Abstract
BackgroundProtein kinase CK2 is a pleiotropic serine/threonine protein kinase with hundreds of reported substrates, and plays an important role in a number of cellular processes. The cellular functions of Plasmodium falciparum CK2 (PfCK2) are unknown. The parasite's genome encodes one catalytic subunit, PfCK2α, which we have previously shown to be essential for completion of the asexual erythrocytic cycle, and two putative regulatory subunits, PfCK2β1 and PfCK2β2.ResultsWe now show that the genes encoding both regulatory PfCK2 subunits (PfCK2β1 and PfCK2β2) cannot be disrupted. Using immunofluorescence and electron microscopy, we examined the intra-erythrocytic stages of transgenic parasite lines expressing hemagglutinin (HA)-tagged catalytic and regulatory subunits (HA-CK2α, HA-PfCK2β1 or HA-PfCK2β2), and localized all three subunits to both cytoplasmic and nuclear compartments of the parasite. The same transgenic parasite lines were used to purify PfCK2β1- and PfCK2β2-containing complexes, which were analyzed by mass spectrometry. The recovered proteins were unevenly distributed between various pathways, with a large proportion of components of the chromatin assembly pathway being present in both PfCK2β1 and PfCK2β2 precipitates, implicating PfCK2 in chromatin dynamics. We also found that chromatin-related substrates such as nucleosome assembly proteins (Naps), histones, and two members of the Alba family are phosphorylated by PfCK2α in vitro.ConclusionsOur reverse-genetics data show that each of the two regulatory PfCK2 subunits is required for completion of the asexual erythrocytic cycle. Our interactome study points to an implication of PfCK2 in many cellular pathways, with chromatin dynamics being identified as a major process regulated by PfCK2. This study paves the way for a kinome-wide interactomics-based approach to elucidate protein kinase function in malaria parasites.
Highlights
Protein kinase CK2 is a pleiotropic serine/threonine protein kinase with hundreds of reported substrates, and plays an important role in a number of cellular processes
Protein kinase CK2 is a pleiotropic serine/threonine Protein kinases (PKs), with over 300 substrates identified in mammalian cells [10] and documented roles in multiple cellular processes, including differentiation, proliferation, survival, translation, apoptosis, transformation, tumorigenesis, RNA synthesis, cell cycle progression, cell morphology and polarity, cellular responses to stress and to DNA damage, and circadian rhythm [11,12]
The enzyme phosphorylates a range of DNA-binding proteins, nuclear oncoproteins, and transcription factors [13], and several studies have highlighted the predominance of CK2 substrates involved in transcription, chromatin structure, and gene expression [10,12,14]
Summary
Protein kinase CK2 is a pleiotropic serine/threonine protein kinase with hundreds of reported substrates, and plays an important role in a number of cellular processes. Protein kinase CK2 (thereafter called CK2) is a pleiotropic serine/threonine PK, with over 300 substrates identified in mammalian cells [10] and documented roles in multiple cellular processes, including differentiation, proliferation, survival, translation, apoptosis, transformation, tumorigenesis, RNA synthesis, cell cycle progression, cell morphology and polarity, cellular responses to stress and to DNA damage, and circadian rhythm [11,12]. The enzyme phosphorylates a range of DNA-binding proteins, nuclear oncoproteins, and transcription factors [13], and several studies have highlighted the predominance of CK2 substrates involved in transcription, chromatin structure, and gene expression [10,12,14] Taken together, these studies highlight transcriptional regulation as one of the major roles of the enzyme [15]. The requirement for viability of the b subunits of CK2 varies across species: disruption of the single metazoan b subunit is lethal in mice [20] and Caenorhabditis elegans [21], whereas disruption of either or both of the b subunits in Saccharomyces cerevisiae is not lethal [14,22,23]
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