Worm Breeder's Gazette 15(3): 18 (June 1, 1998)
These abstracts should not be cited in bibliographies. Material contained herein should be treated as personal communication and should be cited as such only with the consent of the author.
Dept. of Cell Biology, Biozentrum, University of Basel, Klingelbergstrasse 70, CH-4056 BASEL, Switzerland
The transcription factor Ceh-14 is a member of the LIM class of homeobox genes, which contain two tandem LIM domains and a homeodomain. It has been shown that the LIM domains (zinc binding structures similar to Zinc fingers) are preventing the homeodomain from binding DNA. The LIM domains themselves are protein-protein interaction domains. Proteins specifically binding LIM domains were cloned and shown to activate the DNA binding of the homeodomain (Agulnik et al. Nature 384, 1996, 270-72). The LIM interacting proteins (Ldp) are very well conserved between different animal phyla. An Ldp homologue with two splice variants has been uncovered by the C. elegans genome project (C.e. ldp-1a and 1b). To understand whether the activation of Ceh-14 is accomplished by C.e. Ldp-1 we performed a yeast two hybrid assay. Using the Ceh-14 tandem LIM domains as a bait (fused to the GAL4 DNA binding domain) we showed that both splice variants of C.e. Ldp-1 (fused to GAL4 activation domain) bind the Ceh-14 tandem LIM domains (Fig. 1). To control the specificity of the binding we changed crucial cysteine residues to serine, which is very similar sterically, but cannot bind the zinc ion and thus cannot participate in forming the ternary complex with the other three residues. The metal binding is essential for the tertiary structure ( Kosa et al. Biochemistry 33, 1994, 468-77). Point mutations of this type in either LIM domain abolish the interaction with Ldp-1 (Fig. 1). Furthermore we showed that the interaction between LIM and Ldp's is conserved between C. elegans and vertebrates since Xenopus Ldp (clone kindly provided by Dr. Alan Agulnik) can also specifically bind Ceh-14 (Fig. 1). We are now examining the expression pattern of C.e. ldp-1 using promotor-gfp constructs. Although detailed identification of the cells expressing C.e. ldp-1 remains to be done, we observe coexpression of ceh-14 and ldp-1. Furthermore ldp-1 is expressed in most neuronal and non-neuronal cells in which other LIM homeobox genes are known to be expressed.