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 approximately 50%, 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.