An inverted CAV1 (caveolin 1) topology defines novel autophagy-dependent exosome secretion from prostate cancer cells

Ariotti, Nicholas; Wu, Yeping; Okano, Satomi; Gambin, Yann; Follett, Jordan; Rae, James; Ferguson, Charles; Teasdale, Rohan D.; Alexandrov, Kirill; Meunier, Frédéric A.; Hill, Michelle M.; Parton, Robert G.

doi:10.1080/15548627.2020.1820787

Cited by 23 publications

(18 citation statements)

References 81 publications

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“…Indeed, recent studies implicate intracellular CAV1 in regulating various organelle compartments, with particular emphasis on roles in autophagosomes and cancer. 39,50,51 The premise that membrane remodelling drives exosomal miRNA sorting is consistent with our prior observa-tion that CAVIN1 attenuates non-caveolar CAV1 in prostate cancer but CAVIN1 itself was not detected in PC3 EVs. 2 In addition to CAV1, other metastasis-promoting genes like Ras family proteins and tetraspanins also impact membrane composition, [52][53][54] hence, membrane remodelling appears to be a common mechanism underlying release of pro-metastatic EVs in cancers.…”

Section: Discussionsupporting

confidence: 89%

“…This is shown in PC3 cells treated with hnRNPK-targeting (si-hnRNPK) or non-targeting siRNA (si-NegCont) lung epithelial cells. 38 While we observed that CAV1 is indeed released in PC3 EV via the endosomal-pathway, 39 its release is not altered in PC3-CAVIN1 cells. 4 To test the possibility that CAV1 binds to hnRNPK in the PC3 cell model, we conducted a coimmunoprecipitation experiment to find hnRNPK binding partners in both PC3-CONT and PC3-CAVIN1 whole cell lysates.…”

Section: 6mentioning

confidence: 62%

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Caveolin‐1‐driven membrane remodelling regulates hnRNPK‐mediated exosomal microRNA sorting in cancer

Robinson

Ruelcke

Blythe

et al. 2021

Clinical & Translational Med

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Background Caveolae proteins play diverse roles in cancer development and progression. In prostate cancer, non‐caveolar caveolin‐1 (CAV1) promotes metastasis, while CAVIN1 attenuates CAV1‐induced metastasis. Here, we unveil a novel mechanism linking CAV1 to selective loading of exosomes with metastasis‐promoting microRNAs. Results We identify hnRNPK as a CAV1‐regulated microRNA binding protein. In the absence of CAVIN1, non‐caveolar CAV1 drives localisation of hnRPNK to multi‐vesicular bodies (MVBs), recruiting AsUGnA motif‐containing miRNAs and causing their release within exosomes. This process is dependent on the lipid environment of membranes as shown by cholesterol depletion using methyl‐β‐cyclodextrin or by treatment with n‐3 polyunsaturated fatty acids. Consistent with a role in bone metastasis, knockdown of hnRNPK in prostate cancer PC3 cells abolished the ability of PC3 extracellular vesicles (EV) to induce osteoclastogenesis, and biofluid EV hnRNPK is elevated in metastatic prostate and colorectal cancer. Conclusions Taken together, these results support a novel pan‐cancer mechanism for CAV1‐driven exosomal release of hnRNPK and associated miRNA in metastasis, which is modulated by the membrane lipid environment.

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Section: Discussionsupporting

confidence: 89%

Section: 6mentioning

confidence: 62%

Caveolin‐1‐driven membrane remodelling regulates hnRNPK‐mediated exosomal microRNA sorting in cancer

Robinson

Ruelcke

Blythe

et al. 2021

Clinical & Translational Med

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“…To further determine the level of GFP association with EVs and the localization of GFP in EVs, FCS analysis of EVs with or without proteinase K treatment was performed. FCS is a highly sensitive method to obtain hydrodynamic diameters, enzyme kinetics, and number of cargo loading/release or surface functionalization on nanoparticles (Ariotti et al., 2021 ; Massi et al., 2020 ; Rigler et al., 1993 ; Rigler & Meier, 2006 ). Incubation of EVs with proteinase K is a common method to study localization of proteins in/on EVs (Ariotti et al., 2021 ; Bonsergent et al., 2021 ; Charoenviriyakul et al., 2018 ).…”

