Oral Presentation ASDR-AWTRS-MEPSA 2018 Joint Meeting

MITF regulates dynamic heterogeneity and structural integrity in melanoma (#11)

Loredana LS Spoerri 1 , Kimberley KAB Beaumont 2 , Crystal CAT Tonnessen 1 , David DSH Hill 2 , Russell RJJ Jurek 3 , Gency GG Gunasingh 1 , Sheena SMD Daignault 1 , Farzana FA Ahmed 1 , Aaron AGS Smith 4 , Wolfgang WW Weninger 2 5 , Nikolas NKH Haass 1 2 5
  1. Diamantina Institute (TRI), University of Queensland, Brisbane
  2. The Centenary Institute, Newtown, NSW, Australia
  3. CSIRO Astronomy & Space Sciences, Australia Telescope National Facility, Epping, NSW, Australia
  4. Dermatology Research Centre, Translational Research Institute, School of Medicine, The University of Queensland, Brisbane, Qld, Australia
  5. Discipline of Dermatology, University of Sydney, Sydney, NSW, Australia

Differential tumour cell behaviour caused by environmental conditions, termed dynamic heterogeneity, is a prime source for drug resistance. We utilise real-time cell cycle imaging (FUCCI) to study melanoma heterogeneity. As distinct proliferative and invasive capabilities reflect variable drug sensitivities, identifying these different responses is crucial to design effective therapies. Mouse xenograft tumours generated from cell lines with high microphthalmia-associated transcription factor (MITF) level displayed a homogeneous distribution of cycling cells throughout. In contrast, tumours generated from cell lines with low MITF levels were composed of clusters of cycling cells and clusters of G1-arrested cells. The proliferating areas were in close proximity to blood vessels, presumably characterized by oxygen/nutrient availability. Melanoma spheroids recapitulated the in vivo cycling behaviour, considering that here oxygen and nutrients are supplied by diffusion. MITF was undetectable within the hypoxic G1-arrested spheroid core, indicating hypoxia-induced MITF downregulation. Furthermore, modulation of MITF expression impacted spheroid morphology, with overexpression giving rise to flatter structures whereas knock down to smaller aggregates with unaffected sphericity. The loss of morphological integrity caused by increased MITF expression did not reduce spheroids’ inner hypoxic level, dismissing the hypothesis that these compromised structures could be more permeable to oxygen resulting in decreased hypoxia induced G1-arrest. Proteomic analysis revealed that modulation of MITF level induced differences in cell-cell and cell-ECM adhesion. In addition, inhibition of the Rho/ROCK signalling pathway, known to control cell contractility, partially mimicked the morphology and cell cycle effects of MITF overexpression. We conclude that MITF protects from cell cycle arrest induced by oxygen deprivation. We hypothesise that high MITF levels prevent cell cycle arrest by reducing the cell-intrinsic propensity to arrest in response to low oxygen via a mechanism involving cell-cell/ECM crosstalk.