title: Generation of rodent malaria parasites with a high mutation rate by destructing proofreading activity of DNA polymerase δ.
abstract: Plasmodium falciparum malaria imposes a serious public health concern throughout the tropics. Although genetic tools are principally important to fully investigate malaria parasites, currently available forward and reverse tools are fairly limited. It is expected that parasites with a high mutation rate can readily acquire novel phenotypes/traits; however, they remain an untapped tool for malaria biology. Here, we generated a mutator malaria parasite (hereinafter called a 'malaria mutator'), using site-directed mutagenesis and gene transfection techniques. A mutator Plasmodium berghei line with a defective proofreading 3' → 5' exonuclease activity in DNA polymerase δ (referred to as PbMut) and a control P. berghei line with wild-type DNA polymerase δ (referred to as PbCtl) were maintained by weekly passage in ddY mice for 122 weeks. High-throughput genome sequencing analysis revealed that two PbMut lines had 175-178 mutations and a 86- to 90-fold higher mutation rate than that of a PbCtl line. PbMut, PbCtl, and their parent strain, PbWT, showed similar course of infection. Interestingly, PbMut lost the ability to form gametocytes during serial passages. We believe that the malaria mutator system could provide a novel and useful tool to investigate malaria biology.
authors.affiliation: Laboratory of Malariology, Research Institute for Microbial Diseases, Osaka University, Suita, Osaka 565-0871, Japan Department of Molecular Protozoology, Research Institute for Microbial Diseases, Osaka University, Suita, Osaka 565-0871, Japan.
authors.forename: Hajime
authors.initials: H
authors.lastname: Honma
authors.affiliation: Department of Parasitology, Graduate School of Medicine, Gunma University, Maebashi, Gunma 371-8511, Japan makotohirai@gunma-u.ac.jp kztanabe@biken.osaka-u.ac.jp.
authors.forename: Makoto
authors.initials: M
authors.lastname: Hirai
authors.affiliation: Department of Genome Informatics, Research Institute for Microbial Diseases, Osaka University, Suita, Osaka 565-0871, Japan.
authors.forename: Shota
authors.initials: S
authors.lastname: Nakamura
authors.affiliation: National Research Center for Protozoan Diseases, Obihiro University of Agriculture and Veterinary Medicine, Inada, Obihiro, Hokkaido 080-8555, Japan.
authors.forename: Hassan
authors.initials: H
authors.lastname: Hakimi
authors.affiliation: National Research Center for Protozoan Diseases, Obihiro University of Agriculture and Veterinary Medicine, Inada, Obihiro, Hokkaido 080-8555, Japan.
authors.forename: Shin-Ichiro
authors.initials: S
authors.lastname: Kawazu
authors.affiliation: Department of Molecular Protozoology, Research Institute for Microbial Diseases, Osaka University, Suita, Osaka 565-0871, Japan.
authors.forename: Nirianne M Q
authors.initials: NM
authors.lastname: Palacpac
authors.affiliation: Department of Parasitology, Graduate School of Medicine, Gunma University, Maebashi, Gunma 371-8511, Japan.
authors.forename: Hajime
authors.initials: H
authors.lastname: Hisaeda
authors.affiliation: Division of Medical Zoology, Department of Infection and Immunity, Jichi Medical University, Shimotsuke, Tochigi 329-0498, Japan.
authors.forename: Hiroyuki
authors.initials: H
authors.lastname: Matsuoka
authors.affiliation: Laboratory of Tropical Medicine and Parasitology, Institute of International Education and Research, Dokkyo Medical University, Shimotsuga, Tochigi 321-0293, Japan.
authors.forename: Satoru
authors.initials: S
authors.lastname: Kawai
authors.affiliation: Department of International Affairs and Tropical Medicine, Tokyo Women's Medical University School of Medicine, Kawada-cho, Shinjuku, Tokyo 162-8666, Japan.
authors.forename: Hiroyoshi
authors.initials: H
authors.lastname: Endo
authors.affiliation: Department of Genome Informatics, Research Institute for Microbial Diseases, Osaka University, Suita, Osaka 565-0871, Japan.
authors.forename: Teruo
authors.initials: T
authors.lastname: Yasunaga
authors.affiliation: Faculty of Medicine, University of Tsukuba, Ibaragi 305-8575, Japan.
authors.forename: Jun
authors.initials: J
authors.lastname: Ohashi
authors.affiliation: Department of International Affairs and Tropical Medicine, Tokyo Women's Medical University School of Medicine, Kawada-cho, Shinjuku, Tokyo 162-8666, Japan Department of Molecular and Cellular Parasitology, Juntendo University School of Medicine, Hongo, Bunkyo, Tokyo 113-8421, Japan.
authors.forename: Toshihiro
authors.initials: T
authors.lastname: Mita
authors.affiliation: Department of Molecular Protozoology, Research Institute for Microbial Diseases, Osaka University, Suita, Osaka 565-0871, Japan.
authors.forename: Toshihiro
authors.initials: T
authors.lastname: Horii
authors.affiliation: Neo-Morgan Laboratory, Inc., Nogawa, Miyamae, Kawasaki, Kanagawa 216-0001, Japan.
authors.forename: Mitsuru
authors.initials: M
authors.lastname: Furusawa
authors.affiliation: Laboratory of Malariology, Research Institute for Microbial Diseases, Osaka University, Suita, Osaka 565-0871, Japan Department of Molecular Protozoology, Research Institute for Microbial Diseases, Osaka University, Suita, Osaka 565-0871, Japan makotohirai@gunma-u.ac.jp kztanabe@biken.osaka-u.ac.jp.
authors.forename: Kazuyuki
authors.initials: K
authors.lastname: Tanabe
mesh_terms: D000818:Animals; D004258:DNA Polymerase III / Q000235:genetics* / Q000378:metabolism; D005260:Female; D008288:Malaria / Q000469:parasitology; D051379:Mice; D059645:Mutation Rate; D010962:Plasmodium berghei / Q000235:genetics* / Q000254:growth & development / Q000378:metabolism; D012733:Sex Differentiation
chemical_list: D004258:DNA Polymerase III
keywords: DNA polymerase δ; Plasmodium; genome sequencing; mutator