Mirk/Dyrk1B Maintains the Viability of Quiescent Pancreatic Cancer Cells by Reducing Levels of Reactive Oxygen Species

Abstract

The kinase Mirk/dyrk1B mediated the clonogenic growth of pancreatic cancer cells in earlier studies. It is now shown that Mirk levels increased 7-fold in SU86.86 pancreatic cancer cells when over a third of the cells were accumulated in a quiescent G 0 state, defined by Hoechst/Pyronin Y staining. Depletion of Mirk by a doxycycline-inducible short hairpin RNA increased the G 0 fraction to f50%, suggesting that Mirk provided some function in G 0 . Mirk reduced the levels of reactive oxygen species (ROS) in quiescent cultures of SU86.86 cells and of Panc1 cells by increasing transcription of the antioxidant genes ferroxidase, superoxide dismutase (SOD)2, and SOD3. These genes were functional antioxidant genes in pancreatic cancer cells because ectopic expression of SOD2 and ferroxidase in Mirk-depleted cells lowered ROS levels. Quiescent pancreatic cancer cells quickly lost viability when depleted of Mirk because of elevated ROS levels, exhibiting up to 4-fold less colony-forming activity and 4-fold less capability for dye exclusion. As a result, reduction of ROS by N-acetyl cysteine led to more viable cells. Mirk also destabilizated cyclin D1 and D3 in quiescent cells. Thus, quiescent pancreatic cancer cells depleted of Mirk became less viable because they were damaged by ROS, and had increased levels of G 1 cyclins to prime cells to escape quiescence. [Cancer Res 2009;69(8):3317-24]

Molecular mechanisms of cisplatin resistance

“…Dual-specificity Y-phosphorylation regulated kinase 1B (DYRK1B, also known as MIRK) is upregulated in multiple solid tumors (Friedman, 2007) and exerts prosurvival functions by increasing the expression of antioxidant enzymes such as ferroxidase, superoxide dismutase 2 and superoxide dismutase 3 (Deng et al, 2009). In NSCLC and ovarian cancer cells, DYRK1B depletion has been shown to potentiate the effects of subapoptotic cisplatin concentrations by favoring the establishment of lethal oxidative stress (Gao et al, 2009;Hu and Friedman, 2010).…”

TRPM7 regulates quiescent/proliferative metabolic transitions in lymphocytes

“…Cellular RNA levels can be defined in relation to their cell cycle position by sequential staining of cells with DAPI (binds to DNA), followed by Pyronin Y (PY) staining (binds to RNA only, after DNA binding sites are saturated with DAPI). 17 In order to assess RNA levels and cell cycle status of TRPM7-KO lymphocytes transitioned to regular media for 26 hrs, we compared them with sequentially labeled WT cells An additional fundamental property of cellular quiescence is the nature of cellular energy metabolism. In lymphocytes, energy metabolism in quiescence is dominated by respiration, and thus quiescent lymphocytes exhibit low rates of glucose utilization and oxygen consumption (reviewed in ref.…”

Transient arrest in a quiescent state allows ovarian cancer cells to survive suboptimal growth conditions and is mediated by both Mirk/dyrk1b and p130/Rb2

“…RNAi duplexes and GC-matched control RNAi duplexes were transfected into cells with lipofectamine 2000 in serum-containing growth medium as described. 7 …”

“…7,38,39 Mirk was shown to also alter ovarian cancer cell cycling through cyclin D1. Efficient depletion of Mirk in SKOV3, OVCAR3 and TOV21G cells by either of two synthetic RNAi duplexes led to a 2-fold increase in cyclin D1 levels after 2-3 days of culture (data not shown).…”