Open AccessCaveolin‐1 down‐regulation is required for Wnt5a‐Frizzled 2 signalling in Ha‐Ras V12 ‐induced cell transformation
Author(s) -
Lin HsiuKuan,
Lin HsiHui,
Chiou YuWei,
Wu ChingLung,
Chiu WenTai,
Tang MingJer
Publication year - 2018
Publication title -
journal of cellular and molecular medicine
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.44
H-Index - 130
eISSN - 1582-4934
pISSN - 1582-1838
DOI - 10.1111/jcmm.13531
Subject(s) - microbiology and biotechnology , exosome , caveolin 1 , microvesicles , cell culture , biology , rna interference , transformation (genetics) , chemistry , microrna , rna , gene , biochemistry , genetics
Caveolin‐1 (Cav1) is down‐regulated during MK 4 ( MDCK cells harbouring inducible Ha‐Ras V12 gene) transformation by Ha‐Ras V12 . Cav1 overexpression abrogates the Ha‐Ras V12 ‐driven transformation of MK 4 cells; however, the targeted down‐regulation of Cav1 is not sufficient to mimic this transformation. Cav1‐silenced cells, including MK 4/shCav1 cells and MDCK /shCav1 cells, showed an increased cell area and discontinuous junction‐related proteins staining. Cellular and mechanical transformations were completed when MDCK /shCav1 cells were treated with medium conditioned by MK 4 cells treated with IPTG ( MK 4+I‐ CM ) but not with medium conditioned by MK 4 cells. Nanoparticle tracking analysis showed that Ha‐Ras V12 ‐inducing MK 4 cells increased exosome‐like microvesicles release compared with their normal counterparts. The cellular and mechanical transformation activities of MK 4+I‐ CM were abolished after heat treatment and exosome depletion and were copied by exosomes derived from MK 4+I‐ CM ( MK 4+I‐ EX s). Wnt5a, a downstream product of Ha‐Ras V12 , was markedly secreted by MK 4+I‐ CM and MK 4+I‐ EX s. Suppression of Wnt5a expression and secretion using the porcupine inhibitor C59 or Wnt5a si RNA inhibited the Ha‐Ras V12 ‐ and MK 4+I‐ CM ‐induced transformation of MK 4 cells and MDCK /shCav1 cells, respectively. Cav1 down‐regulation, either by Ha‐Ras V12 or targeted sh RNA , increased frizzled‐2 (Fzd2) protein levels without affecting its mRNA levels, suggesting a novel role of Cav1 in negatively regulating Fzd2 expression. Additionally, silencing Cav1 facilitated the internalization of MK 4+I‐ EX s in MDCK cells. These data suggest that Cav1‐dependent repression of Fzd2 and exosome uptake is potentially relevant to its antitransformation activity, which hinders the activation of Ha‐Ras V12 ‐Wnt5a‐Stat3 pathway. Altogether, these results suggest that both decreasing Cav1 and increasing exosomal Wnt5a must be implemented during Ha‐Ras V12 ‐driven cell transformation.