Trafficking of galectin-3 through endosomal organelles of polarized and non-polarized cells

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SummaryThe role of post-translational tubulin modifications in the development and maintenance of a polarized epithelium is not well understood. We studied the balance between detyrosinated (detyr-) and tyrosinated (tyr-) tubulin in the formation of MDCK cell monolayers. Increased quantities of detyrosinated microtubules were detected during assembly into confluent cell sheets. These tubules were composed of alternating stretches of detyr-and tyr-tubulin. Constant induction of tubulin tyrosination, which decreased the levels of detyr-tubulin by overexpression of tubulin tyrosine ligase (TTL), disrupted monolayer establishment. Detyr-tubulin-depleted cells assembled into isolated islands and developed a prematurely polarized architecture. Thus, tubulin detyrosination is required for the morphological differentiation from non-polarized cells into an epithelial monolayer. Moreover, membrane trafficking, in particular to the apical domain, was slowed down in TTL-overexpressing cells. This effect could be reversed by TTL knockdown, which suggests that detyr-tubulin-enriched microtubules serve as cytoskeletal tracks to guide membrane cargo in polarized MDCK cells.

Section: Distribution Of Detyrosinated and Tyrosinated Tubulin In Polsupporting

confidence: 65%

Section: Microscopymentioning

confidence: 99%

Tubulin detyrosination promotes monolayer formation and apical trafficking in epithelial cells

Zink

Große

Freikamp

et al. 2012

Self Cite

“…Internalized Gal3 interaction with glycosylated cargo in apical recycling endosomes sorts glycoproteins to the apical membrane (Delacour et al, 2007;Schneider et al, 2010). Similarly, in the absence of the basolateral sorting adaptor AP-1B, Gal4 promotes transcytosis and apical sorting of the transferrin receptor through apical recycling endosomes (Perez Bay et al, 2014).…”

Section: Galectin Traffickingmentioning

confidence: 99%

“…In polarized MDCK cells, Gal3 is endocytosed at the apical membrane following activation of a raft-dependent pathway to recycling endosomes (Schneider et al, 2010;Straube et al, 2013). Internalized Gal3 interaction with glycosylated cargo in apical recycling endosomes sorts glycoproteins to the apical membrane (Delacour et al, 2007;Schneider et al, 2010).…”

Section: Galectin Traffickingmentioning

confidence: 99%

The galectin lattice at a glance

Nabi

Shankar

Dennis

2015

276

230

Galectins are a family of widely expressed β-galactoside-binding lectins in metazoans. The 15 mammalian galectins have either one or two conserved carbohydrate recognition domains (CRDs), with galectin-3 being able to pentamerize; they form complexes that crosslink glycosylated ligands to form a dynamic lattice. The galectin lattice regulates the diffusion, compartmentalization and endocytosis of plasma membrane glycoproteins and glycolipids. The galectin lattice also regulates the selection, activation and arrest of T cells, receptor kinase signaling and the functionality of membrane receptors, including the glucagon receptor, glucose and amino acid transporters, cadherins and integrins. The affinity of transmembrane glycoproteins to the galectin lattice is proportional to the number and branching of their N-glycans; with branching being mediated by Golgi N-acetylglucosaminyltransferasebranching enzymes and the supply of UDP-GlcNAc through metabolite flux through the hexosamine biosynthesis pathway. The relative affinities of glycoproteins for the galectin lattice depend on the activities of the Golgi enzymes that generate the epitopes of their ligands and, thus, provide a means to analyze biological function of lectins and of the 'glycome' more broadly.

“…One potential sorting receptor is galectin-3, which binds to galactose and utilizes alternative pathways independent of the passage through the ER and the Golgi (Boscher et al, 2011;Hughes, 1999). Galectin-3 accumulates in acidified subapical endosomal compartments but not in the TGN of COS-1 and MDCK cells (Schneider et al, 2010). It was also shown that galectin-3 is secreted apically in filter-grown MDCK cells (Lindstedt et al, 1993;Schneider et al, 2010) and that depletion of galectin-3 from MDCK cells results in missorting of apical membrane proteins, such as lactose-phlorizin hydrolase (LPH), p75 and gp114 (Delacour et al, 2006).…”

Section: Possible Mechanism Of the Apical Secretion Of Wnt11mentioning

confidence: 99%

Apicobasal secretion of Wnt11 and Wnt3a in polarized epithelial cells is regulated by distinct mechanisms

Yamamoto

Awada

Hanaki

et al. 2013

SummaryWnts are glycan-and lipid-modified morphogens that are important for cellular responses, but how Wnts are secreted in polarized epithelial cells remains unclear. Although Wntless (Wls) has been shown to interact with Wnts and support their secretion, the role of Wls in the sorting of Wnts to the final destination in polarized epithelial cells have not been clarified. Glycosylation was shown to be important for the sorting of some transmembrane and secreted proteins, but glycan profiles and their roles in the polarized secretion of Wnts has not yet been demonstrated. Here we show the apical and basolateral secretion of Wnts is regulated by different mechanisms. Wnt11 and Wnt3a were secreted apically and basolaterally, respectively, in polarized epithelial cells. Wls was localized to the basolateral membrane. Mass-spectrometric analyses revealed that Wnt11 is modified with complex/hybrid(Asn40)-, highmannose(Asn90)-and high-mannose/hybrid(Asn300)-type glycans and that Wnt3a is modified with two high-mannose-type glycans (Asn87 and Asn298). Glycosylation processing at Asn40 and galectin-3 were required for the apical secretion of Wnt11, whereas clathrin and adaptor protein-1 were required for the basolateral secretion of Wnt3a. By the fusion of the Asn40 glycosylation site of Wnt11, Wnt3a was secreted apically. The recycling of Wls by AP-2 was necessary for the basolateral secretion of Wnt3a but not for the apical secretion of Wnt11. These results suggest that Wls has different roles in the polarized secretion of Wnt11 and Wnt3a and that glycosylation processing of Wnts decides their secretory routes.