The body's immediate reaction to serious trauma, denoted the Acute Phase Response (APR), coordinates a wide variety of inflammatory signals, and produces a common response, in the massive induction of protective proteins by the liver. While the APR is designed for survival, severe or prolonged activation with its interruption of normal homeostatic functions, likely contributes to organ failure and death in some critically ill patients. Evidence suggests that the APR's repression of steady-state liver function is a byproduct of its mode of induction. Further evidence suggests that this is a highly regulated process, which shares pathways important to early cell type development, and with some signals associated with the proliferative response. Evidence is presented that injury induced regulation of differentiated genes may be mediated through phosphorylation of liver-specific transcription factors, particularly HNF-4. Better understanding of the mechanisms regulating this process would have therapeutic importance in advantageous support of the acute phase response.
The first aim will dissect where on HNF-4 phosphorylation occurs. This will be done by phosphopeptide mapping. We will also identify the kinases and thus the signal transduction pathways that are involved in initiating the APR. This will generate original materials, in the form of altered HNF-4 molecules, to test the importance of this phosphorylation to the APR.
The second aim i s to trace the effect of this phosphorylation on the biochemical activities of HNF-4, its DNA binding and site selection, and changes in its interactions and activities in the cell. This will be carried out using diagnostic chromatin immunoprecipitation.
The third Aim will use DNA microarrays, to produce a picture of the genes regulated by HNF-4 before and after injury. An in vitro model of acute phase induction will also be used to allow direct manipulation of HNF-4 and to measure its effects on acute phase transcriptional events. These studies will elucidate the mechanisms of HNF-4 functions and test, the importance of HNF-4's modifications. It will establish what transcriptional events are common to the early phase of the APR, when normal liver function is modified, and the liver prepares for subsequent massive modulation of protein production.
