Invasion, the process during which cells cross tissue borders, needs to be tightly regulated since uncontrolled invasion can lead to metastatic cancer growth. During C. elegans larval development, a specialized cell in the somatic gonad called anchor cell (AC) breaks two basal laminae and then invades the adjacent vulval tissue in order to form a connection between the developing vulva and uterus. Multiple signals from the vulval cells and the ventral nerve cord regulate AC invasion, however, how these signals are integrated by the AC is largely unknown. We have identified
madd-2, a conserved RING finger gene and E3 ubiquitin ligase, as a regulator of AC invasion.
madd-2 has independently been identified by both the P. Roy lab as a gene acting downstream of the UNC-40 netrin receptor in controlling muscle arm extensions and the C. Bargmann & M. Tessier-Lavigne groups that showed that the same is true in axon branching. The human
madd-2 homologue, MID1, is mutated in most cases of Opitz syndrome, a disease characterized by ventral midline defects. Analysis of basal laminae breaching in
madd-2 mutants demonstrates that the timing of AC invasion is changed.
madd-2 single mutants show delayed invasion, but AC invasion in double mutants for
madd-2 and either
unc-40 or
unc-6 occurs precociously. We thus postulate that AC invasion is not only activated at the late L3 stage, when invasion normally begins, but also repressed at earlier stages. Additionally, by examining markers for the invasive membrane, we find that AC morphology is altered in
madd-2 mutants. The AC in
madd-2 mutants forms ectopic protrusions at random locations, suggesting a loss of directed invasion. RNAi mediated knock-down of the proposed GEF K07D4.7, a homolgue of human Ephexin, suppresses the ectopic AC protrusions of
madd-2 mutants but does not affect AC shape in wild-type animals. Thus, the GEF K07D4.7 might be negatively regulated by
madd-2, directly or indirectly, to prevent the formation of ectopic AC protrusions at the time of invasion.