Figure 4. Anchor Cell Expressed VAB-10A/ Plakin Is Required for BM-BM Adhesion(A) An animal treated with
vab-10a RNAi shows a lack of adhesion between the gonadal and ventral BMs (laminin::GFP, green, top; white, bottom) under the AC (mCherry::PLCdPH, magenta) prior to invasion (red arrowheads delineate BM under the AC, distance between BMs = 0.61 +/- 0.01, n = 9, p< 0.0005 compared to wild-type, Student's t test). (B) A ventral perspective of laminin::GFP in a wildtype (top) and
vab-10a RNAi treated animal (bottom) shows delayed ventral BM breach afterreduction of
vab-10a (indicated by increased signal under AC, see Figure 3A for schematic; 19/23 animals with 1-5
mm2 gonadal BM holes fail to breach ventral BM, p < 0.005 compared to wildtype, Fisher's exact test). (C) GFP inserted in frame in the
vab-10a genomic locus revealed that VAB-10A::GFP accumulates at the AC-BM interface (arrowhead, fluorescence top, overlaid on differential interference contrast [DIC], bottom, AC outlined with dashed line), and the site of epidermal-cuticle contact (arrow). (D) A DIC image overlaid with a marker for the AC (mCherry::PLCdPH, magenta) shows a
vab10(e698) mutant (top) that failed to invade (BM is not breached as indicated by intact phase-dense line, arrowhead). Expression of VAB-10A in the AC (
cdh-3 >
vab-10a::GFP, inset) rescued the invasion defect (arrowhead indicates BM breach; n = 41/44 animals with complete invasion). (E) Hemicentin punctae (green) are contained within the region of AC-BM contact (outlined by dotted line, AC in magenta). Following treatment with RNAi targeting
vab-10a, the hemicentin punctae were no longer contained within the AC footprint (bottom, arrowheads). (F) The graph quantifies the percentage of punctae in wild-type and
vab-10 RNAi treated animals outside the AC footprint (*p < 0.05, Fisher's exact test, R100 animals for each treatment, error bars show 95% confidence interval with a continuity correction). Scale bars, 5 um.