share domains (colored squares) with Fas1p (yellow circle). Right: an example of some of the phosphorylation sites in Swe1p (red residues).
Proteins that share domains with Fas1p
Swe1p protein sequence and phophorylation sites highlighted in red.
YGM Early Registration Deadline is Approaching!
June 23, 2014
There are only a few days left to register for the Yeast Genetics Meeting at the early registration rate. After midnight on Thursday, June 26, the fees will increase by $75. Conference housing is filling up fast too. This is a meeting you don’t want to miss, so don’t delay!
Like People, Prions Need Intimate Contact to Spread
June 19, 2014
In the Matrix Trilogy, the delicate balance of a virtual world is upset by a rogue computer program that goes by the name of Agent Smith. This program finds and touches other agent programs, converting them into copies of itself. Eventually, all the agent programs are copies of Agent Smith and only the hero Neo can save humanity in an epic battle within the virtual world of the Matrix.
Prions are the Agent Smiths of cells. They convert healthy proteins into prions just by touching them. Image by Marcin Wichary
A new study out in GENETICS by Li and Du provides additional evidence that prions in the yeast Saccharomyces cerevisiae work similarly to Agent Smith, in that they spread through a direct contact model. These prions are proteins that have entered a rogue conformation, and they end up converting all copies of the same protein into a similar rogue conformation. The proteins change from a hardworking Agent Smith trying to do its job into something that mucks up the working of a cell. And the results, at least in humans, can be as catastrophic for the cell as Agent Smith was for the Matrix.
Mad cow disease, for example, is caused by prions converting the prion protein (PrP) in the brain cells of people from a useful conformation to a dangerous one that spreads. As the conformation spreads throughout the cell, these prions form amyloid fibrils that eventually kill the cell. When enough brain cells are killed, the person dies.
The authors chose to work in yeast because unlike in people, there are multiple examples of proteins in yeast that can go prion. The list includes Sup35p, Ure2p, Rnq1p, Swi1p, Cyc8p, Mot3p, Sfp1p, Mod5p and Nup100p. As you might guess from the sheer number of these prion-ready proteins, prions actually do more than kill a cell in yeast; they can serve useful functions. Scientists have yet to identify any useful functions for the prion form of PrP in people.
Having multiple prions in a cell allowed Li and Du to perform some experiments to try to distinguish between two models of prion conformation spreading. In the first, called the cross-seeding model, the prion acts very much like Agent Smith in that it needs to contact a “healthy” protein to convert it into a prion. In the second model, the titration model, factors in the cell that prevent prion formation are titrated out when prions form. As the factors are taken out of commission, prions are free to form.
The main evidence in this study that supports the cross-seeding model has to do with the localization of pre-existing prions during the de novo formation of a new prion. Li and Du found that the prion [SWI+] localized to newly forming [PSI+] prions but not to already formed [PSI+] prions. This is not the result we would expect if prion formation were due to titrating out of inhibitors of prion formation. If that were the mechanism, then there would be no reason for [SWI+] to colocalize with newly forming [PSI+]. These experiments are like having a google map of the Matrix where we could see Smiths converting other agents by touch and then moving on and touching other agents.
Work like this is important for helping to find treatments for prion associated diseases and, perhaps, other amyloid fibril forming diseases like Huntington’s or Alzheimer’s. Scientists need to focus on the amyloid fiber forming proteins themselves instead of trying, for example, to ramp up the activity of factors that inhibit formation. Scientists probably need to eliminate Agent Smith to prevent the destruction of the Matrix and all of mankind.
This is how prions turn other proteins into copies of themselves:
by D. Barry Starr, Ph.D., Director of Outreach Activities, Stanford Genetics