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Collagen triple helix repeat containing 1 increases melanoma cell migration, adhesion and survival through modulation of the actin cytoskeleton

University of British Columbia

Background: Collagen Triple Helix Repeat Containing 1 (CTHRC1) is a recently discovered extracellular protein that can bind and activate Wnt signaling pathway. In previous gene expression profiling experiments, it was found to be aberrantly upregulated in metastatic melanoma and its expression level was correlated with melanoma progression and metastasis. Objective: The purpose of this study is to understand the functional impact of CTHRC1 on cancer using melanoma cell lines as a model. Experimental Methods: We transfected two melanoma cell lines, MMAN and MMRU, with plasmid vectors to create stable clones with high and low CTHRC1 expression to study the functional effects of CTHRC1 in vitro. Using these two cell lines, we assayed for melanoma migration, adhesion and survival using scratch wound healing assay, attachment assay and cell cycle analysis, respectively. In addition, the cells were stained for F-actin with AlexaFluor 594 labeled phalloidin to observe for actin organization. Results: Using these two pairs of cell lines, we have found that CTHRC1 expression increased melanoma cell migration, enhanced melanoma cell adhesion to both tissue culture plastic and matrigel, and protected melanoma cells from serum deprivation induced apoptosis. Further, it was demonstrated that CTHRC1’s pro-survival effect was dependent on cell adhesion, as the protection effect was lost when melanoma cells were cultured in suspension. Immunofluorescent staining of F-actin revealed that CTHRC1 expression increased the formation structures such as focal complexes, lamellipodia and filopodia. Discussion: The increased formation of the adhesion structures may be the key to CTHRC1 associated cell migration, adhesion and survival. These structures are likely regulated by the Rho family of proteins that act downstream of the Wnt/PCP pathway, with which CTHRC1 has been previously demonstrated to be involved as a co-receptor. Conclusion: Results from this study suggest that CTHRC1 expression promotes cellular behaviours associated with tumour metastases. Therefore, inhibition of this protein may be able to block melanoma metastasis and may have value as a potential therapeutic.

Thesis/Dissertation

application/pdf

2009-06-11

Attribution-NonCommercial-NoDerivatives 4.0 International

http://hdl.handle.net/2429/8929

Experimental Medicine

University of British Columbia

UBCV

http://creativecommons.org/licenses/by-nc-nd/4.0/