E-proteins are bHLH transcription factors with a very broad tissue distribution. Activation of a target gene by E-proteins usually is achieved by formation of a heterodimeric complex with other, tissue specific bHLH transcription factors. Common partners are the MyoD family members in muscle and achaete-scute family members in neurons. In contrast to vertebrates where at least three E-protein members have been identified, a single member is present in C.elegans called CeE/DA (the product of
hlh-2) (Krause et al., 1997). CeE/DA shows highest identity to the single E-protein present in the fly, the product of the daughterless gene. Surprisingly, CeE/DA is not a favoured heterodimer partner for CeMyoD (
hlh-1), as shown by in vitro binding studies and non-overlapping reporter construct expression and immunostainning patterns for the two proteins (Krause et al., 1997). However, other bHLH transcription factors in C.elegans appear to interact with CeE/DA. We have focused our studies on two such factors; HLH-3, an achaete-scute family member (several other members have also been described in C.elegans) and the recently described homologue of Drosophila Twist, CeTwist (the product of
hlh-8) (Harfe et al., 1998). To investigate developmental processes affected by the various bHLH heterodimeric complexes, we have used both in vitro gel-shift binding studies and in vivo approaches, including immunostainning and reporter constructs. In gel-shift studies, we have observed the formation of homo- and heterodimers and binding to a cannonical E-box, CAGGTG. Homodimers of CeTwist also bind to the "NdE-box" (CATATG), a cis-acting regulatory sequence identified upstream of
egl-15 and
ceh-24 (an FGF receptor and NK-2 class homeodomain, respectively). Formation of heterodimers with CeE/DA, as opposed to homodimers, seems to be favoured by CeTwist. Moreover, simultaneous overexpression of CeE/DA and CeTwist can drive expression from promoter elements of these two putative target genes at very high levels compared with expression of either factor alone (Harfe et al., 1998). We have also shown that a deletion of the basic region of CeE/DA can act as a dominant negative in gel shift binding assays and we are exploring in vivo functions with this mutant form in transgenic animals. Implications for developmental processes shall be discussed further. REFERENCES Harfe, B. & Fire, A. (1998), Development 125, 421-429 Harfe, B., Vaz Gomes, A., Kenyon, C., Krause, M. & Fire, A. (1998) Genes and Development (in press) Krause, M., Park., M., Zhang,J.-M., Yuan, J., Harfe, Xu, S.-Q., Greenwald, I., Cole, M., Paterson, B. & Fire, A. (1997), Development 124, 2179-2189