Caenorhabditis elegans embryonic elongation is driven by cell shape changes that cause a contraction of the epidermal cell layer enclosing the embryo. We have previously shown that this process requires a Rho-associated kinase (LET-502) and is opposed by the activity of a myosin phosphatase regulatory subunit (MEL-11). We now extend our characterization and show that mel-11 activity is required both in the epidermis during embryonic elongation and in the spermatheca of the adult somatic gonad. let-502 and mel-11 reporter gene constructs show reciprocal expression patterns in the embryonic epidermis and the spermatheca, and mutations of the two genes have opposite effects in these two tissues. These results are consistent with let-502 and mel-11 mediating tissue contraction and relaxation, respectively. We also find that mel-11 embryonic inviability is genetically enhanced by mutations in a Rac signaling pathway, suggesting that Rac potentiates or acts in parallel with the activity of the myosin phosphatase complex. Since Rho has been implicated in promoting cellular contraction, our results support a mechanism by which epithelial morphogenesis is regulated by the counteracting activities of Rho and Rac.

We have identified two genes associated with the hypodermal cell shape changes that occur during elongation of the Caenorhabditis elegans embryo. The first gene, called let-502, encodes a protein with high similarity to Rho-binding Ser/Thr kinases and to human myotonic dystrophy kinase (DM-kinase). Strong mutations in let-502 block embryonic elongation, and let-502 reporter constructs are expressed in hypodermal cells at the elongation stage of development. The second gene, mel-11, was identified by mutations that act as extragenic suppressors of let-502. mel-11 encodes a protein similar to the 110-to 133-kD regulatory subunits of vertebrate smooth muscle myosin-associated phosphatase (PP-IM). We suggest that the LET-502 kinase and the MEL-11 phosphatase subunit act in a pathway linking a signal generated by the small GTP-binding protein Rho to a myosin-based hypodermal contractile system that drives embryonic elongation. LET-502 may directly regulate the activity of the MEL-11 containing phosphatase complex and the similarity between LET-502 and DM-kinase suggests a similar function for DM-kinase.

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Each of 22 amino acids in the proposed alpha‐helix‐turn‐alpha‐helix operator binding motif of the Tn10 encoded Tet repressor was replaced by alanine and one residue was replaced by valine to determine their role in tet operator recognition by a ‘loss of contact’ analysis with 16 operator variants. One class of amino acids consisting of T27 and R28 in the first alpha‐helix and L41, Y42, W43 and H44 in the recognition alpha‐helix are quantitatively most important for wild‐type operator binding. These residues are probably involved in the structural architecture of the motif. A second class of residues is quantitatively less important for binding, but determines specificity by forming base pair specific contacts to three positions in tet operator. This property is most clearly demonstrated for Q38 and P39 and to a lesser extent for T40 at the N‐terminus of the recognition alpha‐helix. The contacted operator base pairs indicate that the N‐terminus of the recognition alpha‐helix is located towards the palindromic center in the repressor‐operator complex. Although the orientation of the recognition alpha‐helix in the Tet repressor‐tet operator complex is inversed as compared with the lambda‐ and 434 repressor‐operator complexes, the reduced operator binding of the TA27 mutation in the first alpha‐helix suggests that the hydrogen bonding networks connecting the two alpha‐helices may be similar in these proteins.(ABSTRACT TRUNCATED AT 250 WORDS)