Dynamic regulation of C/EBPs and HP-1alpha during cell differentiation

MARÍA ANDREA DESBATS; MARÍA SOLEDAD LLARRULL; LUCIANA PAOLA PRENDES; GRACIELA PIWIEN PILLIPUK

Bariloche

Simposio; ICGEB (Gene Expression and RNA processing) y EURASNET (Cell Biology, signalling and Alternative Splicing); 2007

ICGEB (Gene Expression and RNA processing) y EURASNET (Cell Biology, signalling and Alternative Splicing)

Cellular differentiation is a complex process in which a specific subset of genes is activated and the remainder genes are silenced. Gene expression patterns, which define cellular identity, are controlled at different levels, including the interaction of protein complexes with chromatin and changes in chromatin structure. It is well established that C/EBPs play a key role during diverse differentiation processes, e.g. adipogenesis. We observed by Immuno-FISH that during differentiation of 3T3-L1 preadipocytes in adipocytes, C/EBPb, C/EBPd, and C/EBPa concentrate in pericentromeric heterocromatin co-localizing with HP-1a. To elucidate the molecular mechanism that regulates such nuclear distribution, we investigated whether C/EBPb interacts with C/EBP sites present in satellite DNA. Chromatin immunoprecipitation assays showed that binding of C/EBPb to satellite DNA increased as cells differentiate into adipocytes. Surprisingly, we found that C/EBPb delocalized from pericentric heterochromatin by deletion of its N-terminal transactivation domain, suggesting that its proper nuclear distribution is also regulated by its interaction with factors enriched in heterochromatin. Therefore, we investigated whether C/EBPb is recruited to heterocromatic domains by HP-1a. C/EBPb co-immunoprecipitated with HP-1a, and GST pull down assays showed their direct interaction. Importantly, decondensation of heterochromatin upon treatment with deacetilase inhibitors, such as Trichostatin A or sodium butirate, delocalized both C/EBPb and HP-1a from pericentromeric domains, suggesting that integrity of chromatin structure is required for the proper subnuclear localization of C/EBPb. Moreover, HP-1a restrains C/EBPb transcriptional capacity in a concentration dependent manner, raising the possibility that C/EBPb may recruit HP-1a to promoter regions for silencing target genes during cell differentiation. In summary, upon induction of adipogenesis C/EBPb localizes in heterochromatin driven by its binding to satellite DNA, interaction with HP-1a and the integrity of chromatin structure. Importantly, C/EBPb-HP-1 complexes may participate in the control of C/EBP target genes during cell differentiation.