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NUTRITION RESEARCH Why This Too, Too Solid Flesh Won't Melt Genes Controlling Fat Cell Differentiation Identified in Mice Fat is eating us alive, at least in the developed world. Too many fat cells and the medical problems they can drag in their wake make obesity a leading cause of death in America. Yet fat is a two-way street, or as Gökhan Hotamisligil sees it, a door that swings both in and out. Hotamisligil, HSPH associate professor of nutrition, points out that even in this overstuffed nation, there are many—HIV sufferers, anorexics, cancer patients to name but a few—who would profit from a trip through the fat cell door in the other direction. Controlling that door has long been the Holy Grail of obesity research, and while Hotamisligil says we are still a long way from having a working key, his lab has discovered what he believes to be a genetic lock, two genes that control the extent of differentiation from adipose stem cells into fat cells. Genetic locksmiths of the fat cell door: Qiang Tong (left) and Gökhan Hotamisligil. Photo by Graham Ramsay The Fat Cell Gate In the Oct. 6 Science, Hotamisligil and HSPH postdoc Qiang Tong report that two genes in mice, GATA-2 and GATA-3, are the gatekeepers that control the transition of preadipocytes to adipocytes in white adipose tissue. Mammals have two general classes of fat cell, brown and white. It is our white adipose cells that haunt us after the holidays. They engorge themselves with fat, storing excess energy as triglycerides and releasing them as free fatty acids to cover caloric deficiencies. (Brown adipose cells do the opposite, dissipating energy through thermogenesis.) The two GATA genes downregulate white adipocyte differentiation. When expressed, they inhibit the change; when silenced, they release the precursor cells from developmental arrest. "There is a pool of preadipocytes in the body and this pool actually remains more or less stable," said Hotamisligil. "Only in extreme conditions can you add to this pool. For adults, we have no concrete figures, but the available evidence indicates that the number of cells that can become adipocytes is more or less constant unless you are exposed to extreme conditions. For example, if you develop morbid obesity, then you will start to generate more of these cells." The reverse, however, is not true. Starved adipocytes release all their stored triglycerides, but their numbers remain fairly constant. Resume overeating and they resume storing fat. Once committed, adipocytes apparently do not revert. The critical factor, said Hotamisligil, is what percentage of the adipocyte precursor pool crosses over. "If all of them turn to adipocytes, than you will become obese. If a healthy proportion of them turns into adipocytes, then you will be what's called in quotes 'normal.' If none of them turns into adipocytes, then you will be abnormally thin or lipodystrophic. This transition between preadipocyte to adipocyte was known, but what was not known was what triggered the change." When Hotamisligil and Tong knocked out GATA-3, the "gate" was open for preadipocytes to differentiate into the mature adipocytes, shown here, engorged with fat. Finding Fat-to-Be The problem was identifying preadipocytes in mammals, where they are present in huge numbers in adipose tissue along with vascular endothelial cells, macrophages, and other precursor cells. Differentiated adipocytes float off in solution, said Hotamisligil, to form what's called in the lab the "fat cake." The remaining fraction is difficult to sort out. "A preadipocyte is indistinguishable from a fibroblast—indistinguishable by morphology, indistinguishable by all the molecular markers. It hasn't been possible to find a gene that would distinguish a preadipocyte from an ordinary fibroblast. No marker has been identified to tell what this cell type is." In Drosophila, though, it was another story. The Drosophila srp, or serpent, gene is a member of the GATA family of transcription factors. In fruit flies, serpent was known to contribute to the formation of "fat body." Looking to Drosophila fat body to study mammal fat cells seemed at first a "crazy idea," Hotamisligil conceded. "But I was thinking that if fat body is a primitive form of fat, then this gene could have been preserved during evolution. It turns out that in the early portions of evolution, fat body was the equivalent of many things, not only fat but also liver, also in the hematopoietic system of the fly. But this serpent gene turns out to be quite functionally conserved in mammals. Of course, it was only one gene in Drosophila. It became six genes in mice. Two of these GATAs have strong biology in the development of adipocytes." Working with murine adipose tissue, the researchers examined all six GATAs and then screened their expression libraries, zeroing in on GATA-2 and GATA-3 as the ones predominately present in white adipose tissue. Floating off the mature adipocytes in fat cake, they found that the two GATAs were strongly expressed only in the fractionate containing the preadipocytes. The researchers then enriched the preadipocytes in culture and, by up- and down-regulating the expression of the two GATAs, they found that they could turn fat cell differentiation on and off. The Path Toward Fat These are heady times in adipocyte research, according to Hotamisligil. Recent discoveries have unraveled the downstream development of adipocytes once they begin to differentiate. That pathway is controlled by the coordinated actions of PPAR-gamma2, a peroxisome proliferator–activated receptor, and the C/EBP family of transcription factors. Tong and Hotamisligil were able to connect their new piece of the upstream pathway directly through PPAR-gamma2, showing that the two GATAs acted by binding PPAR-gamma2, directly suppressing its activity. In effect, GATA-2 and GATA-3 were the brakes on differentiation; when they were turned off, the preadipocyte was free to roll down the pathway to fat cell maturity. The identification of these two genes in adipogenesis extends our knowledge of the molecular pathway, Hotamisligil said. Having a new way of flagging preadipocytes also opens new questions to experimentation; for example, the effect of aging on their total numbers and on their ratio to adipocytes. "But the more important aspect here is that we've put our foot into a very important door. As we study the mechanism that controls this door, we will find other ways to open this lock and other ways to close it. Now we know that this is the door and this is the handle, but there are many ways to jam a lock." —John Fleischman