Variation in cadherins and catenins expression is linked to both proliferation and transformation of Rhabdomyosarcoma

Abstract

Cadherins are a family of transmembrane glycoproteins that mediate Ca 2 þ -dependent homophilic cell-cell adhesion and play a crucial role in cell differentiation. E-cadherin-mediated cell-cell adhesion is lost during the development of most epithelial cancers. This study examines cadherin-dependent adhesion in cell lines derived from rhabdomyosarcoma (RMS), a highly malignant softtissue tumor committed to the myogenic lineage, but arrested prior to terminal differentiation. We analysed the expression of cadherins and associated catenins at the mRNA and protein levels as well as their localization in nine RMS-derived cell lines relative to normal myoblasts. We show a decrease in the expression of cadherins and catenins in all RMS-derived cell lines compared to control cells. This decrease in the expression of N-and M-cadherin was confirmed in RMS biopsies. In contrast, R-cadherin is found expressed in RMS, whereas it is normally absent in normal myoblasts. We show that a decrease of R-cadherin expression using RNA interference inhibits cell proliferation of the RD cell line. In addition to their diminished expression, cadherins and catenins do not localize to intercellular contacts in embryonal RMS (ERMS), whereas specific persistent localization is seen in alveolar RMS (ARMS)-derived cell lines. Thus, RMS exhibit defects in the expression of molecules of the cadherin family. Defects in the localization of these adhesion molecules at the sites of cell-cell contact are specifically observed in the ERMS subtype. In addition, our data suggest that R-cadherin is a specific diagnostic marker for RMS and is also an important factor of RMS cell proliferation.

RhoA GTPase Regulates M-Cadherin Activity and Myoblast Fusion

“…Cells cotransfected with empty GFP and control R-cadherin RNAi were able to fuse, whereas cells cotransfected with empty GPF and M-cadherin RNAi did not fuse ( Figure 3B). Our R-cadherin RNAi was previously validated in a rhabdomyosarcoma-derived cell line (Charrasse et al, 2004), and it does not perturb the expression of the myogenic markers described above (our unpublished data). Quantification of the number of transfected cells that have fused in these two conditions is shown in Figure 3C.…”

“…Short interfering RNA (siRNA) constructs were made in pSUPER polymerase III expression vector (Charrasse et al, 2004). To suppress endogenous Mcadherin expression, oligonucleotide GATCCCCCAGCAGCTAGGCAGTGTCAttcaagagaTGACACTGCCTA GCTGCTGTTTTTGGAAA was inserted into pSUPER.…”

“…Bold letters correspond to oligonucleotides 735-753 of the mouse M-cadherin cDNA sequence (NM_007662). As a control, R-cadherin siRNA was used (Charrasse et al, 2004). Cells (25,000) were plated in six-well Limbro 1 d before transfection.…”

“…Anti-Mcadherin was revealed by either an Alexa Fluor 546-or an Alexa Fluor 488-conjugated goat anti-rabbit antibody (Molecular Probes, Interchim, Montluçon, France). Cells were analyzed as previously described (Charrasse et al, , 2003(Charrasse et al, , 2004.…”

RAGE signaling deficiency in rhabdomyosarcoma cells causes upregulation of PAX7 and uncontrolled proliferation

“…Aside from low RAGE levels and/or signaling in ERMSs, other factors and/or conditions might cause ERMSs to express high PAX7 levels. For example, altered expression levels of N-cadherin (Charrasse et al, 2004) or Cdo (Wegorzewska et al, 2003) in ERMSs might contribute to high PAX7 levels because signaling of N-cadherin (Lovett et al, 2006) and Cdo (Takaesu et al, 2006) impacts upon p38 MAPK.…”