The malarial CDK Pfmrk and its effector PfMAT1 phosphorylate DNA replication proteins and co-localize in the nucleus

Authors

Dayadevi Jirage, Division of Experimental Therapeutics, Walter Reed Army Institute of Research, Silver Spring, MD
Yueqin Chen, Division of Experimental Therapeutics, Walter Reed Army Institute of Research, Silver Spring, MD
Diana Caridha, Division of Experimental Therapeutics, Walter Reed Army Institute of Research, Silver Spring, MD
Michael T. O’Neil, Division of Experimental Therapeutics, Walter Reed Army Institute of Research, Silver Spring, MD
Fredrick Eyase, United States Army Medical Research Unit-Kenya, Walter Reed Project, Kisumu, MRU
William H. Witola, University of Chicago Medical Center
Choukri Ben Mamoun, Yale School of Medicine, Section of Infectious Diseases
Norman C. Waters, Division of Experimental Therapeutics, Walter Reed Army Institute of Research, Silver Spring, MDFollow

Date of this Version

2010

Citation

Molecular & Biochemical Parasitology 172 (2010) 9–18

Comments

U.S. Government Work

Abstract

Cyclin-dependent kinases (CDKs) have an established role in metazoans and yeast in DNA replication, transcription and cell cycle regulation. Several CDKs and their effectors have been identified in the malaria parasite Plasmodium falciparum and their biological functions are beginning to be investigated. Here we report results from the functional characterization of Pfmrk and its effector PfMAT1. We validated the interactions between Pfmrk and PfMAT1 and pinpointed their intracellular location. Co-immunoprecipitation studies demonstrated physical interaction between the two proteins and identified the C-terminal domain of PfMAT1 as the Pfmrk activator domain. Immunofluorescence analyses using GFP and RFP-tagged versions of Pfmrk and PfMAT1, respectively, demonstrated the co-localization of these two proteins to the parasite nucleus. Bacterial two-hybrid screen of a P. falciparum cDNA library using Pfmrk as the bait identified two plasmodial DNA replication proteins, PfRFC-5 and PfMCM6, as interactors with Pfmrk. We demonstrate that that these two proteins are substrates of Pfmrk-mediated phosphorylation and that PfMAT1 confers substrate specificity to the Pfmrk kinase complex. Collectively, these data suggest a role for Pfmrk in the nucleus of the parasite presumably in regulation of the DNA replication machinery.