Inner workings of molecular thermostat point to pathways to fight diseases such as diabetes and obesity

PHILADELPHIA—Researchers at the University of PennsylvaniaSchool of Medicine have discovered a molecular circuit involving heme thathelps maintain proper metabolism in the body, providing new insights intometabolic disorders such as obesity and diabetes.

Heme is best known as the oxygen-carrying component ofhemoglobin, the protein that makes blood red, but it also plays a role inchemical detoxification and energy metabolism within the cell. Heme levels aretightly maintained, and with good reason: Too little heme prevents cell growthand division; excessive amounts of heme are toxic.

The work builds on 2007 findings from the same team, led byDr. Mitchell Lazar, director of Penn's Institute for Diabetes, Obesity and Metabolism,showing that a protein called rev-erb coordinates the daily cycles of heme.

The new research, published recently online in Genes& Development, makes it clear thatrev-erb, by controlling the production of heme, also plays a key role inmaintaining the body's correct metabolism.

According to Lazar, this happens through a molecular pathwaythat allows the cell to monitor and adjust internal heme levels, creating morewhen heme levels fall, and slowing it down when levels rise.

The circuit is a negative feedback loop, with rev-erb as itscentral component, explains Lazar.

"Rev-erb is a thermostat for heme," he says.

When heme levels are high, rev-erb is activated, reducingheme, which leads the cell back towards a normal state. On the other hand, whenheme levels are low, Rev-erb alpha activity is low, and this permits the cellto make more heme, again leading back toward a normal state. Maintaining thisstasis allows energy metabolism to occur, but avoids harm to the cell due toexcessive levels of heme.

According to Lazar, the findings have huge implications fordrug discovery and development.

"Rev-erb alpha, the heme receptor, is a member of thesuperfamily of nuclear receptors that includes the receptors for a variety ofsmall molecules including hormones, vitamins, and many drugs," Lazar says."Thus it is very possible that its activity can be modified by a smallmolecule."

Understanding the control of heme levels is likely to berelevant to several diseases.

For example, obesity is a condition where fat tissue buildsup due to low-energy expenditure relative to energy intake. Lazar notes thatproteins such as rev-erb that help maintain a cell's proper metabolism andenergy balance point to their role in such metabolic disorders as obesity anddiabetes and suggest ways to intervene.

Rev-erb is a transcription factor, a protein that binds toDNA in front of, or within, genes to alter their expression. Rev-erb acts as repressor of gene expression,that is, gene expression goes down when it binds to DNA.

Lazar has been studying the protein for nearly 20 years, yethe never really knew how it worked. What he did know was that, as a member of afamily of nuclear receptor proteins, rev-erb could bind DNA and likely had an intracellular bindingpartner.

"We spent manyyears understanding its molecular properties, but didn't have a clue as to itsphysiology until Ulli Schibler in Geneva discovered that Rev-erb plays a rolein the circadian clock," he says. "That was key to moving ahead with our work,but we still did not know if rev-erb had a ligand that regulated its activity.Our discovery that heme is this key ligand regulator in 2007, simultaneouslywith another group, was key to moving this forward."

Lazar points out that understanding the control of hemelevels is key in the process.

"Heme is a critical part of many intracellular enzymes,including key components in the electron transport chain," he says. "Rev-erbsensing of heme levels seems to help to keep heme levels constant, and this inturn regulates the availability of heme for these critical biochemicalprocesses."

Typical nuclear receptor proteins are like sensors,registering a specific molecular event and responding accordingly, generally byaltering gene expression patterns. So, Lazar asked, "What is the purpose ofhaving a system that responds to changes in cellular heme levels?" Hehypothesized that the sensor could act to regulate heme itself.

Working with cultured human and mouse cells his team, led byfirst author, graduate student Nan Wu, monitored heme levels as rev-erbabundance changed. What they found confirmed the protein's role in hemeregulation: when overexpressed, heme levels dropped; when suppressed, hemelevels rose.

"That was consistent with the hypothesis," says Lazar. "Thequestion was how does heme do this?"

To figure that out, the team looked for Rev-erb binding sites within the sequences ofgenes known to control heme biosynthesis and found one in PGC-1 alpha, atranscription factor that stimulates the production of heme. Since rev-erbactivity is controlled by heme itself, the net effect is that, as heme levelsrise, PGC-1 alpha gets repressed, and heme synthesis drops off.

The team also demonstrated the physiological consequence ofdisrupting this pathway.

"We reasoned, if heme levels get too low, cells won't likeit," Lazar says.

Lazar points out that until now, no one knew there even wasa mechanism for keeping heme levels in this narrow range.

"We've shown that it exists and have defined molecularplayers that make it work," he says.

In so doing, he and his team have linked heme biosynthesiswith both energy metabolism and the body's internal clock. Rev-erb is anegative regulator of genes involved in energy metabolism. It also, along withPGC-1 and heme, rises and falls over a 24-hour period and even regulates someof the cogs within the clock itself.

The next step, Lazar says, is to determine whether thepathway can be exploited in the clinic.

"We will determine whether rev-erb regulates genes in acircadian manner in a manner that is regulated by heme, and how this integratesmetabolism with circadian rhythm," he says.

Lazar's team team showed that downregulating heme stifledcell division and metabolism, while upregulating heme enhanced them. Ittherefore is possible, he says, that by pharmacologically "tickling" rev-erb orits other cellular partners to believe the cell has more or less heme than itactually does, researchers may be able to either boost or suppress metabolism accordingly,opening the door to potential therapies for cancer and obesity.

The research was supported by the National Institute ofDiabetes and Digestive and Kidney Diseases. Lei Yin, Elyisha A. Hanniman andShree Joshi, all from Penn, are co-authors.

"There's still a lot more to be discovered about rev-erb'sbiological functions," concludes Lazar. "Continued success will entail findingthe target genes and the physiological pathways regulated by rev-erb, and thendetermining how heme levels both regulate these processes and are themselvesregulated."