Plasmodium sporozoites require the protein B9 to invade hepatocytes

Abstract

Plasmodium sporozoites are transmitted to a mammalian host during blood feeding by an infected mosquito and invade hepatocytes for initial replication of the parasite in the liver. This leads to the release of thousands of merozoites into the blood circulation and initiation of the pathogenic blood stages of malaria. Merozoite invasion of erythrocytes has been well characterized at the molecular and structural levels. In sharp contrast, the molecular mechanisms of sporozoite invasion of hepatocytes are poorly characterized. Here we report a new role during sporozoite entry for the B9 protein, a member of the 6-cysteine domain protein family. Using genetic tagging and gene deletion approaches in rodent malaria parasites, we show that B9 is secreted from sporozoite micronemes and is required for productive invasion of hepatocytes. Structural modelling indicates that the N-terminus of B9 forms a beta-propeller domain structurally related to CyRPA, a cysteine-rich protein forming an invasion complex with Rh5 and RIPR in P. falciparum merozoites. We provide evidence that the beta-propeller domain of B9 is essential for protein function during sporozoite entry and interacts with P36 and P52, both also essential for productive invasion of hepatocytes. Our results suggest that, despite using distinct sets of parasite and host entry factors, Plasmodium sporozoites and merozoites may share common structural modules to assemble protein complexes for invasion of host cells.

Mid-Liver Stage Arrest of Plasmodium falciparum Schizonts in Primary Porcine Hepatocytes

“…Sporozoites with P52/p36 deletions are incapable of productively invading hepatocytes resulting in near-complete attenuation during early liver stage in Pb and Py , as well as Pf ( Ishino et al, 2005 ; van Dijk et al, 2005 ; Labaied et al, 2007 ; VanBuskirk et al, 2009 ; Ploemen et al, 2012 ; Manzoni et al, 2017 ). Finally, the function of B9 remains more controversial in different Plasmodia species: while required in Pb for productive hepatocyte invasion, this is not the case for Pf invasion into human hepatocytes despite structural similarity ( Annoura et al, 2014 ; Fernandes et al, 2021 ). As schizonts are detected in porcine hepatocytes on day three and (to a lesser amount) on day five, this challenges this prevailing theory that a functional, intact, PVM is required for progression beyond early-stage Pf infection.…”

Plasmodium 6-Cysteine Proteins: Functional Diversity, Transmission-Blocking Antibodies and Structural Scaffolds

“…PfB9 has three 6-cysteine domains and a predicted GPI anchor ( Figure 1B ). It is predicted to localize to the plasma membrane of liver stage parasites ( Annoura et al., 2014 ) and the micronemes of sporozoites in P. berghei ( Fernandes et al., 2021 ). The function of PfB9 has not been precisely defined, but knockout parasites display growth arrest in the liver and a lack of parasitophorous vacuole markers, indicating a critical role for PfB9 in liver-stage development and parasitophorous vacuole integrity ( Annoura et al., 2014 ).…”