Mucolipin‐2 Localizes to the Arf6‐Associated Pathway and Regulates Recycling of GPI‐APs

Abstract

In mammals, the mucolipin family includes three members mucolipin-1, mucolipin-2, and mucolipin-3 (MCOLN1-3). While mutations in MCOLN1 and MCOLN3 have been associated with mucolipidosis type IV and the varitint-waddler mouse phenotype, respectively, little is known about the function and cellular distribution of MCOLN2. Here we show that MCOLN2 traffics via the Arf6-associated pathway and colocalizes with major histocompatibility protein class I (MHCI) and glycosylphosphatidylinositol-anchored proteins (GPI-APs), such as CD59 in both vesicles and long tubular structures. Expression of a constitutive active Arf6 mutant, or activation of endogenous Arf6 by transfection with EFA6 or treatment with aluminum fluoride, caused accumulation of MCOLN2 in enlarged vacuoles that also contain MHCI and CD59. In addition, overexpression of MCOLN2 promoted efficient activation of Arf6 in vivo, thus suggesting that MCOLN2 may have a role in the traffic of cargo through the Arf6-associated pathway. In support of this we found that overexpression of a MCOLN2 inactive mutant decreases recycling of CD59 to the plasma membrane. Therefore, our results indicate that MCOLN2 localizes to the Arf6-regulated pathway and regulates sorting of GPI-APs.

Heteromultimeric TRPML channel assemblies play a crucial role in the regulation of cell viability models and starvation-induced autophagy

“…Recent studies have demonstrated that the KK 3114 Journal of Cell Science 123 (18) mutations in TRPML2 and TRPML3 exert a dominant-negative inhibitory effect on the function of their respective native channels. This effect was attributed to the interactions of TRPML2-KK and TRPML3-KK with and neutralization of the activity of their corresponding native channel (Karacsonyi et al, 2007;Kim et al, 2009). In a similar manner, we hypothesized that if the TRPML(P) and TRPML(P-KK) mutants were to form channel assemblies with each other, regardless of channel identity, then the channel-inactive TRPML(P-KK) mutants should exert a dominant-negative effect on constitutively active TRPML(P) mutants by rendering the complex inactive.…”

“…Furthermore, recombinant as well as native TRPML channels also physically interact with each other in homo-and heteromultimeric combinations (CurcioMorelli et al, 2010;Venkatachalam et al, 2006;Zeevi et al, 2009). In addition, TRPML2 and TRPML3 traffic, in part, to lysosomes (Karacsonyi et al, 2007;Kim et al, 2009;Martina et al, 2009;Zeevi et al, 2009) where, like TRPML1, they also appear to regulate lysosomal function, in an as-yet-unknown manner. Indeed, gene-specific knockdown of TRPML2 or TRPML3 leads to lysosomal inclusions reminiscent of those found in MLIV patient TRPML1-null cells (Zeevi et al, 2009), suggesting that TRPMLs might dynamically interact with each other to sustain lysosomal integrity.…”

“…Although TRPMLs have shown to be distinct from each other in their primary intracellular distributions (Karacsonyi et al, 2007;Kim et al, 2009;Martina et al, 2009;Pryor et al, 2006;Vergarajauregui and Puertollano, 2006), all three channels do also colocalize with each other, in dual combinations, in a subset of intracellular vesicles (Zeevi et al, 2009). Furthermore, recombinant as well as native TRPML channels also physically interact with each other in homo-and heteromultimeric combinations (CurcioMorelli et al, 2010;Venkatachalam et al, 2006;Zeevi et al, 2009).…”

“…MLIV pathogenesis is directly linked to TRPML1 loss-of-function within the late endosomal/ lysosomal compartments where the protein is generally found (Manzoni et al, 2004;Pryor et al, 2006;Vergarajauregui and Puertollano, 2006) and, hence, several lysosomal regulatory roles have been proposed for TRPML1 in these compartments (Dong et al, 2008;Dong et al, 2009;LaPlante et al, 2006;Miedel et al, 2008;Piper and Luzio, 2004;Pryor et al, 2006;Soyombo et al, 2006;Treusch et al, 2004;Venugopal et al, 2009;Vergarajauregui et al, 2008). TRPML2 features cell surface and recycling endosomal localization and, as a result, has been implicated in recycling of glycosylphosphatidylinositol (GPI)-anchored proteins from recycling endosomes to the cell surface along the Arf6-regulated pathway (Karacsonyi et al, 2007). TRPML3 appears to play roles in the regulation of endocytosis and autophagy that correspond with localization of the protein to the cell surface and early endosomes under basal conditions, and autophagosome localization under stress conditions (Kim et al, 2009;Martina et al, 2009).…”

ERK MAP kinase-activated Arf6 trafficking directs coxsackievirus type B3 into an unproductive compartment during virus host-cell entry

“…Recently, ADP-ribosylation factor 6 (Arf6)-dependent endocytosis has been shown to traffic GPI-linked proteins (Karacsonyi et al, 2007), such as the CVB3 co-receptor DAF. Arf6-mediated endocytosis is a lipid-raft and non-raft endocytosis pathway that cycles at the plasma membrane and is involved in the recycling of major histocompatibility complex class I to and from the cell surface.…”