Laboratory of Protozoology
Graduate School of Science
Professor

Shiroh Iwanaga

iwanaga

Specially Appointed
Assistant Professor

Mai Nakashima

mainakashima

URL: https://malaria.biken.osaka-u.ac.jp/

Malaria is one of the world's threemajor infectious diseases caused by
Plasmodium species. Malaria parasite
isaneukaryotic
parasitethatparasitizes the host body in a morediverse range of ways than viral orbacterial pathogens. Their parasitic
strategies were established after along evolutionary struggle with hostand are beyond our imagination. Weare interested in how parasitism isestablished through the interaction
between molecules of parasites andhost cells, including immune cells. We
have developed our own genetic
manipulation techniques, such as
CRISPR/Cas, which can be used as
tools, and are working to reveal theelegant parasitization strategies.
Development of Plasmodium
artificial chromosomes and
CRISPR/Cas9 system

Genetic engineering technology formalaria parasite is essential for its
molecular biological research. However,parasite has been known as one of themost difficult organisms to geneticallymanipulate due to its parasitism in hostcells (erythrocytes), which makes genetransfer difficult, and the low efficiency ofincorporating foreign genes into its
chromosomes. On the other hand, we
identified centromeres responsible forchromosome segregation based onparasiteʼs genome information andsucceeded in creating an artificial
chromosome by combining telomere andreplication origin with centromere. The
artificial chromosome behaves like real
chromosome within parasites and aremaintained stably as they are equallydistributed into daughter parasites, whichenables efficient production of transgenicparasites. In addition, it has been foundthat under culture, the mature schizont
degrades the erythrocyte membrane justbefore its egress and temporarily remains
in the thin membrane, and by utilizing
this phenomenon, highly efficienttransfection method has beensuccessfully developed. Furthermore, bymounting CRISPR/Cas9 on artificial
chromosomes and combining it with thehigh-efficiency transfection method, we
have succeeded in developing a methodthat dramatically improves the efficiency
of foreign gene incorporation. It is now
possible to produce recombinantparasites in about two weeks with anefficiency of more than 90%, thus solving
a conventional technical limitation.
Interaction between surface
antigens of infected erythrocyte
and host receptors

Of human malaria parasites, P.
falciparum causes the most severe
symptoms in humans. During erythrocyte
infection, P. falciparum transports to andpresents its own proteins on the surface
of infected erythrocytes. PfEMP1 andRIFIN have been identified as
erythrocyte surface molecules. CD36,
ICAM1 and EPCR on vascularendothelial cells have been identified asreceptors for PfEMP1. P. falciparum
binds to and embolisms cerebral vascularendothelium via binding of PfEMP1 tothese molecules, causing severe coma
and brain damage (cerebral malaria).
Our collaborative work on RIFIN has
also shown that LILRB1, an
immunosuppressive receptor widely
expressed on immune cells, is a receptor
for RIFIN, and that parasite uses RIFN
to suppress host immunity in favor ofparasitization.Approximately 60 PfEMP1genes and 150-200 RIFIN genes areencoded on the genome of P. falciparum.
However, only ~20 surface antigens havebeen found to have a known function,
while the rest remain unknown. We are

currently analyzing the functions andhost receptors of remained PfEMP1 andRIFINs comprehensively using newtechnologies such as Cas12-Ultra andCas12k and artificial chromosome
technology. Through these studies, wehope to elucidate the elegant parasitic
strategies of parasites, leading to thedevelopment of vaccines and treatments
for cerebral malaria.

(A)
(A)
(B)

(B)

Red blood cell infected with P. falciparum
GFP-expressed sporozites

Host cell(C)
Infected RBC (iRBC)(D)（C）Image of Host-iRBC interaction（D）IFA of Surface antigens on iRBCs

Basic biological research on infectious diseases
can be based on academic interest andapplication. If you are interested in both ofthem, letʼs work on malaria research together
with us!

Department of Biological Sciences
Graduate School of Science, Osaka University
Yamadaoka 3-1. Suita, Osaka, 565-0871, Japan
TEL: +81-6-6879-8363, 8364
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