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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.

View Article: PubMed Central - HTML - PubMed

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

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Recruitment of cortactin to late endosomes depends on Arp2/3 complex activity. (A) Representative images show cortactin (green), actin (red), and Rab7 (blue) localization after 2 h treatment of MDA-MB-231 cells with 800 nM YM201636 ± 200 µM CK-666. Bars, 20 µm (5 µm for magnifications). (B) Images from cells treated with YM201636 and 100 or 200 µM CK-666 were analyzed for percentage of colocalization of cortactin or actin with Rab7. The DMSO and YM201636 datasets (no CK-666) are the same data as that shown in Fig. 3 ( C and D). Data shown as box and whiskers plots with the box indicating the 25th and 75th percentiles, solid line indicating the median, and whiskers indicating the 95% confidence intervals. 3 independent experiments, n ≥ 50 cells for each condition. ****, P < 0.0001.
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fig4: Recruitment of cortactin to late endosomes depends on Arp2/3 complex activity. (A) Representative images show cortactin (green), actin (red), and Rab7 (blue) localization after 2 h treatment of MDA-MB-231 cells with 800 nM YM201636 ± 200 µM CK-666. Bars, 20 µm (5 µm for magnifications). (B) Images from cells treated with YM201636 and 100 or 200 µM CK-666 were analyzed for percentage of colocalization of cortactin or actin with Rab7. The DMSO and YM201636 datasets (no CK-666) are the same data as that shown in Fig. 3 ( C and D). Data shown as box and whiskers plots with the box indicating the 25th and 75th percentiles, solid line indicating the median, and whiskers indicating the 95% confidence intervals. 3 independent experiments, n ≥ 50 cells for each condition. ****, P < 0.0001.

Mentions: One mechanism by which cortactin could promote accumulation of actin on endosomes is by regulating branched actin network assembly by the Arp2/3 complex (Uruno et al., 2001; Weaver et al., 2001). To determine whether Arp2/3-mediated branched actin nucleation is required for recruiting cortactin to endosomal membranes, we treated cells with the Arp2/3 complex inhibitor CK-666 (Nolen et al., 2009; Hetrick et al., 2013). Treatment with CK-666 resulted in significantly reduced cortactin localization on Rab7+ endosomes (Fig. 4, A and B). Moreover, the accumulation of cortactin that occurs with inhibition of PI(3,5)P2 synthesis by YM201636 was abolished in CK-666–treated cells. Likewise, Arp2/3 inhibition with CK-666 treatment inhibited actin accumulation on Rab7+ late endosomes in both control and YM201636-treated cells (Fig. 4, A and C). These data suggest a model in which cortactin is recruited to late endosomes by interactions with branched actin networks and removed by interaction with PI(3,5)P2.


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

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

Recruitment of cortactin to late endosomes depends on Arp2/3 complex activity. (A) Representative images show cortactin (green), actin (red), and Rab7 (blue) localization after 2 h treatment of MDA-MB-231 cells with 800 nM YM201636 ± 200 µM CK-666. Bars, 20 µm (5 µm for magnifications). (B) Images from cells treated with YM201636 and 100 or 200 µM CK-666 were analyzed for percentage of colocalization of cortactin or actin with Rab7. The DMSO and YM201636 datasets (no CK-666) are the same data as that shown in Fig. 3 ( C and D). Data shown as box and whiskers plots with the box indicating the 25th and 75th percentiles, solid line indicating the median, and whiskers indicating the 95% confidence intervals. 3 independent experiments, n ≥ 50 cells for each condition. ****, P < 0.0001.
© Copyright Policy - openaccess
Related In: Results  -  Collection

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fig4: Recruitment of cortactin to late endosomes depends on Arp2/3 complex activity. (A) Representative images show cortactin (green), actin (red), and Rab7 (blue) localization after 2 h treatment of MDA-MB-231 cells with 800 nM YM201636 ± 200 µM CK-666. Bars, 20 µm (5 µm for magnifications). (B) Images from cells treated with YM201636 and 100 or 200 µM CK-666 were analyzed for percentage of colocalization of cortactin or actin with Rab7. The DMSO and YM201636 datasets (no CK-666) are the same data as that shown in Fig. 3 ( C and D). Data shown as box and whiskers plots with the box indicating the 25th and 75th percentiles, solid line indicating the median, and whiskers indicating the 95% confidence intervals. 3 independent experiments, n ≥ 50 cells for each condition. ****, P < 0.0001.
Mentions: One mechanism by which cortactin could promote accumulation of actin on endosomes is by regulating branched actin network assembly by the Arp2/3 complex (Uruno et al., 2001; Weaver et al., 2001). To determine whether Arp2/3-mediated branched actin nucleation is required for recruiting cortactin to endosomal membranes, we treated cells with the Arp2/3 complex inhibitor CK-666 (Nolen et al., 2009; Hetrick et al., 2013). Treatment with CK-666 resulted in significantly reduced cortactin localization on Rab7+ endosomes (Fig. 4, A and B). Moreover, the accumulation of cortactin that occurs with inhibition of PI(3,5)P2 synthesis by YM201636 was abolished in CK-666–treated cells. Likewise, Arp2/3 inhibition with CK-666 treatment inhibited actin accumulation on Rab7+ late endosomes in both control and YM201636-treated cells (Fig. 4, A and C). These data suggest a model in which cortactin is recruited to late endosomes by interactions with branched actin networks and removed by interaction with PI(3,5)P2.

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