Anopheles Midgut FREP1 Mediates Plasmodium Invasion

Zhang, Genwei; Niu, Guodong; Franca, Caio M.; Dong, Yuemei; Wang, Xiaohong; Butler, Noah S.; Dimopoulos, George; Li, Jun

doi:10.1074/jbc.m114.623165

Cited by 63 publications

(96 citation statements)

References 43 publications

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“…gambiae [14]. FREP1 was able to bind to P. falciparum [12]. Here, we determined the interaction between FREP1 and P. berghei .…”

Section: Resultsmentioning

confidence: 99%

“…Here, we determined the interaction between FREP1 and P. berghei . FREP1 protein was expressed in High Five insect cells and purified using Ni-NTA column as we reported previously [12]. P. berghei -infected mouse blood cell lysate was used to coat microplates for enzyme-linked immunosorbent assays (ELISA), with uninfected mouse blood cell lysate was used as a control.…”

Section: Resultsmentioning

confidence: 99%

“…We labelled polyclonal Ab against human α-tubulin with CF™ 568 dye, and followed the nonpermeable approach for immunofluorescence assays (IFA) [12] to investigate the interaction between Ab and ookinetes. P. falciparum ookinetes were cultured in vitro, deposited onto slide coverslips, and fixed with paraformaldehyde to acquire impermeable cells.…”

Section: Anti-α-tubulin Ab Bound To Ookinete Apical Invasion Apparatusmentioning

confidence: 99%

“…gambiae in Kenya [11]. Further investigation revealed that FREP1 bound to P. falciparum ookinetes to mediate their invasion [12]. Preventing Plasmodia from binding to FREP1 by small molecules inhibited malaria transmission [13].…”

Section: Introductionmentioning

confidence: 99%

“…Further investigation revealed that FREP1 bound to P. falciparum ookinetes to mediate their invasion [12]. Preventing Plasmodia from binding to FREP1 by small molecules inhibited malaria transmission [13]. Notably, Ab against FREP1 could inhibit multiple species of Plasmodium (e.g.…”

Section: Introductionmentioning

confidence: 99%

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Malaria transmission assisted by interaction betweenPlasmodium α-tubulin-1 andAnophelesFREP1 protein

Zhang

Niu

Perez

et al. 2019

Preprint

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Passage of Plasmodium through a mosquito midgut is essential for malaria transmission. FREP1, a peritrophic matrix protein in a mosquito midgut, binds to the parasite and mediates Plasmodium infection in Anopheles. The FREP1-mediated Plasmodium invasion pathway is highly conserved across multiple species of Plasmodium and Anopheles. Through pulldown, nine P. berghei proteins were co-precipitated with FREP1-conjugated beads. After cloning these nine genes from P. berghei and expressing them in insect cells, six of them were confirmed to interact with recombinant FREP1 protein. Among them, α-tubulin-1 and heat shock protein 70 (Hsp70) were highly conserved in Plasmodium species with >95% identity. Thus, P. falciparum α-tubulin-1 and Hsp70 were cloned and expressed in E. coli to stimulate antibody (Ab) in mice. Our results showed that anti-serum against P. falciparum α-tubulin-1 significantly inhibited P. falciparum transmission to An. gambiae, while Ab against P. falciparum Hsp70 serum did not. The polyclonal Ab against human α-tubulin did not interfere formation of ookinetes, however, significantly reduced the number of P. falciparum oocysts in An. gambiae midguts. Moreover, fluorescence microscope assays showed that anti-α-tubulin Ab bound to impermeable Plasmodium ookinete apical invasive apparatus. Therefore, we propose that the interaction between Anopheles FREP1 protein and Plasmodium α-tubulin-1 directs the ookinete invasive apparatus towards midgut peritrophic matrix for the efficient passage of the parasite. Anopheles FREP1 and Plasmodium α-tubulin-1 are potential targets for blocking malaria transmission to the mosquito host.AUTHOR SUMMARYThe molecular mechanisms of malaria transmission to mosquito are not well-understood. FREP1 proteins in mosquito midget PM has been proved to mediate malaria transmission by binding to parasite ookinetes. Here we reported that Plasmodium parasite α-tubulin-1 is an FREP1 binding partner. We initially identified the α-tubulin-1 through the FREP1-pulldown assay; Then we cloned P. falciparum α-tubulin-1, and demonstrated that the insect cell expressed recombinant Plasmodium α-tubulin-1 bound to FREP1 in vitro; Next, mouse anti-serum against P. falciparum α-tubulin-1 was found to inhibit P. falciparum transmission to An. gambiae. P. falciparum α-tubulin-1 shares >84% identical amino acid sequence with human α-tubulin, purified Ab against human α-tubulin significantly inhibited malaria transmission. Anti-human α-tubulin Ab did not interfere the gametocyte-to-ookinetes conversion. Final, we found that anti-α-tubulin Ab bound to the apical end of impermeable ookinetes. Structurally, ookinete invasive apparatus locates at the apical opening. Therefore, we propose that the interaction between Anopheles midgut FREP1 protein and Plasmodium apical α-tubulin-1 directs the ookinete invasive apparatus towards midgut PM for the efficient parasite invasion.

