Ott Lab News

Three scientists, including Gladstone’s Melanie Ott, MD, PhD, have been chosen to receive the 2014 Avant-Garde Award for HIV/AIDS Research from the National Institute on Drug Abuse (NIDA), part of the National Institutes of Health. The three scientists will each receive $500,000 per year for five years to support their research.

Project: A new model of accelerated immune aging in HIV-infected drug users

Dr. Ott will investigate the role of an enzyme (SIRT-1) in slowing accelerated immune aging resulting from either long-term HIV infection or regular drug use. Because SIRT-1 appears to protect against overworked immune activation that can eventually exhaust immune cell functions, new therapies aimed at this enzyme could delay immune aging and its related health risks in HIV-infected drug users.

“The goal of our research is to transform our understanding of how HIV and drug abuse affect the immune system and the aging process,” Ott said. “We hope to identify novel links between HIV, abused substances and the biological pathways of aging that lead to potential therapeutic strategies to slow the accelerated immune aging in this patient population.”

“These innovative approaches can shed light on mechanisms through which HIV damages or circumvents the immune system, and how these effects interact with those of drugs of abuse” said NIDA Director Nora D. Volkow, M.D. “By learning more about these underlying processes, not only might this research slow the progression and transmission of HIV infection, but it could improve the long-term health of those already infected.”

November 7, 2013—One of biology’s most fundamental processes is something called transcription. It is just one step of many required to build proteins—and without it life would not exist. However, many aspects of transcription remain shrouded in mystery. But now, scientists at the Gladstone Institutes are shedding light on key aspects of transcription, and in so doing are coming even closer to understanding the importance of this process in the growth and development of cells—as well as what happens when this process goes awry.

In the latest issue of Molecular Cell, researchers in the laboratory of Gladstone Investigator Melanie Ott, MD, PhD, describe the intriguing behavior of a protein called RNA polymerase II (RNAPII). The RNAPII protein is an enzyme, a catalyst that guides the transcription process by copying DNA into RNA, which forms a disposable blueprint for making proteins. Scientists have long known that RNAPII appears to stall or “pause” at specific genes early in transcription. But they were not sure as why.

“This so-called ‘polymerase pausing’ occurs when RNAPII literally stops soon after beginning transcription for a short period before starting up again,” explained Dr. Ott. “All we knew was that this behavior was important for the precise transcription of DNA into RNA, so we set out to understand how, when and—most importantly—why.”

The research team focused their efforts on a segment of RNAPII called the C-terminal domain, or CTD. This section is most intimately involved with transcription regulation. Previous research had found that CTD’s chemical structure is modified before and during transcription. However, the combinations of modifications as well as precisely how they influence or control transcription remained unclear. So in laboratory experiments on cells extracted from mammals, the researchers took a closer look.

The first breakthrough came when the research team identified a new type of modification, known asacetylation, which regulated transcription.

“Our next breakthrough occurred when we pinpointed the precise locations on the CTD where acetylation occurred—and realized it was unique to higher eukaryotes,” explained Sebastian Schröder, PhD, the paper’s first author. “We then wanted to see how this mammalian-specific acetylation fit into the realm of polymerase pausing.”

Now that the team knew where the CTD became acetylated, their next goal was to find out when. Clues to the timing of acetylation came in experiments where they mutated RNAPII so that CDT was unable to become acetylated. In these cases, the length of polymerase pausing dropped, and the necessary steps for the completion of transcription failed to occur. Additional experiments revealed the elusive timeline of acetylation and transcription.

“RNAPII binds to DNA to prepare for transcription. Shortly after that we see polymerase pausing—at which point the CTD becomes highly acetylated,” continued Dr. Shröder. “Soon after the pause, CTD is thendeacetylated—the original modification is reversed—and transcription continues without a hitch.”

Polymerase pausing is not unique to mammals—in fact it was characterized in HIV, the virus that causes AIDS, many years ago—but the fact that the CTD becomes acetylated just before or during the time when transcription is paused appears to be unique. Drs. Ott and Schröder argue that CTD acetylation is a stabilizer, preparing RNAPII for efficient completion of transcription and slowing down the process to make sure everything is functioning correctly—not unlike the final ‘systems check’ a pilot must perform before takeoff.

These findings offer important insight into the relationship between acetylation and transcription. And given the importance of transcription in the growth and maturation of cells in general, the team’s result stands to inform scientists about a variety of cellular processes. These include, for example, the mechanisms behind stem-cell development and what happens when normal cellular growth spirals out of control, such as in cancer.

“However, there is still much we don’t know about acetylation as it relates to transcription,” said Dr. Ott. “For example, if CTD acetylation is important for stabilizing transcriptional pausing, why do we also see CTD acetylation at non-paused genes, although at different locations? Further, we believe there may be other steps in the transcription cycle that depend upon acetylation. Our most immediate goal is to find them. By doing so, we hope to deepen our understanding of one of nature’s most elegant biological processes.”

Publication: Schröder S, Herker E, Itzen F, He D, Thomas S, Gilchrist DA, Kaehlcke K, Cho S, Pollard KS, Capra JA, Schnolzer M, Cole PA, Geyer M, Bruneau BG, Adelman K, Ott M (2013) Acetylation of RNA Polymerase II Regulates Growth-Factor-Induced Gene Transcription in Mammalian Cells. Mol Cell 52:314–324.