CLONING OF A CDC2-RELATED PROTEIN-KINASE FROM TRYPANOSOMA-CRUZI THAT INTERACTS WITH MAMMALIAN CYCLINS

Two cdc2-related protein kinases (crk), tzcrk3 and tzcrk1, from the pr otozoan parasite Trypanosoma cruzi were cloned. tzcrk3 encodes a 35 kD a protein sharing 51.5% amino acid identity with human cdc2 and 82% id entity with Trypanosoma brucei CRK3. tzcrk1 encodes a 33 kDa protein s haring 52.7% identity with human cdc2 and a high degree of identity (> 78%) with T. brucei CRK1, Leishmania mexicana CRK1 and Trypanosoma co ngolense CRK1. A recombinant TzCRK1 protein was able to phosphorylate histone HI and retinoblastoma protein. Western blotting using a polycl onal antibody raised against the recombinant TzCRK1 protein showed tha t the kinase is present in all life cycle stages of the parasite. A PS TAIRE antiserum detected proteins of 32, 33 and 35 kDa, with different ial expression in the life cycle of the parasite. Transfection of COS- 7 cells with tzcrk1 demonstrated for the first time that a CRK protein can bind mammalian cyclins; TzCRK1 co-immunoprecipitated with cyclins E, D3 and A suggesting a role for this kinase in cell cycle control. These results indicate that T. cruzi might have cyclin homologues that control the activity of the CRK proteins and that a complex mechanism would exist in order to regulate the kinases involved in the cell cyc le and the differentiation processes of the parasite. (C) 1998 Elsevie r Science B.V. All rights reserved.