We are examining the initial steps that regulate the connection of the uterus and vulva during C. elegans development. The initial contact between the developing uterus and vulva is established by the anchor cell (AC), which crosses the basement membranes separating both tissues and specifically attaches to the inner descendants of the 1 o fated vulval precursor cell, P6.p, during the mid- to late L3 stage. Following attachment, the AC extends cellular processes from its basolateral surface and invades between the inner descendants of the P6.p. cell, positioning itself at the apex of the developing vulva. Using mutants lacking vulval induction, we show that underlying vulval cells trigger AC attachment. We have further discovered that AC attachment is not stimulated by 2 o fated vulval cells (the vulval cells that flank the centrally located 1 o cells), but rather appears to be specific for the 1 o vulval cells. Using a basement membrane marker, we show that isolated 1 o cells do not break down the basement membrane in the absence of the AC, suggesting that 1 o vulval cells do not stimulate attachment by removing the basement membrane. Instead we provide evidence indicating that the 1 o vulval cells secrete a signal that triggers AC invasive behavior: when the AC is placed at a distance from the 1 o vulval cells, it sends out cellular extensions toward the 1 o vulval cells that are similar to those seen during normal invasion. We further show that the competence of the AC to respond to this signal is regulated: AC attachment is either absent or delayed in the heterochronic mutant
lin-28 , which causes precocious vulval development. To understand the molecular mechanisms that regulate AC attachment and invasion, we are examining many known mutants. Through these studies we have found that the
evl-5 mutant, originally isolated in a screen for sterile mutants with everted vulvae (Seydoux et al., (1993) Dev. Biol. 157(2): 423-36), has a defect in AC attachment to the vulval epithelium. In
evl-5 animals vulval induction and AC positioning over the P6.p cell is normal; however, AC attachment either does not occur or is severely delayed. Visualization of AC behavior in this mutant revealed the extension of cellular processes from the AC toward the vulva, indicating that the AC is still attracted to the developing vulva. However, in many cases the AC processes appeared to flatten or broaden out at the basement membrane of the developing gonad, suggesting that the AC may not be able to cross this basement membrane. We are currently molecularly cloning
evl-5 .