Forum for Science, Industry and Business

Sponsored by:     3M 
Search our Site:

 

Next-gen reappraisal of interactions within a cancer-associated protein complex

16.01.2014
Application of global sequencing technology reveals how an activator of gene expression stays focused

At a glance, DNA is a rather simple sequence of A, G, C, T bases, but once it is packaged by histone proteins into an amalgam called chromatin, a more complex picture emerges. Histones, which come in four subtypes—H2A, H2B, H3, and H4—can either coil DNA into inaccessible silent regions or untwist it to allow gene expression. To further complicate things, small chemical flags, such as methyl groups, affect whether histones silence or activate genes.


This image shows the context-dependent function of COMPASS.

Credit: Image courtesy of Dr. Ali Shilatifard, Stowers Institute for Medical Research

Among activator histones is a form of H3 decorated at a precise location (defined as H3K4) with three methyl groups (known as "H3K4me3"). Researchers knew previously that the presence of H2B exhibiting a single ubiquitin molecule stimulated the methylase that modifies H3K4, thereby increasing H3K4me3 levels. But how the methylase's activity was directed toward the appropriate targets had remained unclear.

Now, using biochemical, structural, and global sequencing techniques, researchers in the lab of Ali Shilatifard, Ph.D., an Investigator at the Stowers Institute for Medical Research, reveal an unanticipated mechanism underlying H3K4 trimethylation. Their study, published in the January 15, 2014 issue of Genes & Development, explains why H3K4me3 is deposited adjacent to a target gene promoter rather than haphazardly across the entire gene. This finding is significant because mutations in the human gene encoding the methylase responsible for H3K4me3 are associated with childhood leukemias, among other malignancies.

The work also illustrates the way powerful new genome-wide sequencing methodologies are impacting all molecular biology, including cancer research. "Here, we show that one cannot rely on methods that simply measure overall bulk H3K4me3 levels in vitro," says Shilatifard. "Only genome-wide sequencing could have revealed that H3K4 trimethylation was promoter-specific in non-mutant yeast."

This means that many assumptions about gene expression may need to be retested using next-generation approaches. "The old technologies were like observing just one region of the earth from a distant telescope in space and then making assumptions about what the entire earth looked like," Shilatifard says. "With new technologies, we can now see the whole planet."

The methylase in question, named SET1 in yeast and MLL in mammals, is part of a protein aggregate called COMPASS, for COMplex of Proteins ASsociated with Set1. Shilatifard was the first to define the role of COMPASS in chromatin modification. "Over a decade ago, our lab used yeast to show that COMPASS was an H3 methylase," he says. "Since these fundamental systems are highly conserved from yeast to Drosophila to humans, we took advantage of the awesome power of yeast genetics to identify what regulates H3K4 methylation activity."

Part of the paper addresses SET1/MLL regulation by different proteins within yeast COMPASS. Investigators knew that if you lopped off more than half of SET1's front end, levels of DNA-bound trimethylated H3K4 in cells harboring the remaining "stub" were equal to those in cells containing the full-length protein when analyzed in bulk. This led some to presume that the entire front end of SET1/MLL, as well as factors that interact with it, must not be needed to regulate H3K4me3 activity.

The new paper shows that this presumption was not correct. The Shilatifard team first employed biochemical methods to capture every piece of DNA bound to H3K4me3 in the genome of yeast harboring either full-length SET1 or the stub missing the front end. They then sequenced all of those DNA fragments and mapped their position in the yeast genome.

Significantly, they found that even though H3K4me3 levels in bulk were equivalent in normal and mutant cells, H3K4me3 was differentially distributed throughout the genome: in normal cells, H3K4me3 complexes sat primarily on DNA regions that switch adjacent genes on or off, control regions called promoters. By contrast, the DNA of cells harboring the stub exhibited DNA-binding H3K4me3 complexes in the middle of or between genes.

These discoveries, combined with other findings, call for re-interpretation of data suggesting that the stub is all you need for H3K4 trimethylation. Instead, the new work shows that COMPASS factors that bind to the SET1/MLL front end limit H3K4me3 deposition to the correct genomic sites (that is, to the promoter regions), while factors that bind the SET1/MLL stub increase the protein's half-life. This partially explains earlier misinterpretations: highly stable stubs of SET1 "promiscuously" methylated the wrong parts of the genome when the regulatory front end of the protein was missing. The paper also addressed how H2B ubiquitin modification machineries stimulate the entire process.

Understanding COMPASS regulation is essential, as genes encoding factors in the complex are mutant in numerous cancers. For example, chromosomal translocations involving a gene encoding one MLL protein occur frequently in human leukemias, hence the designation MLL, which stands for Mixed Lineage Leukemia protein. Other MLL proteins are strongly implicated as tumor suppressors in human cancers such as lymphoma and pediatric brain tumors.