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“…gambiae [14]. FREP1 was able to bind to P. falciparum [12]. Here, we determined the interaction between FREP1 and P. berghei .…”

Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

Section: Anti-α-tubulin Ab Bound To Ookinete Apical Invasion Apparatusmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Malaria transmission assisted by interaction betweenPlasmodium α-tubulin-1 andAnophelesFREP1 protein

Zhang

Niu

Perez

et al. 2019

Preprint

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Active genetics comes alive

Bier

2022

BioEssays

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Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)-based "active genetic" elements developed in 2015 bypassed the fundamental rules of traditional genetics. Inherited in a super-Mendelian fashion, such selfish genetic entities offered a variety of potential applications including: gene-drives to disseminate gene cassettes carrying desired traits throughout insect populations to control disease vectors or pest species, allelic drives biasing inheritance of preferred allelic variants, neutralizing genetic elements to delete and replace or to halt the spread of gene-drives, splitdrives with the core constituent Cas9 endonuclease and guide RNA (gRNA) components inserted at separate genomic locations to accelerate assembly of complex arrays of genetic traits or to gain genetic entry into novel organisms (vertebrates, plants, bacteria), and interhomolog based copying systems in somatic cells to develop tools for treating inherited or infectious diseases. Here, we summarize the substantial advances that have been made on all of these fronts and look forward to the next phase of this rapidly expanding and impactful field.

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Structure of Aedes aegypti carboxypeptidase B1‐inhibitor complex uncover the disparity between mosquito and non‐mosquito insect carboxypeptidase inhibition mechanism

et al. 2021

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Metallocarboxypeptidases (MCPs) in the mosquito midgut play crucial roles in infection, as well as in mosquito dietary digestion, reproduction, and development. MCPs are also part of the digestive system of plant‐feeding insects, representing key targets for inhibitor development against mosquitoes/mosquito‐borne pathogens or as antifeedant molecules against plant‐feeding insects. Notably, some non‐mosquito insect B‐type MCPs are primarily insensitive to plant protease inhibitors (PPIs) such as the potato carboxypeptidase inhibitor (PCI; MW 4 kDa), an inhibitor explored for cancer treatment and insecticide design. Here, we report the crystal structure of Aedes aegypti carboxypeptidase‐B1 (CPBAe1)‐PCI complex and compared the binding with that of PCI‐insensitive CPBs. We show that PCI accommodation is determined by key differences in the active‐site regions of MCPs. In particular, the loop regions α6‐α7 (Leu242‐Ser250) and β8‐α8 (Pro269‐Pro280) of CPBAe1 are replaced by α‐helices in PCI‐insensitive insect Helicoverpa zea CPBHz. These α‐helices protrude into the active‐site pocket of CPBHz, restricting PCI insertion and rendering the enzyme insensitive. We further compared our structure with the only other PCI complex available, bovine CPA1‐PCI. The potency of PCI against CPBAe1 (Ki = 14.7 nM) is marginally less than that of bovine CPA1 (Ki = 5 nM). Structurally, the above loop regions that accommodate PCI binding in CPBAe1 are similar to that of bovine CPA1, although observed changes in proteases residues that interact with PCI could account for the differences in affinity. Our findings suggest that PCI sensitivity is largely dictated by structural interference, which broadens our understanding of carboxypeptidase inhibition as a mosquito population/parasite control strategy.

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Anopheles Midgut FREP1 Mediates Plasmodium Invasion

Cited by 63 publications

References 43 publications

Malaria transmission assisted by interaction betweenPlasmodium α-tubulin-1 andAnophelesFREP1 protein

Malaria transmission assisted by interaction betweenPlasmodium α-tubulin-1 andAnophelesFREP1 protein

Active genetics comes alive

Structure of Aedes aegypti carboxypeptidase B1‐inhibitor complex uncover the disparity between mosquito and non‐mosquito insect carboxypeptidase inhibition mechanism

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