Molecular Mechanisms of C/EBP Alpha Mediated Growth Arrest

Cincinnati Childrens Hosp Med Ctr, Cincinnati OH

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Abstract

ABSTRACT Biology of the liver is regulated by a complex network of many signal transduction pathways. The understanding of molecular basis of biological processes in the liver is highly significant for the support of healthy life in humans. This renewal of R01 application investigates molecular mechanisms of liver growth and differentiation, liver regeneration after surgical resections and liver recovery after chemical injuries. Two members of C/EBP family, C/EBPa and C/EBPb, are critical regulators of liver biology and liver functions. In the course of previous studies, we found that phosphorylation of C/EBPa at S193 increases its hetero- dimerization with C/EBPb and subsequent interactions of the C/EBPa/b dimers with chromatin remodeling proteins p300 and HDAC1. While C/EBP-p300 complexes activate promoters of target genes, C/EBP-HDAC1 complexes mainly repress the promoters. Therefore, amounts of C/EBPa/b-p300 and C/EBPa/b-HDAC1 complexes are changed during distinct biological processes in appropriate ratios. The main hypothesis of this application is that the network of C/EBP proteins and chromatin remodeling proteins regulates post-natal liver growth and differentiation, liver regeneration after surgical resections and liver injury after CCl4 treatments. We generated 4 genetically modified mouse models: C/EBPa-S193D mice (phosphor- mimicking mutation), C/EBPa-S193A mice (no phosphorylation on S193), transgenic mice which express dominant negative p300 (dnp300) and double knockin-transgenic S193D-dnp300 mice. We found that C/EBPa-S193A mutant does not interact with HDAC1; however, S193D mutation increases interactions of C/EBPa with HDAC1. Our data show that proliferation of the liver is inhibited in S193D mice, while liver proliferation is increased in S193A mice. Specific Aim 1 will test the hypothesis that a balance of C/EBPa/b- p300 and C/EBPa/b-HDAC1 complexes is required for normal post-natal liver development. Specific Aim 2 will test the hypothesis that C/EBPa controls entry of hepatocytes into cell cycle and termination of liver regeneration after partial hepatectomy (PH) through interactions with HDAC1. Specific Aim 3 will examine the hypothesis that severe liver injury in young S193D and old WT mice after treatments with CCl4 is caused by C/EBPa/b-HDAC1-mediated repression of telomere reverse transcriptase, TERT. This aim is based on the observations that a) CCl4 causes severe liver damage and increased apoptosis in young S193D and old WT mice (both have high levels of C/EBPa-HDAC1 complexes); b) that basal levels of TERT are reduced in young S193D mice and in old WT mice; and c) that the TERT promoter contains binding sites for C/EBP proteins. In summary, the elucidation of the mechanisms of liver proliferation during post-natal development and after surgical resections as well as mechanisms of liver recovery after drug-mediated injury will have high impact on the field of liver biology and will provide a step for the further studies in clinical settings.