| Abstract |
The causative agent for the debilitating disease malaria is the Plasmodium parasite.
Plasmodium spp is the only genus of Apicomplexa which possesses an additional
gene encoding actin besides the major actin isoform actin I. This second actin has
been termed actin II. It has been found to be expressed in male gametocytes and
ookinetes in the murine model strain P. berghei. Disruption resulted in an
impairment of the male gametocyte to exflagellate and a 50‐fold decrease in
ookinete formation, while females remained fertile.
In this study we explored the role of actin II in the mosquito stages. By performing
genetic crosses with a knockout strain of actin II (actII(‐)) and the wild type (wt) we
determined that heterozygous oocysts were formed, but did not develop further
than ~ three days, while the homozygous wt ookinetes developed into mature
oocysts. To analyse this defect further, we attempted to alter expression levels of
actin II by exchanging its promoter region with promoter regions from proteins
known to be expressed in the male gametocyte. Two mutant strains, cdpk4p‐actII
and setp‐actin II were constructed. While the cdpk4‐actII parasites did not show
any detectable phenotype, the setp‐actII mutant displayed a growth defect in the
asexual stages, even though its driving promoter is exclusively active in the sexual
stages. We also studied a pure heterozygous ookinete population with a parasite
strain producing only males and the actII(‐) female and observed that actin II was
not expressed in these ookinetes. The resulting oocysts were growth arrested and
did not undergo DNA replication. We followed the expression of GFP tagged actin
II and showed that the highest protein expression occurs within the first few hours
of zygote formation. These results show that actin II is a sex‐specific protein
required in the male gametogenesis however it is inherited from the female
gamete during zygote formation to complete the transmission cycle through the
mosquito.
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Characterisation of a second actin gene : analysing its function in the Plasmodium berghei mosquito midgut stages
