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PI(3,5)P2 controls endosomal branched actin dynamics by regulating cortactin-actin interactions.

Hong NH, Qi A, Weaver AM - J. Cell Biol. (2015)

Bottom Line: These findings suggest that PI(3,5)P2 formation on endosomes may remove cortactin from endosome-associated branched actin.Conversely, inhibition of Arp2/3 complex activity greatly reduced cortactin localization to late endosomes.These data suggest a model in which PI(3,5)P2 binding removes cortactin from late endosomal branched actin networks and thereby promotes net actin turnover.

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Affiliation: Department of Cancer Biology, Vanderbilt University Medical Center, Nashville, TN 37232.

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Model of endosomal branched actin network regulation by PI(3,5)P2. (A) Branched actin nucleation is initiated by the WASH complex which is bound to the surface of late endosomes by signaling molecules that likely include PI(3)P (Jia et al., 2010). WASH-induced activation of the Arp2/3 complex recruits cortactin to nascent branch points (Fig. 4), where it synergistically promotes actin assembly. (B) Conversion of PI(3)P to PI(3,5)P2 is accomplished by the enzymatic activity of PIKfyve within the three-member PIKfyve complex that includes the scaffold protein Vac14 and the opposing 5′ phosphatase Fig. 4 (Shisheva, 2008; Dove et al., 2009). (C) PI(3,5)P2 binds to cortactin, releasing actin filaments (Fig. 5) and potentially the WASH complex from endosomes (Fig. 8). (D) Without cortactin binding to the branchpoint, Arp2/3 complex loses affinity for the mother filament (Uruno et al., 2001) causing debranching (Weaver et al., 2001; Fig. 6). Disassembly of branched actin networks leads to diminished recruitment of cortactin. (E) Conversion of PI(3,5)P2 back to PI(3)P should release cortactin from the endosome surface unless new branched actin networks are available for rebinding. Interconversion of PI(3)P and PI(3,5)P2 by the PIKfyve complex may facilitate dynamic cycling of actin assembly and disassembly through control of cortactin–actin interactions.
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fig9: Model of endosomal branched actin network regulation by PI(3,5)P2. (A) Branched actin nucleation is initiated by the WASH complex which is bound to the surface of late endosomes by signaling molecules that likely include PI(3)P (Jia et al., 2010). WASH-induced activation of the Arp2/3 complex recruits cortactin to nascent branch points (Fig. 4), where it synergistically promotes actin assembly. (B) Conversion of PI(3)P to PI(3,5)P2 is accomplished by the enzymatic activity of PIKfyve within the three-member PIKfyve complex that includes the scaffold protein Vac14 and the opposing 5′ phosphatase Fig. 4 (Shisheva, 2008; Dove et al., 2009). (C) PI(3,5)P2 binds to cortactin, releasing actin filaments (Fig. 5) and potentially the WASH complex from endosomes (Fig. 8). (D) Without cortactin binding to the branchpoint, Arp2/3 complex loses affinity for the mother filament (Uruno et al., 2001) causing debranching (Weaver et al., 2001; Fig. 6). Disassembly of branched actin networks leads to diminished recruitment of cortactin. (E) Conversion of PI(3,5)P2 back to PI(3)P should release cortactin from the endosome surface unless new branched actin networks are available for rebinding. Interconversion of PI(3)P and PI(3,5)P2 by the PIKfyve complex may facilitate dynamic cycling of actin assembly and disassembly through control of cortactin–actin interactions.

Mentions: Activation of Arp2/3 complex occurs upon binding of an activator in the WASP family of proteins (Millard et al., 2004). At endosomes, the WASP family member WASH appears to be the main activator of Arp2/3 complex (Derivery et al., 2009; Gomez and Billadeau, 2009; Duleh and Welch, 2010) and should act in concert with cortactin to promote endosomal branched actin assembly. Consistent with those studies, WASH, cortactin, and actin show strong colocalization on endosomal membranes (Fig. 8). The FAM21 component of WASH has been found to bind multiple phosphoinositides by dot plot, including the major endosomal lipids PI(3)P and PI(3,5)P2 (Jia et al., 2010). However, the consequence of that interaction is unclear. To assess the effect of PI(3,5)P2 on WASH localization, MDA-MB-231 and SCC61 cells were treated with YM201636 and immunostained for WASH and cortactin and colocalized with either actin (Fig. 8 A) or mRFP-Rab7 (Fig. 8 B). Inhibition of PI(3,5)P2 synthesis led to an increase in the localization of WASH to cortactin+ and actin+ structures at Rab7+ endosomes (Fig. 8), suggesting that similarly to cortactin, PI(3,5)P2 does not recruit WASH to endosomal membranes but instead may promote its removal. However, the increase in WASH localization to Rab7+ endosomes was small compared with the increase in cortactin localization in YM201636-treated cells (40% increase in WASH vs. 300% increase in cortactin colocalization with Rab7, compare median values in Figs. 8 C and 3 C). As with endogenous Rab7 (Fig. 3 E), there was no difference in mRFP-Rab7 area with YM201636 treatment (Fig. 8 D). Overall, PI(3,5)P2 plays a concerted role to remove branched actin regulators from late endosomes and promote disassembly of endosome-associated actin assemblies via cortactin (see model in Fig. 9).


