The 6 touch receptor neurons in C. elegans are AVM, ALML ALMR, PVM, PLML and PLMR (Chalfie and Sulston, Dev. Biol. 82:358). Several genes are known to affect the generation, differentiation, and function of these cells. Mutants of the gene
mec-7 lack the large microtubules (15 protofilaments), a specific feature of the touch cells, and this results in touch insensitivity. Since normal sized microtubules (11 protofilaments) are still produced in these cells, as well as in all other cells of
mec-7 animals, the
mec-7 , -tubulin is therefore specifically required for the generation of large microtubules. A likely possibility is that
mec-7 expression is confined to the touch cells. In order to test this hypothesis, we brought a reporter gene under the control of the
mec-7 promoter and generated transgenic animals. Using Andy Fire's Ppd16.51 vector, we placed the lacZ coding region under the control of a 1.65 kb fragment that includes 0.85 kb upstream of the
mec-7 start codon and 0.80 kb of the coding region. This plasmid, named pNW115, was coinjected with pRF4, a marker plasmid carrying a dominant mutation of the
rol-6 gene ( Mello et al., WBG 11( 1):18). Transgenic rollers from the F1 were stained with X-Gal and expression of the
mec-7-lacZ was found in all of the touch cells. Except for some occasional staining in a lateral pair of neurons just posterior to the pharynx (tentatively identified as FLP neurons), no other cells seem to express
mec-7. Since pNW115 lacks the SV40 nuclear localization sequence, staining was sometimes observed along the axons, helping to confirm the identity of the neurons. The roller and -Gal staining phenotypes were found to be unstable in the subsequent generations (each roller yielding ~50% rollers), implying that the exogenous DNA is maintained as an extrachromosomal element. This element, named evEX1, is likely to contain copies of both plasmids in a tandem array. As a consequence of this instability, each transgenic animal can be assumed to be a mosaic of cells containing evEX1 and of cells lacking it. Indeed, in most animals, only a fraction of the touch cells are stained. Among adult animals exhibiting at least one stained neuron, the percentages of stained cells of each type were: AVM: 30%; ALML or ALMR: 67%; simultaneous staining of both ALMs: 24%; PVM: 47% PLML or PLMR: 79%; simultaneous staining of both PLMs: 34%. FLP neurons stained at a frequency of 3%. Expression was observed at all larval stages and in embryos too.
unc-86 lineage mutants lack several neurons including the six touch cells and the FLPs. We genetically transferred evEX1 in
unc-86 animals and found no expression of
mec-7. When passed back into the wild type, this element re-expressed lac-Z in the touch neurons. The
mec-3 gene product, which is required for the maturation of the touch cells, is expressed only in these cells (with the exception of the FLPs and the PVDs) (Way and Chalfie, Genes & Dev. 3:1823), and is likely to be a transcription factor. Therefore, an attractive assumption is that
mec-3 directly activates the
mec-7 gene. We transferred evEX1 into
mec-3 mutants and surprisingly, we found expression of
mec-7 at almost normal levels in larvae, but at a lower level in adults. Because of these results and because expression of
mec-7 was not observed in PVD neurons, we conclude that
mec-3 is not required to turn on
mec-7, but is needed (directly or indirectly) for its maintenance in the touch cells. With the goal of identifying the activator(s) of
mec-7, we are currently studying its expression in other mutants affecting the generation and the maturation of the touch neurons.