« Predict Toxicity from Structure
Empagliflozin »
Cells Rhythmically Regulate Their Genes
November 17, 2015 by larryhbern
Larry H. Bernstein, MD, FCAP, Curator
LPBI
http://www.technologynetworks.com/Genotyping/news.aspx?ID=184486
Study led by researchers at Caltech shows that pulsing can allow two proteins to interact with each other in a rhythmic fashion that allows them to control genes.
Even in a calm, unchanging environment, cells are not static. Among other actions, cells activate and then deactivate some types of transcription factors—proteins that control the expression of genes—in a series of unpredictable and intermittent pulses. Since discovering this pulsing phenomenon, scientists have wondered what functions it could provide for cells.
Now, a new study from Caltech researchers shows that pulsing can allow two proteins to interact with each other in a rhythmic fashion that allows them to control genes. Specifically, when the expression of the transcription factors goes in and out of sync, gene expression also goes up and down. These rhythms of activation, the researchers say, may also underlie core processes in the cells of organisms from across the kingdoms of life.
"The way transcription factor pulses sync up with one another in time could play an important role in allowing cells to process information, communicate with other cells, and respond to stress," says paper coauthor Michael Elowitz, a professor of biology and biological engineering and an investigator with the Howard Hughes Medical Institute.
http://www.technologynetworks.com/images/videos/News%20Images/PT/Msn2_green_Mig1_red-NEWS-WEB.jpg
The research was led by Caltech postdoctoral scholar Yihan Lin. Other Caltech authors of the paper are Assistant Professor of Chemistry Long Cai; Chang Ho Sohn, a staff scientist in the Cai lab; and Elowitz's former graduate student Chiraj K. Dalal (PhD '10), now at UC San Francisco.
Cai, Dalal, and Elowitz reported a functional role for transcription factor pulsing in 2008. In the meantime, researchers worldwide have been steadily uncovering similar surges of protein activity across diverse cell types and genetic systems.
Realizing that many different factors are pulsing in the same cell even in unchanging conditions, the Caltech scientists began to wonder if cells might adjust the relative timing of these pulses to enable a novel sort of time-based regulation. To find out, they set up time-lapse movies to follow two pulsing proteins and a target gene in real time in individual yeast cells.
The team tagged two central transcription factors named Msn2 and Mig1 with green and red fluorescent proteins, respectively. When the transcription factors are activated, they move into the nucleus, where they influence gene expression. This movement—as well as the activation of the factors—can be visualized because the fluorescent markers concentrate within the small volume of the nucleus, causing it to glow brightly, either green, red, or both. The color choice for the fluorescent tags was symbolic: Msn2 serves as an activator, and Mig1 as a repressor. "Msn2, the green factor, steps on the gas and turns up gene expression, while Mig1, the red factor, hits the brakes," says Elowitz.
When the scientists stressed the yeast cells by adding heat, for example, or restricting food, the pulses of Msn2 and Mig1 changed their timing with respect to one another, with more or less frequent periods of overlap between their pulses, depending upon the stressing stimulus.
Generally, when the two transcription factors pulsed in synchrony, the repressor blocked the ability of the activator to turn on genes. "It's like someone simultaneously pumping the gas and brake pedals in a car over and over again," says Elowitz.
But when they were off-beat, with the activator pulsing without the repressor, gene expression increased. "When the cell alternates between the brake and the gas—the Msn2 transcription factor in this case—the car can move," says Elowitz. As a result of these stress-altered rhythms, the cells successfully produced more (or fewer) copies of certain proteins that helped the yeast cope with the unpleasant situation.
Previously, researchers have thought that the relative concentrations of multiple transcription factors in the nucleus determine how they regulate a common gene target—a phenomenon known as combinatorial regulation. But the new study suggests that the relative timing of the pulses of transcription factors may be just as important as their concentration.
"Most genes in the cell are regulated by several transcription factors in a combinatorial fashion, as parts of a complex network," says Cai. "What we're now seeing is a new mode of regulation that controls the pulse timing of transcription factors, and this could be critical to understanding the combinatorial regulation in genetic networks."
