Plasmodium falciparum Prp16 homologue and its role in splicing |
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| Authors: | Prashant Kumar Singh Shivani Kanodia Chethan Jambanna Dandin Usha Vijayraghavan Pawan Malhotra |
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| Institution: | 1. Malaria Research Group, International Centre for Genetic Engineering and Biotechnology, New Delhi 110067, India;2. Department of Microbiology and Cell Biology, Indian Institute of Science, Bangalore 560012, India;1. Department of Biochemistry and Molecular Biology, Johns Hopkins School of Public Health, Baltimore, MD 21205, USA;2. The Johns Hopkins Malaria Research Institute, Baltimore, MD 21205, USA;3. Department of Molecular and Comparative Pathobiology, Johns Hopkins School of Medicine, Baltimore, MD 21205, USA;4. Department of Molecular Microbiology and Immunology, Johns Hopkins School of Public Health, Baltimore, MD 21205, USA;5. Department of Medicine, Johns Hopkins School of Medicine, Baltimore, MD, 21205, USA;1. Genetics Unit, Shriners Hospital for Children, Canada H3G 1A6;2. Department of Surgery, McGill University, Montreal (Quebec), Canada H3A 1A1;1. Institute of Parasitology, McGill University, Canada;2. QIMR Berghofer Medical Research Institute, Australia;3. Department of Biochemistry and Molecular Biology, Bio21 Institute, University of Melbourne, Australia;4. ARC Centre of Excellence for Coherent X-Ray Science, Melbourne, Australia;5. School of Science and Health, University of Western Sydney, Australia;6. Australian Institute for Bioengineering & Nanotechnology, University of Queensland, Australia;7. The Australian National Fabrication Facility, Queensland Node, Brisbane, Australia;8. School of Veterinary Sciences, University of Queensland, Australia;9. School of Natural Sciences, Griffith University, Brisbane, Queensland 4111, Australia;10. Eskitis Institute for Drug Discovery, Griffith University, Queensland, Australia;11. School of Medicine, University of Queensland, Australia;12. School of Biological Sciences, Queen’s University Belfast, Northern Ireland, United Kingdom;1. ICREA at Barcelona Centre for International Health Research (CRESIB, Hospital Clinic - Universitat de Barcelona), 08036 Barcelona, Spain;2. International Centre for Genetic Engineering and Biotechnology (ICGEB), New Delhi 110067, India |
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| Abstract: | Large numbers of Plasmodium genes have been predicted to have introns. However, little information exists on the splicing mechanisms in this organism. Here, we describe the DExD/DExH-box containing Pre-mRNA processing proteins (Prps), PfPrp2p, PfPrp5p, PfPrp16p, PfPrp22p, PfPrp28p, PfPrp43p and PfBrr2p, present in the Plasmodium falciparum genome and characterized the role of one of these factors, PfPrp16p. It is a member of DEAH-box protein family with nine collinear sequence motifs, a characteristic of helicase proteins. Experiments with the recombinantly expressed and purified PfPrp16 helicase domain revealed binding to RNA, hydrolysis of ATP as well as catalytic helicase activities. Expression of helicase domain with the C-terminal helicase-associated domain (HA2) reduced these activities considerably, indicating that the helicase-associated domain may regulate the PfPrp16 function. Localization studies with the PfPrp16 GFP transgenic lines suggested a role of its N‐terminal domain (1–80 amino acids) in nuclear targeting. Immunodepletion of PfPrp16p, from nuclear extracts of parasite cultures, blocked the second catalytic step of an in vitro constituted splicing reaction suggesting a role for PfPrp16p in splicing catalysis. Further we show by complementation assay in yeast that a chimeric yeast-Plasmodium Prp16 protein, not the full length PfPrp16, can rescue the yeast prp16 temperature‐sensitive mutant. These results suggest that although the role of Prp16p in catalytic step II is highly conserved among Plasmodium, human and yeast, subtle differences exist with regards to its associated factors or its assembly with spliceosomes. |
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