Mmip-2, a novel RING finger protein that interacts with mad members of theMyc oncoprotein network

Mad proteins are basic-helix-loop - helix -leucine zipper (bHLH-ZIP)-contai ning members of the myc oncoprotein network. They interact with the bHLH-ZI P protein max, compete for the same DNA binding sites as myc-max heterodime rs and down-regulate myc-responsive genes. Using the bHLH-ZIP domain of mad 1 as a yeast two-hybrid 'bait', we identified Mmip-2, a novel RING finger p rotein that interacts with all mad members, but weakly or not at all with c -myc, max or unrelated bHLH or bZIP proteins. The mad1-Mmip-2 interaction i s mediated by the ZIP domain in the former protein and by at least two regi ons in the latter which do not include the RING finger. Mmip-2 can disrupt max-mad DNA binding and can reverse the suppressive effects of mad proteins on c-myc-responsive target genes and on c-myc+ras-mediated focus formation in fibroblasts. Tagging with spectral variants of green fluorescent protei n showed that Mmip-2 and mad proteins reside in separate cytoplasmic and nu clear compartments, respectively. When co-expressed, however, the proteins interact and translocate to the cellular compartment occupied by the more a bundant protein. These observations suggest a novel way by which Mmip-2 can modulate the transcriptional activity of myc oncoproteins.