The knob protein KAHRP assembles into a ring-shaped structure that underpins virulence complex assembly

Area of Science:

• Cell Biology
• Parasitology
• Microscopy

Background:

• Plasmodium falciparum infection causes severe malaria by inducing infected red blood cells (RBCs) to adhere to blood vessels.
• This adhesion is mediated by protein complexes, known as knobs, on the RBC surface, which display the P. falciparum Erythrocyte Membrane Protein-1 (PfEMP1) virulence antigen.

Purpose of the Study:

• To spatially and temporally map the delivery and assembly of knob components, specifically KAHRP and PfEMP1, to the RBC membrane skeleton.
• To elucidate the role of KAHRP in knob structure formation and the involvement of host cell actin remodeling in the assembly process.

Main Methods:

• Correlative STochastic Optical Reconstruction Microscopy-Scanning Electron Microscopy (STORM-SEM) was developed and utilized.
• Electron tomography of negatively-stained membranes was employed to visualize scaffold structures.
• Experiments involving cytochalasin D were performed to assess the role of actin remodeling.

Main Results:

• KAHRP is delivered as individual modules that assemble into ring-shaped structures around dimpled disks on the RBC membrane.
• A spiral scaffold underpins the assembled knobs, and KAHRP's C-terminal region is essential for proper knob and scaffold formation.
• Host cell actin remodeling is critical for knob assembly, as demonstrated by cytochalasin D treatment blocking the process.
• PfEMP1 appears to be delivered to the RBC membrane and subsequently inserted into existing knob structures.

Conclusions:

• KAHRP plays a vital role in the physical assembly of the knob structure, including the underlying spiral scaffold.
• Host cell actin remodeling is an important factor in the assembly of the P. falciparum virulence complex.
• PfEMP1 is likely inserted laterally into the knob structure after its delivery to the RBC membrane.