{"id":8870,"date":"2016-05-31T09:28:05","date_gmt":"2016-05-31T09:28:05","guid":{"rendered":"http:\/\/revoscience.com\/en\/?p=8870"},"modified":"2016-05-31T09:28:05","modified_gmt":"2016-05-31T09:28:05","slug":"8870","status":"publish","type":"post","link":"https:\/\/www.revoscience.com\/en\/8870\/","title":{"rendered":"Scientists illuminate a hidden regulator in gene transcription"},"content":{"rendered":"

New super-resolution technique visualizes important role of short-lived enzyme clusters.\u00a0<\/strong><\/em><\/p>\n

\"MIT<\/a>
MIT scientists developed a super-resolution imaging technique, using a combination of multi-colored lasers and mirrors, to visualize very tiny, transient phenomena, such as enzyme clustering on genes.
Photo: Jose-Luis Olivares\/MIT<\/figcaption><\/figure>\n

CAMBRIDGE, Mass. —\u00a0Gene transcription is the process by which DNA is copied and synthesized as messenger RNA (mRNA) \u2014 which delivers its genetic blueprints to the cell\u2019s protein-making machinery.<\/p>\n

Now researchers at MIT and the Howard Hughes Medical Institute (HHMI) have identified a hidden, ephemeral phenomenon in cells that may play a major role in jump-starting mRNA production and regulating gene transcription.<\/p>\n

In a paper published in the online journal\u00a0eLife<\/em>, the researchers report using a new super-resolution imaging technique they\u2019ve developed, to see individual mRNA molecules coming out of a gene in a live cell. Using this same technique, they observed that, just before mRNA\u2019s appearance, the enzyme RNA polymerase II (Pol II) gathers in clusters on the same gene for just a few brief seconds before scattering apart.<\/p>\n

When the researchers manipulated the enzyme clusters in such a way that they stayed together for longer periods of time, they found that the gene produced correspondingly more molecules of mRNA. Clusters of Pol II therefore may play a central role in triggering mRNA production and controlling gene transcription.<\/p>\n

Ibrahim Ciss\u00e9, assistant professor of physics at MIT, explains that because of their transient nature, enzyme clusters have largely been regarded as a mystery, and scientists have questioned whether such clustering is purposeful or merely coincidental. These new results, he says, suggest that, although short-lived, enzyme clustering can have a significant impact on major biological processes.<\/p>\n

\u201cWe think these weak and transient clusters are a fundamental way for the cell to control gene expression,\u201d says Ciss\u00e9, who is senior author on the paper. \u201cIf a small mutation changes the cluster\u2019s lifetime ever so slightly, that can also change the gene expression in a major way. It seems to be a very sensitive knob that the cell can tune.\u201d<\/p>\n

What\u2019s more, Ciss\u00e9 says scientists can now explore Pol II clusters as targets to \u201cstall or induce a burst of transcription\u201d and control the expression of certain genes, to explore cancer drugs and other gene therapies.<\/p>\n

The paper\u2019s co-authors include Won-Ki Cho, lead author and postdoc in the Department of Physics; Namrata Jayanth and Jan-Henrik Spille, also postdocs in physics; Takuma Inoue and J. Owen Andrews, graduate students in physics; and William Conway, an undergraduate in physics and biology; as well as researchers from HHMI\u2019s Janelia Research Campus: Brian English, Jonathan Grimm, Luke Lavis, and Timoth\u00e9e Lionnet who is also co-senior author with Ciss\u00e9.<\/p>\n

Imaging at super-resolution<\/strong><\/p>\n

Pol II enzymes only cluster together for very short periods of time, on the order of several seconds. These clusters are also extremely small, on the scale of 100 nanometers in width. Because they are so tiny and fleeting, Pol II clusters and other weak and transient interactions have largely been hidden from view, essentially invisible to conventional imaging techniques.<\/p>\n

To see these interactions, Ciss\u00e9 and his colleagues developed a super-resolution imaging technique to visualize cellular processes at the single-molecule level. The team\u2019s technique builds on two existing super-resolution methods \u2014 photo-activation localization microscopy (PALM) and stochastic optical reconstruction microscopy (STORM). Both techniques involve tagging molecules of interest and lighting them up one by one to determine where each molecule is in space. Scientists can then merge every molecule\u2019s position to create one super-resolution image of the cellular region.<\/p>\n