"There appears to be a layer of time-based regulation in the cell that, because it can only be observed with movies of individual cells, is still largely unexplored," says Lin. "We look forward to learning more about this intriguing and underappreciated form of gene regulation."
In future research, the scientists will try to understand how prevalent this newfound mode of time-based regulation is in a variety of cell types and will examine its involvement in gene regulation systems. In the context of synthetic biology—the harnessing and modification of biological systems for human technological applications—the researchers also hope to develop methods to control such pulsing to program new cellular behaviors.
Combinatorial gene regulation by modulation of relative pulse timing.
Lin Y1,2, Sohn CH3, Dalal CK1,2, Cai L3, Elowitz MB1,2. Author information
Nature. Nov 5, 2015; 527(7576):54-8. doi: 10.1038/nature15710. Epub 2015 Oct 14.
Studies of individual living cells have revealed that many transcription factors activate in dynamic, and often stochastic, pulses within the same cell. However, it has remained unclear whether cells might exploit the dynamic interaction of these pulses to control gene expression. Here, using quantitative single-cell time-lapse imaging of Saccharomyces cerevisiae, we show that the pulsatile transcription factors Msn2 and Mig1 combinatorially regulate their target genes through modulation of their relative pulse timing. The activator Msn2 and repressor Mig1 showed pulsed activation in either a temporally overlapping or non-overlapping manner during their transient response to different inputs, with only the non-overlapping dynamics efficiently activating target gene expression. Similarly, under constant environmental conditions, where Msn2 and Mig1 exhibit sporadic pulsing, glucose concentration modulated the temporal overlap between pulses of the two factors. Together, these results reveal a time-based mode of combinatorial gene regulation. Regulation through relative signal timing is common in engineering and neurobiology, and these results suggest that it could also function broadly within the signalling and regulatory systems of the cell.
Pulsatile Dynamics in the Yeast Proteome
Chiraj K. Dalal,1,2 Long Cai,1,2 Yihan Lin,1 Kasra Rahbar,1 and Michael B. Elowitz1, * 1
Howard Hughes Medical Institute, Division of Biology and Bioengineering, California Institute of Technology, Pasadena, CA 91125, USA
http://dx.doi.org/10.1016/j.cub.2014.07.076
•Pulsing is prevalent in the yeast proteome
•Pulsing is specific to transcription factors
•Pulsing regulates a large fraction of the genome
The activation of transcription factors in response to environmental conditions is fundamental to cellular regulation. Recent work has revealed that some transcription factors are activated in stochastic pulses of nuclear localization, rather than at a constant level, even in a constant environment [ 1–12 ]. In such cases, signals control the mean activity of the transcription factor by modulating the frequency, duration, or amplitude of these pulses. Although specific pulsatile transcription factors have been identified in diverse cell types, it has remained unclear how prevalent pulsing is within the cell, how variable pulsing behaviors are between genes, and whether pulsing is specific to transcriptional regulators or is employed more broadly. To address these issues, we performed a proteome-wide movie-based screen to systematically identify localization-based pulsing behaviors in Saccharomyces cerevisiae. The screen examined all genes in a previously developed fluorescent protein fusion library of 4,159 strains [ 13 ] in multiple media conditions. This approach revealed stochastic pulsing in ten proteins, all transcription factors. In each case, pulse dynamics were heterogeneous and unsynchronized among cells in clonal populations. Pulsing is the only dynamic localization behavior that we observed, and it tends to occur in pairs of paralogous and redundant proteins. Taken together, these results suggest that pulsatile dynamics play a pervasive role in yeast and may be similarly prevalent in other eukaryotic species.
Since most pulsing proteins are members of a pair of paralogous or functionally redundant transcription factors, one explanation for the evolution of pulsing is one in which pulsing is ancient and existed prior to the whole-genome duplication (estimated to be w80 million years ago [20]). Since then, pulsing appears to have been lost only in some proteins (Mig3 and Rtg3), and the paralogs that have retained the ability to pulse have changed in their dynamics (Figure 3). Alternatively, paralogs that both pulse could have acquired pulsatile regulation through shared regulatory inputs that later became pulsatile. Further work analyzing whether proteins orthologous to the pulsing transcription factors described here also pulse, specifically in species that diverged prior to the whole-genome duplication, will distinguish between these hypotheses.
