Plasmodium falciparum suppresses the host immune response by inducing the synthesis of insulin-like peptides (ILPs) in the mosquito Anopheles stephensi

Abstract

The insulin-like peptides (ILPs) and their respective signaling and regulatory pathways are highly conserved across phyla. In invertebrates, ILPs regulate diverse physiological processes, including metabolism, reproduction, behavior, and immunity. We previously reported that blood feeding alone induced minimal changes in ILP expression in Anopheles stephensi. However, ingestion of a blood meal containing human insulin or Plasmodium falciparum, which can mimic insulin signaling, leads to significant increases in ILP expression in the head and midgut, suggesting a potential role for AsILPs in the regulation of P. falciparum sporogonic development. Here, we show that soluble P. falciparum products, but not LPS or zymosan, directly induced AsILP expression in immortalized A. stephensi cells in vitro. Further, AsILP expression is dependent on signaling by the mitogen-activated protein kinase kinase/extracellular signal-regulated kinase (MEK/ERK) and phosphatidylinositol 3′-kinase (PI3K)/Akt branches of the insulin/insulin-like growth factor signaling (IIS) pathway. Inhibition of P. falciparum-induced ILPs in vivo decreased parasite development through kinetically distinct effects on mosquito innate immune responses. Specifically, knockdown of AsILP4 induced early expression of immune effector genes (1–6 hours after infection), a pattern associated with significantly reduced parasite abundance prior to invasion of the midgut epithelium. In contrast, knockdown of AsILP3 increased later expression of the same genes (24 hours after infection), a pattern that was associated with significantly reduced oocyst development. These data suggest that P. falciparum parasites alter the expression of mosquito AsILPs to dampen the immune response and facilitate their development in the mosquito vector.

Two insulin-like peptides differentially regulate malaria parasite infection in the mosquito through effects on intermediary metabolism

“…Accordingly, we used this concentration of ILPs in our feeding assays. Provision of 170 pM ILP4 significantly increased the prevalence (proportion of mosquitoes infected) of infection (Figure 1A) from 57.8 to 80.8% and the intensity (oocysts/midgut) of infection (Figure 1B) from 1.11 to 2.13 oocysts/midgut when compared with controls, as predicted by our recent work [13]. Surprisingly, feeding of 170 pM ILP3 significantly decreased the prevalence of infection from 62.2 to 54.3% (Figure 1C), but had no effect on the intensity of infection relative to controls (Figure 1D).…”

“…We have previously shown that knockdown of either ILP3 or ILP4 can significantly reduce the prevalence and intensity of P. falciparum infection in A. stephensi , suggesting that mosquito ILPs are immunosuppressive [13]. While the primary aim of the present study was to determine the mechanisms by which these ILPs modulate resistance to infection, we first sought to extend previous findings by characterizing the phenotypic effects of these peptides using ILP provisioning as a complement to our knockdown studies.…”

“…We previously showed that knockdown of either of two infection-induced ILPs, ILP3 or ILP4, in the midgut decreased P. falciparum infectivity through kinetically distinct effects on A. stephensi innate immune defenses [13]. Specifically, knockdown of ILP4 increased early expression of antiparasite genes (1–6 h post-infection) and increased killing of ookinetes prior to invasion, whereas knockdown of ILP3 increased anti-parasite gene expression at a later time (24 h post-infection), boosting killing of parasites during and after invasion [13]. While we predicted that decreased P. falciparum infectivity following ILP knockdown was due, at least in part, to increased expression of antiparasite effector genes in the midgut, the specific mechanisms by which ILPs regulate mosquito resistance to infection remained unconfirmed.…”

Abscisic acid induces a transient shift in signaling that enhances NF-κB-mediated parasite killing in the midgut of Anopheles stephensi without reducing lifespan or fecundity

“…2). This repression was consistent with ABA’s effect on infection given that knockdown of ILP3 and ILP4 is also associated with reduced P. falciparum infection in A. stephensi [12]. Addition of 100 nM ABA to a P. falciparum -infected blood meal had no effect on phosphorylation of ERK, JNK or p38 MAPK relative to controls (Fig.…”

“…Data were normalized to transcript levels for A. stephensi ribosomal s7 protein, a18s rDNA, and control levels as previously described [12]. nos , defensin , apl1 , tep1 , and lrim transcript levels were determined by qRT-PCR and analyzed as described previously [1].…”