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October 2003, Vol. 9, No. 3 Biomedical Research Conducting basic biomedical research, such as illuminating the events in cells at a molecular level, is a painstaking endeavor that can take decades to produce major results. But knowledge garnered from this type of study is crucial to the formulation of new drugs and the development of successful strategies for tackling health problems. For more than 15 years, Population Council biomedical researcher James F. Catterall and his colleagues have been studying the genetic mechanisms of action of male sex hormones, or androgens. Catterall is the director of the reproductive physiology and immunology program at the Council’s Center for Biomedical Research. His lab has devised ingenious ways of gleaning information about how androgens regulate the activity of genes and why they affect certain tissues and not others. Scientists in the Catterall lab are studying the effects of androgen in the kidney, rather than in the reproductive system. “We expect androgens to use the same mechanism of action in the kidney as they use in the reproductive system,” says Catterall. By using kidney cells, the research team is able to simplify its studies. For example, germ cells, which exist only in the reproductive system, undergo meiosis, the type of cell division that results in sperm and eggs. At various points in this complex process, cells can have either the normal amount of DNA, twice the normal amount of DNA, or half the normal amount of DNA. The ever-varying amount of genetic material present in these cells complicates the analysis of gene regulation. Cells in the kidney, in contrast, always have the normal amount of DNA; they never undergo meiosis. Catterall and his colleagues study the gene for the kidney androgen-regulated protein, known as the Kap gene. The importance of androgens Genes make the proteins needed for various cellular tasks. All functional genes have a section called a “promoter.” The promoter precedes the part of the gene that contains the code, or recipe, for the gene’s protein. “Essentially, the promoter is a message for the cell’s protein-making apparatus that says ‘start here,’ ” says Dianne O. Hardy, a scientist in Catterall’s lab. Catterall and his colleagues wanted to study the Kap gene promoter to determine what parts of it were involved in the regulation of the gene by androgen. To do this, they created a hybrid gene in the laboratory. They took part of the promoter from the Kap gene and attached it to the genetic code for another protein. Then they genetically engineered mice to carry this hybrid gene along with the normal Kap gene. In these mice, the researchers could compare the activity of the hybrid gene to that of the normal gene when androgen levels were manipulated experimentally. They were able to tell the difference between the activities of the two genes by comparing the amounts of the two different proteins they produce. The protein-coding section of the hybrid gene is known as the “reporter” because its activity in the cell provides information about the functioning of the promoter. Catterall and his team removed a section of the hybrid gene’s promoter known as the L1 repeat. When they compared the hybrid gene's activity to that of the true gene, they found that the reporter gene no longer responded to androgen. This surprised the researchers because the L1 repeat was previously thought to be unnecessary for proper gene function. Now they believe that the L1 sequence is required for Kap expression and its regulation by testosterone. Catterall and his colleagues are conducting research to further quantify the role of the L1 repeat in the Kap gene. In vivo or in vitro? In the future, the investigators hope to determine how androgen exerts its influence in some tissues and not in others. At least two things may play a role in the specificity of androgen action. Variations in the promoter sections of androgen-responsive genes probably play a role. Another factor may be the action of co-regulator substances that are present in some tissues and not others. Tissue-specific complexes made from these substances may modulate hormone action. Sources Ding, Yueming, Robin L. Davisson, Dianne O. Hardy, Li-Ji Zhu, David C. Merrill, James F. Catterall, and Curt D. Sigmund. 1997. “The kidney androgen-regulated protein promoter confers renal proximal tubule cell-specific and highly androgen-responsive expression on the human angiotensinogen gene in transgenic mice,” Journal of Biological Chemistry 272(44): 28142–28148. Outside funding The Andrew W. Mellon Foundation and the National Institutes of Health See Also
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