Fred Schaufele, PhD
Our laboratory studies the mechanisms by which the genome is utilized differently in different cells. To attack this question, we compare transformed progenitor and mature cells from two different tissues that are important in the regulation of metabolism, fat and the pituitary. We seek to define what changes within the progenitor cell when these cells are induced to stop dividing and express the genes characteristic of the mature cell. We also investigate the molecular mechanisms by which gene expression patterns are altered in response to physiologic regulators. Particularly in the adipocyte this includes molecules important to diabetes regulation and treatment: nuclear receptors including PPARgamma that is the receptor for thiazolidenediones, and insulin, which we are beginning to investigate for its effects on the transcription factor C/EBPalpha.
Much of our earlier work centered around the identification of transcription factors and co-factors that control the expression of genes expressed specifically in the mature cells. These regulate the extent to which a gene is expressed. We then investigated the consequences of introducing these factors into the progenitor cells. This has led us to our current studies of the movements of these proteins within the cell. C/EBPalpha and nuclear receptors that cause the progenitor cells to differentiate and/or stop dividing are tagged with molecules that fluoresce light of particular colors. We then follow, by microscopy, the positions from which the light is locally emitted within the cell. We have found that the expression of these transcription factors causes a dramatic rearrangement in the locations of critical co-factors relative to the positions of chromatin within the cell. We refer this new-found organizational capacity of some transcription factors as "intranuclear marshaling".
We also utilize cutting-edge technologies to define precisely how the molecules interact within the living cell. Sometimes, instead of emitting light, one tagged molecule can directly contact and transfer its fluorescence energy to another tagged molecule, which then emits light of a different color. By precisely measuring how much light is transferred from one molecule to another, we define which molecules interact with each other in different positions within the cell. Because this fluorescence resonance energy transfer is highly sensitive to the distance between the molecules, we measure subtle changes of even a few Angstroms within the interactions of the proteins upon changing the cellular environment. We also define the extent to which the structure of these interactions is relatively inflexible or more variable, and again assign that to different parts of the cell. Finally, we collaborate with others within UCSF who are developing novel derivatives of nuclear receptor ligands and study whether these alter these interactions in specific ways. These unique studies allow us to directly watch and follow, in living cells, the dynamic motions within and between molecules, motions that constitute the very essence of life itself.