Transcriptional activity of CCAAT/enhancer-binding proteins is controlled by a conserved inhibitory domain that is a target for sumoylation

CCAAT/enhancer-binding proteins (C/EBPs) are basic region/leucine zipper transcription factors that function as regulators of cell growth and differentiation in numerous cell types. We previously localized transcriptional activation and inhibitory regions in one family member, C/EBPepsilon. Here we describe the further characterization of a C/EBPepsilon inhibitory domain termed regulatory domain 1. We show that functionally related domains are present in C/EBPalpha, C/EBPbeta, and C/EBPdelta. These domains contain an evolutionarily conserved five-amino acid motif (the regulatory domain motif (RDM)) that conforms to the consensus sequence (I/V/ L)KXEP. Mutagenesis studies revealed that the residues at positions 1, 2, and 4 of the RDM are critical for inhibitory domain function. Data base searches identified RDM-like sequences in a number of nuclear proteins. We found that small regions from c-Jun, JunB, and JunD containing this sequence also function as transcriptional inhibitory domains. Importantly, the RDM is similar to the recognition sequence for attachment of the ubiquitin-like protein, small ubiquitin-like modifier-1 (SUMO-1), and the conserved lysine residue of each C/EBP RDM served as an attachment site for SUMO-1. SUMO-1 attachment decreased the inhibitory effect of the C/EBPepsilon regulatory domain, suggesting that sumoylation may play an important role in modulating C/EBPepsilon activity as well as that of the other C/EBP family members.