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A key step of epithelial morphogenesis is the creation of the lumen. Luminogenesis by hollowing proceeds through the fusion of apical vesicles at cell-cell contacts. The small nascent lumens grow through extension, coalescence and enlargement, coordinated with cell division, to give rise to a single central lumen. Here, by using MDCK cells grown in 3D-culture, we show that EFA6A (also known as PSD) participates in luminogenesis. EFA6A recruits α-actinin 1 (ACTN1) through direct binding. In polarized cells, ACTN1 was found to be enriched at the tight junction where it acts as a primary effector of EFA6A for normal luminogenesis. Both proteins are essential for the lumen extension and enlargement, where they mediate their effect by regulating the cortical acto-myosin contractility. Finally, ACTN1 was also found to act as an effector for the isoform EFA6B (also known as PSD4) in the human mammary tumoral MCF7 cell line. EFA6B restored the glandular morphology of this tumoral cell line in an ACTN1-dependent manner. Thus, we identified new regulators of cyst luminogenesis essential for the proper maturation of a newly-formed lumen into a single central lumen.

β2-adrenergic receptor (β2AR), a member of the GPCR (G-Proteins Coupled Receptor) family, is internalized in a ligand- and β-arrestin-dependent manner into early endosomes, and subsequently recycled back to the plasma membrane. Here we report that β-arrestin promotes the activation of the small G protein Arf6, which regulates the recycling and degradation of β2AR. We demonstrate in vitro that the C-terminal region of β-arrestin1 interacted directly and simultaneously with Arf6GDP and its specific exchange factor EFA6, to promote Arf6 activation. Similarly, the ligand-mediated activation of β2AR leads to the formation of Arf6GTP in vivo in a β-arrestin-dependent manner. Expression of either EFA6 or an activated Arf6 mutant caused accumulation of β2AR in the degradation pathway. And this phenotype could be rescued by the expression of an activated mutant of Rab4, suggesting that Arf6 acts upstream of Rab4. We propose a model in which Arf6 plays an essential role for the β2AR desensitization. The ligand-mediated stimulation of β2AR relocates β-arrestin to the plasma membrane, and triggers the activation of Arf6 by EFA6. The activated Arf6 leads to accumulation of β2AR to the degradation pathway, and negatively controls the Rab4-dependent fast recycling to prevent the re-sensitization of β2AR.