title: Src triggers circular ruffling and macropinocytosis at the apical surface of polarized MDCK cells.
abstract: We addressed the role of Src on cortical actin dynamics and polarized endocytosis in MDCK cells harboring a thermosensitive v-src mutant. Shifting monolayers established at 40 degrees C (non-permissive temperature) to 34 degrees C (permissive temperature) rapidly reactivated v-Src kinase, but tight junctions and cell polarity resisted for >6 h. At this interval, activated v-src was recruited on apical vesicles, induced cortactin-associated apical circular ruffles productive of macropinosomes, thereby accelerating apical pinocytosis by approximately fivefold. Ruffling and macropinosome formation were selectively abrogated by inhibitors of actin polymerization, phosphoinositide 3-kinase, phospholipase C, and phospholipase D, which all returned apical pinocytosis to the level observed at 40 degrees C, underscoring the distinct control of apical micropinocytosis and macropinocytosis. Src promoted microtubule-dependent fusion of macropinosomes to the apical recycling endosome (ARE), causing its strong vacuolation. However, preservation of tubulation and apical polarity indicated that its function was not affected. The ARE was labeled for v-src, Rab11, and rabankyrin-5 but not early endosome antigen 1, thus distinguishing two separate Rab5-dependent apical pathways. The mechanisms of Src-induced apical ruffling and macropinocytosis could shed light on the triggered apical enteroinvasive pathogens entry and on the apical differentiation of osteoclasts.
authors.affiliation: CELL Unit, Université catholique de Louvain and Christian de Duve Institute of Cellular Pathology, 1200 Brussels, Belgium.
authors.forename: Marcel
authors.initials: M
authors.lastname: Mettlen
authors.forename: Anna
authors.initials: A
authors.lastname: Platek
authors.forename: Patrick
authors.initials: P
authors.lastname: Van Der Smissen
authors.forename: Sarah
authors.initials: S
authors.lastname: Carpentier
authors.forename: Mustapha
authors.initials: M
authors.lastname: Amyere
authors.forename: Letizia
authors.initials: L
authors.lastname: Lanzetti
authors.forename: Philippe
authors.initials: P
authors.lastname: de Diesbach
authors.forename: Donatienne
authors.initials: D
authors.lastname: Tyteca
authors.forename: Pierre J
authors.initials: PJ
authors.lastname: Courtoy
mesh_terms: D000818:Animals; D002460:Cell Line; D004285:Dogs; D011992:Endosomes / Q000378:metabolism; D015688:Oncogene Protein pp60(v-src) / Q000502:physiology*; D010873:Pinocytosis / Q000502:physiology*; D014617:Vacuoles / Q000378:metabolism
chemical_list: D015688:Oncogene Protein pp60(v-src)