Recent work shows that pulsatile regulation occurs in diverse mammalian systems including NF-AT [9], p53 [10], Erk signaling [11], TGF-b signaling [12], and NF-kB [22–24]. Moreover, many bacterial systems, such as persistence in Mycobacterium smegmatis [25] and bacterial competence [26], sporulation [27], and stress response in Bacillus subtilis [28], employ pulsing. The presence of pulsing in so many systems across a wide range of species suggests that pulsing may be a common solution to many biological problems. For example, pulsing has already been shown to proportionally regulate entire regulons of target genes [2, 7], implement transient differentiation [26, 29], enable a multi-cell-cycle timer [27], and promote bet-hedging [25]. Pulsing may provide a time-based mode of regulation that facilitates these and other functions [1].
Figure 3. Pulsing Is Variable Single-cell traces show that pulses vary from cell to cell (different colors on the same trace), from paralog to paralog (across columns) and from protein to protein (A–L). All traces are from the same movie that generated corresponding filmstrips in Figure 2. All traces have been smoothed. See also Figure S2 and Movie S1. pulsing may be a common solution to many biological problems. For example, pulsing has already been shown to proportionally regulate entire regulons of target genes [2, 7], implement transient differentiation [26, 29], enable a multi-cell-cycle timer [27], and promote bet-hedging [25]. Pulsing may provide a time-based mode of regulation that facilitates these and other functions [1].
Taken together, these observations reveal that pulsatility is surprisingly pervasive in cells. It will now be critical to determine its mechanisms and functions and understand how these dynamics are integrated into the core functions of living cells. Although recent work has provided new insights into Msn2 pulsing [3, 4, 7, 8, 30, 31] and other work has provided a mechanism for pulsatile activation of a sigma factor in bacteria [28], we still lack a full understanding of the mechanisms of pulse generation and modulation for any yeast transcription factor. Do different pulsing systems use a common type of mechanism for pulsing, or are there many distinct mechanisms that can generate similar pulse dynamics? Pulsatility appears to be a core regulatory mechanism in yeast and most likely in other cell types as well [9]. The pulsatile proteins identified here should provide a starting point for understanding the roles that this dynamic regulatory mechanism plays in diverse cell types.
Jose E S Roselino
The title places scientific facts in the correct direction. However, for fast regulatory responses (those that keeps cells alive), no change in gene expression is required.
Posted in Metabolism, Proteins, Proteomics, Signaling & Cell Circuits | Tagged activation of transcription factors, pulsing, regulation | 1 Comment
on November 17, 2015 at 4:14 PM | Reply
This is very insightful. There is no doubt that there is the bias you refer to. 42 years ago, when I was postdocing in biochemistry/enzymology before completing my residency in pathology, I knew that there were very influential mambers of the faculty, who also had large programs, and attracted exceptional students. My mentor, it was said (although he was a great writer), could draft a project on toilet paper and call the NIH. It can't be true, but it was a time in our history preceding a great explosion. It is bizarre for me to read now about eNOS and iNOS, and about CaMKII-á, â, ã, ä – isoenzymes. They were overlooked during the search for the genome, so intermediary metabolism took a back seat. But the work on protein conformation, and on the mechanism of action of enzymes and ligand and coenzyme was just out there, and became more important with the research on signaling pathways. The work on the mechanism of pyridine nucleotide isoenzymes preceded the work by Burton Sobel on the MB isoenzyme in heart. The Vietnam War cut into the funding, and it has actually declined linearly since.
A few years later, I was an Associate Professor at a new Medical School and I submitted a proposal that was reviewed by the Chairman of Pharmacology, who was a former Director of NSF. He thought it was good enough. I was a pathologist and it went to a Biochemistry Review Committee. It was approved, but not funded. The verdict was that I would not be able to carry out the studies needed, and they would have approached it differently. A thousand young investigators are out there now with similar letters. I was told that the Department Chairmen have to build up their faculty. It's harder now than then. So I filed for and received 3 patents based on my work at the suggestion of my brother-in-law. When I took it to Boehringer-Mannheim, they were actually clueless.