The UNC-4 homeodomain protein functions with the UNC-37 Groucho- like co-repressor protein to prevent VA motor neurons from assuming differentiated traits normally reserved for their lineal sister cells, the VB motor neurons. In flies, Groucho binds to specific transcription factors to repress gene expression. A similar mechanism could account for the strong genetic interactions that we observe between unc-4 and unc-37 missense alleles and for the dependence of VA motor neuron differentiation on both unc-4 and unc-37 activity (1). Previous studies, however, have failed to detect physical interactions between UNC-4 and UNC-37. In these experiments, full length UNC-4 and UNC-37 were tested in vitro with recombinant proteins and in the yeast two hybrid assay (2). We have now established that a C-terminal region of UNC-4 displays strong interactions with intact UNC-37 in yeast. Systematic deletions of this region have revealed a 50 A.A. fragment that is necessary for interactions with UNC-37. This subdomain includes an UNC-4 C-terminal sequence (ELD) that is strikingly similar to an Engrailed-like repressor domain that is also found in many other classes of homeoproteins (Winnier, et al, this meeting). Moreover, specific missense mutations in the UNC-4 ELD prevent interactions with UNC-37 in yeast and perturb unc- 4 function in vivo. GFP reporters of the acr-5 and del-1 genes are normally expressed in VB but not VA motor neurons (3). Expression of acr-5::gfp and del-1::gfp is de-repressed in the VA motor neurons by unc-4 null alleles as well as by missense mutations in the UNC-4 ELD. Therefore, we propose that the UNC-4 ELD mediates physical interaction with UNC-37 to negatively regulate genes that are normally expressed in VB motor neurons. Our failure to detect physical interaction between UNC-37 and intact UNC-4 may be indicative of an intramolecular regulatory mechanism whereby an UNC-4 N-terminal region prevents the UNC-4 ELD from binding UNC-37. Interestingly, a recent report describing physical interactions between the canonical Engrailed repressor domain, eh-1, and Groucho did not include experiments using the full length Engrailed protein (4). We speculate that an additional gene product may be required to relieve the presumptive inhibitory effects of the UNC-4 N-terminus on interactions of the UNC-4 ELD with UNC-37. 1. Pflugrad, A., et al. (1997) Development 124, 1699-1709. 2. Meir, J. Y.-J. et al., (1997) C. elegans Meeting. 3. Ross, J. et al., (1997) International C. elegans Meeting. 4. Jimenez et al., (1997) Genes & Development 11.3072-3082.

Affiliation:
- Vanderbilt University Medical Center, Nashville, TN, 37232-2175