While incredibly precise, these imaging techniques rely on the assumption that every molecule remains stationary. Molecules that come and go, and quickly cluster and scatter, are difficult to track. To catch Pol II clusters in action, Ciss\u00e9 and his team tweaked existing super-resolution imaging techniques, looking not just at a single enzyme\u2019s position, but also at how frequently molecules were detected. The higher the frequency of detection, the higher the chance that a cluster has formed.<\/p>\n

The team applied their technique to image cells, using a camera that recorded one frame every 50 milliseconds, running continuously for up to 10,000 frames.<\/p>\n

A transient lifetime<\/strong><\/p>\n

They then created a cell line that included a bright fluorescent tag for mRNA, as well as a fluorescent tag of a different color for Pol II enzymes. The team applied its super-resolution technique to image a particular gene inside the cell, called beta-actin, which has been characterized extensively. In experiments with live cells, the researchers observed that, while previously transcribed mRNA molecules lit up on the gene, new Pol II clusters appeared on the same gene, for about 8 seconds, before disassembling.<\/p>\n

From these experiments, the group was uncertain as to whether the clusters had any impact on mRNA production, since the time it takes from the start of transcription to the complete production of mRNA takes significantly longer \u2014 about 2.5 minutes. Could a cluster, appearing for just a fraction of that time, have any effect on mRNA?<\/p>\n

To answer this question, the team stimulated the cells with a chemical cocktail which they knew would affect gene transcription and mRNA output. In these cells, they found that, just before the mRNA peak appeared, clusters formed on the gene and actually remained stable for as long as 24 seconds \u2014 a fourfold increase in a cluster\u2019s typical lifetime. What\u2019s more, the number of mRNAs increased by a similar amount.<\/p>\n

After repeating the experiment in 207 living cells, the team found that the lifetime of Pol II clusters was directly related to the number of mRNA produced from the same gene.<\/p>\n

Ciss\u00e9 speculates that perhaps Pol II clusters acts as an efficient driver of gene transcription, speeding up an otherwise inefficient process.<\/p>\n

\u201cIt makes sense that you wouldn\u2019t want an efficient initiation process, because you don\u2019t want to randomly turn on any gene just because there was a random collision,\u201d Ciss\u00e9 says. \u201cBut you also want to have a way to change the initiation from an inefficient to an efficient process, for example when you want to express a gene in response to some environmental stimuli. We think that these transient clusters are probably the way that the cell can render transcription initiation efficient.\u201d<\/p>\n

Next, Ciss\u00e9 plans to follow up his studies on Pol II clusters to determine what are the forces holding them together, as well as how they\u2019re formed, and whether other molecular factors cluster with similar effects.<\/p>\n

\u201cI suspect there are new biophysical phenomena that come from weak and transient interactions,\u201d Ciss\u00e9 says. \u201cThis is an underexplored area in biology, and because the interactions are so elusive we understand very little about how the regulatory processes happen inside a living cell.\u201d<\/p>\n

This research was funded, in part, by the National Institutes of Health Director\u2019s New Innovator Award to Ciss\u00e9, and additional support from the National Cancer Institute, the MIT physics department start-up funds, and the Howard Hughes Medical Institute.<\/p>\n","protected":false},"excerpt":{"rendered":"

