Antigenicity and immunogenicity of PvRALP1, a novel Plasmodium vivax rhoptry neck protein.
BACKGROUND
Proteins secreted from the rhoptry in Plasmodium merozoites are associated with the formation of tight junctions and parasitophorous vacuoles during invasion of erythrocytes and are sorted within the rhoptry neck or bulb. Very little information has been obtained to date about Plasmodium vivax rhoptry-associated leucine (Leu) zipper-like protein 1 (PvRALP1; PVX_096245), a putative rhoptry protein. PvRALP1 contains a signal peptide, a glycine (Gly)/glutamate (Glu)-rich domain, and a Leu-rich domain, all of which are conserved in other Plasmodium species.


METHODS
Recombinant PvRALP1s were expressed as full-length protein without the signal peptide (PvRALP1-Ecto) and as truncated protein consisting of the Gly/Glu- and Leu-rich domains (PvRALP1-Tr) using the wheat germ cell-free expression system. The immunoreactivity to these two fragments of recombinant PvRALP1 protein in serum samples from P. vivax-infected patients and immunized mice, including analysis of immunoglobulin G (IgG) subclasses, was evaluated by enzyme-linked immunosorbent assay or protein microarray technology. The subcellular localization of PvRALP1 in blood stage parasites was also determined.


RESULTS
Recombinant PvRALP1-Ecto and PvRALP1-Tr proteins were successfully expressed, and in serum samples from P. vivax patients from the Republic of Korea, the observed immunoreactivities to these proteins had 58.9% and 55.4% sensitivity and 95.0% and 92.5% specificity, respectively. The response to PvRALP1 in humans was predominantly cytophilic antibodies (IgG1 and IgG3), but a balanced Th1/Th2 response was observed in mice. Unexpectedly, there was no significant inverse correlation between levels of parasitaemia and levels of antibody against either PvRALP1-Ecto (R2=0.11) or PvRALP1-Tr (R2=0.14) antigens. PvRALP1 was localized in the rhoptry neck of merozoites, and this was the first demonstration of the localization of this protein in P. vivax.


CONCLUSIONS
This study analysed the antigenicity and immunogenicity of PvRALP1 and suggested that PvRALP1 may be immunogenic in humans during parasite infection and might play an important role during invasion of P. vivax parasites.
Department of Medical Environmental Biology and Tropical Medicine, Kangwon National University School of Medicine, Hyoja2-dong, Chuncheon, Gangwon-do, 200-701, Republic of Korea. chengyang1127@gmail.com.|Laboratory of Malaria and Vector Research, National Institute of Allergy and Infectious Diseases (NIAID), National Institutes of Health (NIH), Rockville, MD, 20852, USA. chengyang1127@gmail.com.
Yang
Y
Cheng
Department of Medical Environmental Biology and Tropical Medicine, Kangwon National University School of Medicine, Hyoja2-dong, Chuncheon, Gangwon-do, 200-701, Republic of Korea. yxlijian@163.com.|Department of Parasitology, College of Basic Medicine, Hubei University of Medicine, Hubei, 442000, China. yxlijian@163.com.
Jian
J
Li
Laboratory of Malaria and Vector Research, National Institute of Allergy and Infectious Diseases (NIAID), National Institutes of Health (NIH), Rockville, MD, 20852, USA. ito.daisuke.mh@ehime-u.ac.jp.|Division of Malaria Research, Proteo-Science Center, Ehime University, Matsuyama, Ehime, 790-8577, Japan. ito.daisuke.mh@ehime-u.ac.jp.
Daisuke
D
Ito
Department of Molecular and Cellular Biochemistry, School of Medicine, Kangwon National University, Chuncheon, Gangwon-do, 200-701, Republic of Korea. Kong0131@hanmail.net.
Deok-Hoon
DH
Kong
Department of Molecular and Cellular Biochemistry, School of Medicine, Kangwon National University, Chuncheon, Gangwon-do, 200-701, Republic of Korea. ksha@kangwon.ac.kr.
Kwon-Soo
KS
Ha
Department of Medical Environmental Biology and Tropical Medicine, Kangwon National University School of Medicine, Hyoja2-dong, Chuncheon, Gangwon-do, 200-701, Republic of Korea. lufeng981@hotmail.com.|Key Laboratory of Parasitic Disease Control and Prevention (Ministry of Health), and Jiangsu Provincial Key Laboratory of Parasite Molecular Biology, Jiangsu Institute of Parasitic Diseases, Wuxi, Jiangsu Province, People's Republic of China. lufeng981@hotmail.com.
Feng
F
Lu
Department of Medical Environmental Biology and Tropical Medicine, Kangwon National University School of Medicine, Hyoja2-dong, Chuncheon, Gangwon-do, 200-701, Republic of Korea. jmswbo@gmail.com.|Department of Clinical Laboratory, The First Affiliated Hospital of Anhui Medical University, Hefei, Anhui, People's Republic of China. jmswbo@gmail.com.
Bo
B
Wang
Mahidol Vivax Research Unit, Faculty of Tropical Medicine, Mahidol University, Bangkok, 10400, Thailand. jetsumon.pra@mahidol.ac.th.
Jetsumon
J
Sattabongkot
Department of Laboratory Medicine, College of Medicine, Korea University Guro Hospital, Seoul, 152-703, Republic of Korea. malarim@korea.ac.kr.
Chae Seung
CS
Lim
Division of Malaria Research, Proteo-Science Center, Ehime University, Matsuyama, Ehime, 790-8577, Japan. tsuboi.takafumi.mb@ehime-u.ac.jp.
Takafumi
T
Tsuboi
Department of Medical Environmental Biology and Tropical Medicine, Kangwon National University School of Medicine, Hyoja2-dong, Chuncheon, Gangwon-do, 200-701, Republic of Korea. ethan@kangwon.ac.kr.
Eun-Taek
ET
Han
D000293:Adolescent; D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D000818:Animals; D004797:Enzyme-Linked Immunosorbent Assay; D006801:Humans; D007074:Immunoglobulin G / Q000276:immunology*; D016780:Malaria, Vivax / Q000276:immunology / Q000469:parasitology; D052940:Merozoites / Q000276:immunology; D051379:Mice; D008807:Mice, Inbred BALB C; D008875:Middle Aged; D010966:Plasmodium vivax / Q000235:genetics* / Q000276:immunology* / Q000378:metabolism; D040081:Protein Array Analysis; D015800:Protozoan Proteins / Q000276:immunology*; D011817:Rabbits; D011994:Recombinant Proteins / Q000276:immunology; D056910:Republic of Korea; D055815:Young Adult
D007074:Immunoglobulin G; D015800:Protozoan Proteins; D011994:Recombinant Proteins