Janet L. Thornton, a Senior Research Technician in the Shilatifard lab, was the study's lead author. Also contributing were Yoh-hei Takahashi, Ph.D., Malcolm Cook, Xin Gao, Ph.D., Ashley R. Woodfin, Jung-Shin Lee, Ph.D., Marc A. Morgan, Ph.D., and Edwin R. Smith, Ph.D., from the Stowers Institute; Georgios Skiniotis, Ph.D., and Gerwin H. Westfield, Ph.D., from the University of Michigan; Jessica Jackson from Saint Louis University School of Medicine; and Jean-Francois Couture, Ph.D., from the University of Ottawa.

Funding for the study came from the Stowers Institute for Medical Research, a Canadian Institutes of Health Research grant (BMA-355900), a National Institute of General Medical Sciences Training Grant (GM008270-23), the Pew Charitable Trusts, and the National Institutes of Health (R01GM069905).

About the Stowers Institute for Medical Research

The Stowers Institute for Medical Research is a non-profit, basic biomedical research organization dedicated to improving human health by studying the fundamental processes of life. Jim Stowers, founder of American Century Investments, and his wife, Virginia, opened the Institute in 2000. Since then, the Institute has spent over 900 million dollars in pursuit of its mission.

Currently, the Institute is home to nearly 550 researchers and support personnel; over 20 independent research programs; and more than a dozen technology-development and core facilities.

Gina Kirchweger | EurekAlert!
Further information:
http://www.stowers.org

More articles from Life Sciences:

nachricht An evolutionary heads-up – The brain size advantage
22.05.2015 | Veterinärmedizinische Universität Wien

nachricht Endocrine disrupting chemicals in baby teethers
21.05.2015 | Goethe-Universität Frankfurt am Main

All articles from Life Sciences >>>

The most recent press releases about innovation >>>

Die letzten 5 Focus-News des innovations-reports im Überblick:

Im Focus: Basel Physicists Develop Efficient Method of Signal Transmission from Nanocomponents

Physicists have developed an innovative method that could enable the efficient use of nanocomponents in electronic circuits. To achieve this, they have developed a layout in which a nanocomponent is connected to two electrical conductors, which uncouple the electrical signal in a highly efficient manner. The scientists at the Department of Physics and the Swiss Nanoscience Institute at the University of Basel have published their results in the scientific journal “Nature Communications” together with their colleagues from ETH Zurich.

Electronic components are becoming smaller and smaller. Components measuring just a few nanometers – the size of around ten atoms – are already being produced...

Im Focus: IoT-based Advanced Automobile Parking Navigation System

Development and implementation of an advanced automobile parking navigation platform for parking services

To fulfill the requirements of the industry, PolyU researchers developed the Advanced Automobile Parking Navigation Platform, which includes smart devices,...

Im Focus: First electrical car ferry in the world in operation in Norway now

  • Siemens delivers electric propulsion system and charging stations with lithium-ion batteries charged from hydro power
  • Ferry only uses 150 kilowatt hours (kWh) per route and reduces cost of fuel by 60 percent
  • Milestone on the road to operating emission-free ferries

The world's first electrical car and passenger ferry powered by batteries has entered service in Norway. The ferry only uses 150 kWh per route, which...

Im Focus: Into the ice – RV Polarstern opens the arctic season by setting course for Spitsbergen

On Tuesday, 19 May 2015 the research icebreaker Polarstern will leave its home port in Bremerhaven, setting a course for the Arctic. Led by Dr Ilka Peeken from the Alfred Wegener Institute, Helmholtz Centre for Polar and Marine Research (AWI) a team of 53 researchers from 11 countries will investigate the effects of climate change in the Arctic, from the surface ice floes down to the seafloor.

RV Polarstern will enter the sea-ice zone north of Spitsbergen. Covering two shallow regions on their way to deeper waters, the scientists on board will focus...

Im Focus: Gel filled with nanosponges cleans up MRSA infections

Nanoengineers at the University of California, San Diego developed a gel filled with toxin-absorbing nanosponges that could lead to an effective treatment for skin and wound infections caused by MRSA (methicillin-resistant Staphylococcus aureus), an antibiotic-resistant bacteria. This "nanosponge-hydrogel" minimized the growth of skin lesions on mice infected with MRSA - without the use of antibiotics. The researchers recently published their findings online in Advanced Materials.

To make the nanosponge-hydrogel, the team mixed nanosponges, which are nanoparticles that absorb dangerous toxins produced by MRSA, E. coli and other...

All Focus news of the innovation-report >>>

Anzeige

Anzeige

Event News

International symposium: trends in spatial analysis and modelling for a more sustainable land use

20.05.2015 | Event News

15th conference of the International Association of Colloid and Interface Scientists

18.05.2015 | Event News

EHFG 2015: Securing health in Europe. Balancing priorities, sharing responsibilities

12.05.2015 | Event News

 
Latest News

Mesoporous Particles for the Development of Drug Delivery System Safe to Human Bodies

22.05.2015 | Materials Sciences

Computing at the Speed of Light

22.05.2015 | Information Technology

Development of Gold Nanoparticles That Control Osteogenic Differentiation of Stem Cells

22.05.2015 | Materials Sciences

VideoLinks
B2B-VideoLinks
More VideoLinks >>>