UNC-6 (netrin) stabilizes oscillatory clustering of the UNC-40 (DCC) receptor to orient polarity.
Bottom Line: By performing live-cell imaging of the DCC orthologue UNC-40 during anchor cell invasion in Caenorhabditis elegans, we have found that UNC-40 clusters, recruits F-actin effectors, and generates F-actin in the absence of UNC-6 (netrin).Together, our data suggest that UNC-6 (netrin) directs polarized responses by stabilizing UNC-40 clustering.We propose that ligand-independent UNC-40 clustering provides a robust and adaptable mechanism to polarize toward netrin.
Affiliation: Department of Biology, Duke University, Durham, NC 27708.Show MeSH
Related in: MedlinePlus
License 1 - License 2
Mentions: To explore whether MADD-2 might mediate UNC-6–independent UNC-40 clustering, we first examined MADD-2 localization. Consistent with direct regulation of UNC-40, MADD-2::GFP was concentrated at regions of the cell where UNC-40 localizes: at the invasive cell membrane in wild-type animals and with ectopic patches of F-actin in unc-6 mutants (Fig. 7 A). To determine whether MADD-2 regulates UNC-6–independent UNC-40 clustering, we examined F-actin dynamics in unc-6 mutants harboring the putative madd-2– allele madd-2(ok2226) (Alexander et al., 2010). In madd-2(ok2226); unc-6(ev400) double mutant animals, dominant F-actin clusters still formed and were localized in random regions in the cell (Fig. 7, B and C). Furthermore, the F-actin patches reached a peak volume comparable to that observed in unc-6 mutants and disassembled at a similar rate (Table 1; Fig. 7, C–E). In madd-2(ok2226); unc-6(ev400) mutants, UNC-40 still colocalized with ectopic F-actin clusters (Fig. 7 F). Strikingly, however, madd-2(ok2226); unc-6(ev400) animals had a threefold decrease in the rate of cluster assembly (Fig. 7, C–E; and Table 1). In addition, the F-actin patches had a significantly longer lifetime before disassembly (>80 min versus ∼26 min in unc-6 mutants alone; Table 1). These results indicate that MADD-2 promotes UNC-6–independent UNC-40 cluster assembly. In addition, these observations suggest that the negative feedback mechanism that triggers cluster disassembly is induced at peak UNC-40 clustering.
Affiliation: Department of Biology, Duke University, Durham, NC 27708.