title: Biophysics of malarial parasite exit from infected erythrocytes.
abstract: Upon infection and development within human erythrocytes, P. falciparum induces alterations to the infected RBC morphology and bio-mechanical properties to eventually rupture the host cells through parasitic and host derived proteases of cysteine and serine families. We used previously reported broad-spectrum inhibitors (E64d, EGTA-AM and chymostatin) to inhibit these proteases and impede rupture to analyze mechanical signatures associated with parasite escape. Treatment of late-stage iRBCs with E64d and EGTA-AM prevented rupture, resulted in no major RBC cytoskeletal reconfiguration but altered schizont morphology followed by dramatic re-distribution of three-dimensional refractive index (3D-RI) within the iRBC. These phenotypes demonstrated several-fold increased iRBC membrane flickering. In contrast, chymostatin treatment showed no 3D-RI changes and caused elevated fluctuations solely within the parasitophorous vacuole. We show that E64d and EGTA-AM supported PV breakdown and the resulting elevated fluctuations followed non-Gaussian pattern that resulted from direct merozoite impingement against the iRBC membrane. Optical trapping experiments highlighted reduced deformability of the iRBC membranes upon rupture-arrest, more specifically in the treatments that facilitated PV breakdown. Taken together, our experiments provide novel mechanistic interpretations on the role of parasitophorous vacuole in maintaining the spherical schizont morphology, the impact of PV breakdown on iRBC membrane fluctuations leading to eventual parasite escape and the evolution of membrane stiffness properties of host cells in which merozoites were irreversibly trapped, recourse to protease inhibitors. These findings provide a comprehensive, previously unavailable, body of information on the combined effects of biochemical and biophysical factors on parasite egress from iRBCs.
authors.affiliation: Singapore-MIT Alliance for Research and Technology Centre, Singapore, Singapore.
authors.forename: Rajesh
authors.initials: R
authors.lastname: Chandramohanadas
authors.forename: YongKeun
authors.initials: Y
authors.lastname: Park
authors.forename: Lena
authors.initials: L
authors.lastname: Lui
authors.forename: Ang
authors.initials: A
authors.lastname: Li
authors.forename: David
authors.initials: D
authors.lastname: Quinn
authors.forename: Kingsley
authors.initials: K
authors.lastname: Liew
authors.forename: Monica
authors.initials: M
authors.lastname: Diez-Silva
authors.forename: Yongjin
authors.initials: Y
authors.lastname: Sung
authors.forename: Ming
authors.initials: M
authors.lastname: Dao
authors.forename: Chwee Teck
authors.initials: CT
authors.lastname: Lim
authors.forename: Peter Rainer
authors.initials: PR
authors.lastname: Preiser
authors.forename: Subra
authors.initials: S
authors.lastname: Suresh
mesh_terms: D000818:Animals; D001703:Biophysics; D004912:Erythrocytes / Q000469:parasitology*; D010963:Plasmodium falciparum / Q000502:physiology*