EVOLUTIONARY RELATIONSHIPS AND FUNCTIONAL CONSERVATION AMONG VERTEBRATE MAX-ASSOCIATED PROTEINS - THE ZEBRA FISH HOMOLOG OF MXI1

In mammals, current evidence supports the view that Myc-responsive act ivities are regulated in part through an intracellular balance between levels of transcriptionally-active Myc/Max heterodimers and those of transcriptionally-inert Max/Max, Mad/Max-and Mxil/Max complexes. To ga in insight into the roles of Mad and Mxi1 in cellular growth and diffe rentiation and to fortify key structure-function relationships from an evolutionary standpoint, low stringency hybridization screens were us ed to identify potential homologs of these Max-associated proteins in the zebra fish genome. A single class of cDNA clones that cross-hybrid ized both to human mad and mxi1 probes was shown to encode a putative protein with significantly greater homology to mammalian Mxi1 than to Mad, particularly in the basic and helix-loop-helix (bHLH) regions. Th e high degree of structural relatedness between vertebrate Mxi1 protei ns apparent in molecular;modelling studies was consistent with the fin dings that the HLH/leucine zipper (LZ) region of zMxi1 exhibited the s ame profile of. dimerization specificities as its mammalian counterpar t in the two-hybrid system and that zmxi1 could, like human mxi1 (Laho z et al., 1994), suppress the oncogenic potential of mouse c-myc in a mammalian cell. Finally, a comparison of steady-state zc-myc and zmxi1 mRNA levels during zebra fish embryogenesis demonstrated (i) high lev els of zc-myc relative to zmxi1 mRNA during initiation of organogenesi s, a period characterized by intense growth and active differentiation and (ii) rising levels of zmxi1 mRNA during progression towards the t erminally differentiated state. These contrasting patterns of developm ental expression together with the capacity of zmxi1 to repress myc-in duced transformation support a model for the regulation, by Max-associ ated proteins, of Myc functions in the control of normal cell developm ent and neoplastic growth.