Section: Resultsmentioning

confidence: 99%

“…FCS is a highly sensitive method to obtain hydrodynamic diameters, enzyme kinetics, and number of cargo loading/release or surface functionalization on nanoparticles (Ariotti et al., 2021 ; Massi et al., 2020 ; Rigler et al., 1993 ; Rigler & Meier, 2006 ). Incubation of EVs with proteinase K is a common method to study localization of proteins in/on EVs (Ariotti et al., 2021 ; Bonsergent et al., 2021 ; Charoenviriyakul et al., 2018 ). The level of GFP association with EVs was first quantified with EVs in PBS.…”

Section: Resultsmentioning

confidence: 99%

Presentation of antigen on extracellular vesicles using transmembrane domains from viral glycoproteins for enhanced immunogenicity

McKay

Samnuan

et al. 2022

J of Extracellular Vesicle

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A vaccine antigen, when launched as DNA or RNA, can be presented in various forms, including intracellular, secreted, membrane‐bound, or on extracellular vesicles (EVs). Whether an antigen in one or more of these forms is superior in immune induction remains unclear. In this study, we used GFP as a model antigen and first compared the EV‐loading efficiency of transmembrane domains (TMs) from various viral glycoproteins, and then investigated whether EV‐bound GFP (EV‐GFP) would enhance immune induction. Our data showed that GFP fused to viral TMs was successfully loaded onto the surface of EVs. In addition, GFP‐bound EVs were predominantly associated with the exosome marker CD81. Immunogenicity study with EV‐GFP‐producing plasmids in mice demonstrated that antigen‐specific IgG and IgA were significantly increased in EV‐GFP groups, compared to soluble and intracellular GFP groups. Similarly, GFP‐specific T cell response‐related cytokines produced by antigen‐stimulated splenocytes were also enhanced in mice immunized with EV‐GFP constructs. Immunogenicity study with purified soluble GFP and GFP EVs further confirmed the immune enhancement property of EV‐GFP in mice. In vitro uptake assays indicated that EV‐GFP was more efficiently taken up than soluble GFP by mouse splenocytes and such uptake was B cell preferential. Taken together, our data indicate that viral TMs can efficiently load antigens onto the EV surface, and that EV‐bound antigen enhances both humoral and cell‐mediated antigen‐specific responses.

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“…The preparation of cells for transmission electron microscopy was performed as described previously [ 45 ]. Briefly, NCI-H460 and A549 cells were plated onto tissue culture-treated plastic.…”

Section: Methodsmentioning

confidence: 99%

βIII-Tubulin Structural Domains Regulate Mitochondrial Network Architecture in an Isotype-Specific Manner

Parker

Teo

Brayford

et al. 2022

Cells

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βIII-tubulin is a neuronal microtubule protein that is aberrantly expressed in epithelial cancers. The microtubule network is implicated in regulating the architecture and dynamics of the mitochondrial network, although the isotype-specific role for β-tubulin proteins that constitute this microtubule network remains unclear. High-resolution electron microscopy revealed that manipulation of βIII-tubulin expression levels impacts the volume and shape of mitochondria. Analysis of the structural domains of the protein identifies that the C-terminal tail of βIII-tubulin, which distinguishes this protein from other β-tubulin isotypes, significantly contributes to the isotype-specific effects of βIII-tubulin on mitochondrial architecture. Mass spectrometry analysis of protein–protein interactions with β-tubulin isotypes identifies that βIII-tubulin specifically interacts with regulators of mitochondrial dynamics that may mediate these functional effects. Advanced quantitative dynamic lattice light sheet imaging of the mitochondrial network reveals that βIII-tubulin promotes a more dynamic and extended reticular mitochondrial network, and regulates mitochondrial volume. A regulatory role for the βIII-tubulin C-terminal tail in mitochondrial network dynamics and architecture has widespread implications for the maintenance of mitochondrial homeostasis in health and disease.

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An inverted CAV1 (caveolin 1) topology defines novel autophagy-dependent exosome secretion from prostate cancer cells

Cited by 23 publications

References 81 publications

Caveolin‐1‐driven membrane remodelling regulates hnRNPK‐mediated exosomal microRNA sorting in cancer

Caveolin‐1‐driven membrane remodelling regulates hnRNPK‐mediated exosomal microRNA sorting in cancer

Presentation of antigen on extracellular vesicles using transmembrane domains from viral glycoproteins for enhanced immunogenicity

βIII-Tubulin Structural Domains Regulate Mitochondrial Network Architecture in an Isotype-Specific Manner

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