MOLECULAR-CLONING AND CELL-CYCLE-DEPENDENT EXPRESSION OF A NOVEL NIMA(NEVER-IN-MITOSIS IN ASPERGILLUS NIDULANS)-RELATED PROTEIN-KINASE (TPNRK) IN TETRAHYMENA CELLS

With the intention of investigating the signal-transduction pathway th at mediates the cold-stress response in Tetrahymena, we isolated a gen e that encodes a novel protein kinase of 561 amino acids, termed Tetra hymena pyriformis NIMA (never-in-mitosis in Aspergillus nidulans)-rela ted protein kinase (TpNrk), by differential display from Tetrahymena c ells exposed to temperature shift-down. TpNrk possesses an N-terminal protein kinase domain that is highly homologous with other NIMA-relate d protein kinases (Neks) involved in the control of the cell cycle. Th e TpNrk protein is 42 %, identical in its catalytic domain with human Nek2, 41 % identical with mouse Nek1 and 37 % with A. nidulans NIMA. I n addition, TpNrk and these NIMA-related kinases have long, basic C-te rminal extensions and are therefore similar in overall structure. In o rder to further explore the function of the TpNrk gene and the associa tion of the cold stress with the cell cycle of Tetrahymena, changes of TpNrk mRNA were determined during the course of the synchronous cell division induced by the intermittent heat treatment. The level of TpNr k transcription increased immediately after the end of the heat treatm ent, with a peak at 30 min, and declined thereafter reaching the minim um level when nearly 80 %, of the cells synchronously entered cell div ision (75 min after the end of heat treatment). The accumulation of Tp Nrk mRNA starting from 0 min to 30 min after the end of the heat treat ment was assumed to be a prerequisite for the start of synchronous cel l division. These results suggest that TpNrk may have a role in the ce ll cycle of Tetrahymena, and that mRNA expression, at least, is under tight cell-cycle control.