ZFIN Image: Liao et al., 2019, Figure 5

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Figure Caption

Figure 5

ALS-Associated Mutations in ANXA11 Disrupt Its Interactions with Lysosomes

(A) Live cell imaging of Opto-mCherry, wild-type Opto-ANXA11, or mutant Opto-ANXA11 with LAMP1-HaloTag in U2OS cells before and after oligomerization induced by exposure to 488nm light. Scale bar: 2 μm. See also Figure S5A.

(B) Quantification of percentage of light-activated Opto-mcherry (CRY2olig-mCherry), wild-type Opto-ANXA11, or mutant Opto-ANXA11 clusters co-localizing with lysosomes at 300s post-488 nm light exposure from (A). n=26-30. One-way ANOVA, ns, not significant. ^∗∗∗∗p < 0.0001. Error bars = SEM. See also Figure S5B.

(C) Extent of co-localization of wild-type or mutant ANXA11 with lysosomes. U2OS cells expressing LAMP1-HaloTag, wild-type ANXA11-mEmerald or mutant ANXA11-mEmerald were imaged 30 minutes after heat shock (43^oC). Scale bar: 2 μm.

(D) Percentage of fluorescence associated with wild-type ANXA11 or mutant ANXA11 that co-localized with lysosomes from (C). n= 22-40. One-way ANOVA, ns, not significant. ^∗∗∗∗p < 0.0001. ^∗p < 0.05. Error bars = SEM.

(E) Extent of co-trafficking of wild-type or mutant ANXA11 with lysosomes in axons. Axons of rat cortical neurons expressing LAMP1-HaloTag and wild-type or mutant ANXA11-mEmerald were imaged for 30 seconds. Kymographs show WT and p.D40G ANXA11 both co-traffic with lysosomes (see arrows) while p.R235Q and p.R346C each disrupt ANXA11 co-trafficking with lysosomes. Scale bar: 10 μm.

(F) Number of puncta containing WT or mutant ANXA11 that co-trafficked with lysosomes as a function of total lysosome number from (E). n= 21-30. One-way ANOVA, ns, not significant. ^∗∗p < 0.01. ^∗p < 0.05. Error bars = SEM.

Acknowledgments

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Reprinted from Cell, 179, Liao, Y.C., Fernandopulle, M.S., Wang, G., Choi, H., Hao, L., Drerup, C.M., Patel, R., Qamar, S., Nixon-Abell, J., Shen, Y., Meadows, W., Vendruscolo, M., Knowles, T.P.J., Nelson, M., Czekalska, M.A., Musteikyte, G., Gachechiladze, M.A., Stephens, C.A., Pasolli, H.A., Forrest, L.R., St George-Hyslop, P., Lippincott-Schwartz, J., Ward, M.E., RNA Granules Hitchhike on Lysosomes for Long-Distance Transport, Using Annexin A11 as a Molecular Tether, 147-164.e20, Copyright (2019) with permission from Elsevier. Full text @ Cell