PI(3,5)P2 controls endosomal branched actin dynamics by regulating cortactin-actin interactions.

Hong NH, Qi A, Weaver AM - J. Cell Biol. (2015)

Model of endosomal branched actin network regulation by PI(3,5)P2. (A) Branched actin nucleation is initiated by the WASH complex which is bound to the surface of late endosomes by signaling molecules that likely include PI(3)P (Jia et al., 2010). WASH-induced activation of the Arp2/3 complex recruits cortactin to nascent branch points (Fig. 4), where it synergistically promotes actin assembly. (B) Conversion of PI(3)P to PI(3,5)P2 is accomplished by the enzymatic activity of PIKfyve within the three-member PIKfyve complex that includes the scaffold protein Vac14 and the opposing 5′ phosphatase Fig. 4 (Shisheva, 2008; Dove et al., 2009). (C) PI(3,5)P2 binds to cortactin, releasing actin filaments (Fig. 5) and potentially the WASH complex from endosomes (Fig. 8). (D) Without cortactin binding to the branchpoint, Arp2/3 complex loses affinity for the mother filament (Uruno et al., 2001) causing debranching (Weaver et al., 2001; Fig. 6). Disassembly of branched actin networks leads to diminished recruitment of cortactin. (E) Conversion of PI(3,5)P2 back to PI(3)P should release cortactin from the endosome surface unless new branched actin networks are available for rebinding. Interconversion of PI(3)P and PI(3,5)P2 by the PIKfyve complex may facilitate dynamic cycling of actin assembly and disassembly through control of cortactin–actin interactions.
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fig9: Model of endosomal branched actin network regulation by PI(3,5)P2. (A) Branched actin nucleation is initiated by the WASH complex which is bound to the surface of late endosomes by signaling molecules that likely include PI(3)P (Jia et al., 2010). WASH-induced activation of the Arp2/3 complex recruits cortactin to nascent branch points (Fig. 4), where it synergistically promotes actin assembly. (B) Conversion of PI(3)P to PI(3,5)P2 is accomplished by the enzymatic activity of PIKfyve within the three-member PIKfyve complex that includes the scaffold protein Vac14 and the opposing 5′ phosphatase Fig. 4 (Shisheva, 2008; Dove et al., 2009). (C) PI(3,5)P2 binds to cortactin, releasing actin filaments (Fig. 5) and potentially the WASH complex from endosomes (Fig. 8). (D) Without cortactin binding to the branchpoint, Arp2/3 complex loses affinity for the mother filament (Uruno et al., 2001) causing debranching (Weaver et al., 2001; Fig. 6). Disassembly of branched actin networks leads to diminished recruitment of cortactin. (E) Conversion of PI(3,5)P2 back to PI(3)P should release cortactin from the endosome surface unless new branched actin networks are available for rebinding. Interconversion of PI(3)P and PI(3,5)P2 by the PIKfyve complex may facilitate dynamic cycling of actin assembly and disassembly through control of cortactin–actin interactions.
Mentions: Activation of Arp2/3 complex occurs upon binding of an activator in the WASP family of proteins (Millard et al., 2004). At endosomes, the WASP family member WASH appears to be the main activator of Arp2/3 complex (Derivery et al., 2009; Gomez and Billadeau, 2009; Duleh and Welch, 2010) and should act in concert with cortactin to promote endosomal branched actin assembly. Consistent with those studies, WASH, cortactin, and actin show strong colocalization on endosomal membranes (Fig. 8). The FAM21 component of WASH has been found to bind multiple phosphoinositides by dot plot, including the major endosomal lipids PI(3)P and PI(3,5)P2 (Jia et al., 2010). However, the consequence of that interaction is unclear. To assess the effect of PI(3,5)P2 on WASH localization, MDA-MB-231 and SCC61 cells were treated with YM201636 and immunostained for WASH and cortactin and colocalized with either actin (Fig. 8 A) or mRFP-Rab7 (Fig. 8 B). Inhibition of PI(3,5)P2 synthesis led to an increase in the localization of WASH to cortactin+ and actin+ structures at Rab7+ endosomes (Fig. 8), suggesting that similarly to cortactin, PI(3,5)P2 does not recruit WASH to endosomal membranes but instead may promote its removal. However, the increase in WASH localization to Rab7+ endosomes was small compared with the increase in cortactin localization in YM201636-treated cells (40% increase in WASH vs. 300% increase in cortactin colocalization with Rab7, compare median values in Figs. 8 C and 3 C). As with endogenous Rab7 (Fig. 3 E), there was no difference in mRFP-Rab7 area with YM201636 treatment (Fig. 8 D). Overall, PI(3,5)P2 plays a concerted role to remove branched actin regulators from late endosomes and promote disassembly of endosome-associated actin assemblies via cortactin (see model in Fig. 9).

Bottom Line: These findings suggest that PI(3,5)P2 formation on endosomes may remove cortactin from endosome-associated branched actin.Conversely, inhibition of Arp2/3 complex activity greatly reduced cortactin localization to late endosomes.These data suggest a model in which PI(3,5)P2 binding removes cortactin from late endosomal branched actin networks and thereby promotes net actin turnover.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Cancer Biology, Vanderbilt University Medical Center, Nashville, TN 37232.

Show MeSH
Related in: MedlinePlus