CAMBRIDGE, Mass. — Gene transcription is the process by which DNA is copied and synthesized as messenger RNA (mRNA) \u2014 which delivers its genetic blueprints to the cell\u2019s protein-making machinery. <\/p>\n","protected":false},"author":2,"featured_media":8871,"comment_status":"open","ping_status":"open","sticky":false,"template":"","format":"standard","meta":{"footnotes":""},"categories":[17],"tags":[],"class_list":["post-8870","post","type-post","status-publish","format-standard","has-post-thumbnail","hentry","category-research"],"featured_image_urls":{"full":["https:\/\/www.revoscience.com\/en\/wp-content\/uploads\/2016\/05\/MIT-Seeing-Transcription-1_0.jpg",639,426,false],"thumbnail":["https:\/\/www.revoscience.com\/en\/wp-content\/uploads\/2016\/05\/MIT-Seeing-Transcription-1_0-150x150.jpg",150,150,true],"medium":["https:\/\/www.revoscience.com\/en\/wp-content\/uploads\/2016\/05\/MIT-Seeing-Transcription-1_0-300x200.jpg",300,200,true],"medium_large":["https:\/\/www.revoscience.com\/en\/wp-content\/uploads\/2016\/05\/MIT-Seeing-Transcription-1_0.jpg",639,426,false],"large":["https:\/\/www.revoscience.com\/en\/wp-content\/uploads\/2016\/05\/MIT-Seeing-Transcription-1_0.jpg",639,426,false],"1536x1536":["https:\/\/www.revoscience.com\/en\/wp-content\/uploads\/2016\/05\/MIT-Seeing-Transcription-1_0.jpg",639,426,false],"2048x2048":["https:\/\/www.revoscience.com\/en\/wp-content\/uploads\/2016\/05\/MIT-Seeing-Transcription-1_0.jpg",639,426,false],"ultp_layout_landscape_large":["https:\/\/www.revoscience.com\/en\/wp-content\/uploads\/2016\/05\/MIT-Seeing-Transcription-1_0.jpg",639,426,false],"ultp_layout_landscape":["https:\/\/www.revoscience.com\/en\/wp-content\/uploads\/2016\/05\/MIT-Seeing-Transcription-1_0.jpg",639,426,false],"ultp_layout_portrait":["https:\/\/www.revoscience.com\/en\/wp-content\/uploads\/2016\/05\/MIT-Seeing-Transcription-1_0.jpg",600,400,false],"ultp_layout_square":["https:\/\/www.revoscience.com\/en\/wp-content\/uploads\/2016\/05\/MIT-Seeing-Transcription-1_0.jpg",600,400,false],"newspaper-x-single-post":["https:\/\/www.revoscience.com\/en\/wp-content\/uploads\/2016\/05\/MIT-Seeing-Transcription-1_0.jpg",639,426,false],"newspaper-x-recent-post-big":["https:\/\/www.revoscience.com\/en\/wp-content\/uploads\/2016\/05\/MIT-Seeing-Transcription-1_0.jpg",540,360,false],"newspaper-x-recent-post-list-image":["https:\/\/www.revoscience.com\/en\/wp-content\/uploads\/2016\/05\/MIT-Seeing-Transcription-1_0.jpg",95,63,false],"web-stories-poster-portrait":["https:\/\/www.revoscience.com\/en\/wp-content\/uploads\/2016\/05\/MIT-Seeing-Transcription-1_0.jpg",639,426,false],"web-stories-publisher-logo":["https:\/\/www.revoscience.com\/en\/wp-content\/uploads\/2016\/05\/MIT-Seeing-Transcription-1_0.jpg",96,64,false],"web-stories-thumbnail":["https:\/\/www.revoscience.com\/en\/wp-content\/uploads\/2016\/05\/MIT-Seeing-Transcription-1_0.jpg",150,100,false]},"author_info":{"info":["RevoScience"]},"category_info":"Research<\/a>","tag_info":"Research","comment_count":"0","_links":{"self":[{"href":"https:\/\/www.revoscience.com\/en\/wp-json\/wp\/v2\/posts\/8870","targetHints":{"allow":["GET"]}}],"collection":[{"href":"https:\/\/www.revoscience.com\/en\/wp-json\/wp\/v2\/posts"}],"about":[{"href":"https:\/\/www.revoscience.com\/en\/wp-json\/wp\/v2\/types\/post"}],"author":[{"embeddable":true,"href":"https:\/\/www.revoscience.com\/en\/wp-json\/wp\/v2\/users\/2"}],"replies":[{"embeddable":true,"href":"https:\/\/www.revoscience.com\/en\/wp-json\/wp\/v2\/comments?post=8870"}],"version-history":[{"count":0,"href":"https:\/\/www.revoscience.com\/en\/wp-json\/wp\/v2\/posts\/8870\/revisions"}],"wp:featuredmedia":[{"embeddable":true,"href":"https:\/\/www.revoscience.com\/en\/wp-json\/wp\/v2\/media\/8871"}],"wp:attachment":[{"href":"https:\/\/www.revoscience.com\/en\/wp-json\/wp\/v2\/media?parent=8870"}],"wp:term":[{"taxonomy":"category","embeddable":true,"href":"https:\/\/www.revoscience.com\/en\/wp-json\/wp\/v2\/categories?post=8870"},{"taxonomy":"post_tag","embeddable":true,"href":"https:\/\/www.revoscience.com\/en\/wp-json\/wp\/v2\/tags?post=8870"}],"curies":[{"name":"wp","href":"https:\/\/api.w.org\/{rel}","templated":true}]}}