Oral Presentation ASDR-AWTRS-MEPSA 2018 Joint Meeting

High-content imaging of melanocyte-keratinocytes-fibroblast interaction reveals novel phenotypes for melanoma-associated oncogenic mutations (#12)

Dubravka Skalamera 1 , Alexander J Stevenson 1 , Steven Ainger 2 , Richard Sturm A Sturm 2 , Brian Gabrielli 1
  1. Mater Medical Research Institute, TRI, University of Queensland, Woollongabba, QLD, Australia
  2. UQ Diamantina Institute, University of Queensland, Brisbane, Queensland, Australia

Understanding melanocytic cell interactions forms the basis of treatment development for multiple diseases, including cutaneous pigmentation disorders and melanoma. We have established an automated-microscopy platform for high-resolution analysis of melanocyte-keratinocyte-fibroblast interaction in microwell plates. The arrayed nature of this system allows for combinatorial analysis of genotypes underlying skin homeostasis, and was used to analyse the effects of oncogene expression in melanocytic line variants. In coculture with HACAT keratinocytes, wild type melanocytes formed keratinocyte-overlapping dendrites facilitating molecular exchange. The dendritic phenotype was reduced in melanocytes expressing NRAS.Q61K, one of the most common melanoma-associated mutations, which also increased melanocyte migration resulting in keratinocyte avoidance. This mutant also altered interaction with primary dermal fibroblasts, to which melanocytes would not be exposed in intact skin. Under our culture conditions, dermal fibroblasts reduced adult melanocyte proliferation and even triggered apoptosis in a proportion of cells. This effect was abolished by NRAS.Q61K expression, suggesting that in addition to known role in activating mitogen signalling, NRAS.Q61K may also contribute to cancer progression conferring avoidance of restrictive cell-cell interactions. Since melanocyte-keratinocyte-fibroblast interaction forms the bases of skin homeostasis, our findings have implications for management of melanoma as well pigmentation disorders and wound healing.