Nuclear fallout provides a new link between aPKC and polarized cell trafficking

Abstract

BackgroundCell polarity, essential for cell physiology and tissue coherence, emerges as a consequence of asymmetric localization of protein complexes and directional trafficking of cellular components. Although molecules required in both processes are well known their relationship is still poorly understood.ResultsHere we show a molecular link between Nuclear Fallout (Nuf), an adaptor of Rab11-GTPase to the microtubule motor proteins during Recycling Endosome (RE) trafficking, and aPKC, a pivotal kinase in the regulation of cell polarity. We demonstrate that aPKC phosphorylates Nuf modifying its subcellular distribution. Accordingly, in aPKC mutants Nuf and Rab11 accumulate apically indicating altered RE delivery. We show that aPKC localization in the apico-lateral cortex is dynamic. When we block exocytosis, by means of exocyst-sec mutants, aPKC accumulates inside the cells. Moreover, apical aPKC concentration is reduced in nuf mutants, suggesting aPKC levels are maintained by recycling.ConclusionsWe demonstrate that active aPKC interacts with Nuf, phosphorylating it and, as a result, modifying its subcellular distribution. We propose a regulatory loop by which Nuf promotes aPKC apical recycling until sufficient levels of active aPKC are reached. Thus, we provide a novel link between cell polarity regulation and traffic control in epithelia.Electronic supplementary materialThe online version of this article (doi:10.1186/s12915-016-0253-6) contains supplementary material, which is available to authorized users.

Nuf and Rip11 requirement for polarity determinant recycling duringDrosophiladevelopment

“…3H), mimicking the effect of the expression of these phosphorylation mutant versions of Nuf on aPKC distribution in the imaginal disc cells. 15 Our data and that of others, strongly suggest that Rip11 is involved mainly in the recycling of the AJ determinant E-Cad. In Rip11 mutants other polarity determinants such as Crb, Par3 or aPKC are also affected but to a lesser or similar extent than DE-Cad.…”

“…We used the following stocks: Oregon R (wild-type), nuf, 1, 25 UAS-Myc-Nuf S155A and UAS-Myc-Nuf S155D . 15 Null embryos for rip11 were generated by inducing germ line clones (GLC) in hsFLP::FRTG19:: ovo 1-18 / rip11 KG02485 :: FRT19A 29 females, which were heat shocked on second larval instar for 1 hour at 37 C and mated to Oregon R males. nuf embryos were obtained from nuf 1 homozygous females mated to Oregon R males.…”

“…Nuf belongs to the Rab11-family interacting proteins (Rab11-FIP 16 ) and is the Drosophila homolog of FIP3, which functions as Rab11 adaptor to the microtubule motor protein kinesin and the dynein complex. 15,17 We have shown that aPKC directly interacts with and phosphorylates Nuf. Remarkably aPKC phosphorylation of Nuf modifies Nuf sub-cellular localization: a nonphosphorylatable version of Nuf accumulates at the apico-lateral cortex, while a phospho-mimetic protein, although also apically accumulated, avoids this lateral domain.…”

“…Thus the analysis of the nuf and Rip11 mutants corroborates the requirement of the Rab11-RE in the maintenance of apical cell polarity 6 and also suggests a different role of the 2 family of FIPs in the control of the cargos transported by the RE, as described in vertebrates. 31 aPKC maintenance in the membrane is heavily affected in nuf mutant embryos indicating that, as in the imaginal discs, 15 aPKC is recycled in the embryonic epidermis and this recycling is dependent on Nuf. In fact, the overexpression of the non-phosphorylatable version of Nuf (Nuf S155A 15 ) in the embryo enhances the levels of aPKC in the membrane without affecting other apical membrane markers such as DE-Cad (Fig.…”

“…31 aPKC maintenance in the membrane is heavily affected in nuf mutant embryos indicating that, as in the imaginal discs, 15 aPKC is recycled in the embryonic epidermis and this recycling is dependent on Nuf. In fact, the overexpression of the non-phosphorylatable version of Nuf (Nuf S155A 15 ) in the embryo enhances the levels of aPKC in the membrane without affecting other apical membrane markers such as DE-Cad (Fig. 3G) while the non-phosphorylatable version (Nuf S155D) has no effect ( Fig.…”