The Acute Phase reaction characterizes the sequelae to acute traumatic injury with blood loss and other forms of """"""""shock"""""""" that mimic fluid depletion. A major organ to respond to acute shock is the liver. It responds to synthesizing and secreting many proteins including clotting factor I (fibrinogen). The level of mRNA coding for the alpha, beta and gamma fibrinogen genes increases 10 fold after defibrinogenation. In the absence of corticosteroids this response is reduced by 50%. The goal of this project is to understand the coordinate expression of these three genes at the genomic level. Clones of rat genomic DNA have been analyzed by RNA mapping and subsequent DNA sequencing. The regions flanking the 5 feet ends of each gene have been sequenced. A computer assisted analysis reveals two particular areas of homology between these genes. One corresponds to the 21 bp consensus sequence found in other genes that respond to corticosteroids. The other consists of a 84 bp region that may be responsible for the regulation of fibrinogen transcription by Hepatocyte Stimulating factor (HSF). The role that these areas of homology play in transcriptional regulation will be analyzed in two systems. One will test for enhancer and activator functions by subcloning the areas of interest into plasmids that contain the chloramphenicol acetylase gene. These will be used as described by Gruss. Here one can determine if the inserted sequences can enhance transcription in primary rat hapatocyte cultures and HSF. The other system will use a portion of the gamma fibrinogen gene cloned into a specially constructed adenovirus. These recombinant viruses will be used to deliver deletion mutations and site-specific base-substituted mutations into hepatocyte cultures. Thus, the sequences required for the proper transcription of fibrinogen in response to corticosteroids and HSF will be determined.
