SummaryA major part of virulence for Plasmodium falciparum malaria infection, the most lethal parasitic disease of humans, results from increased rigidity and adhesiveness of infected host red cells. These changes are caused by parasite proteins exported to the erythrocyte using novel trafficking machinery assembled in the host cell. To understand these unique modifications, we used a large-scale gene knockout strategy combined with functional screens to identify proteins exported into parasite-infected erythrocytes and involved in remodeling these cells. Eight genes were identified encoding proteins required for export of the parasite adhesin PfEMP1 and assembly of knobs that function as physical platforms to anchor the adhesin. Additionally, we show that multiple proteins play a role in generating increased rigidity of infected erythrocytes. Collectively these proteins function as a pathogen secretion system, similar to bacteria and may provide targets for antivirulence based therapies to a disease responsible for millions of deaths annually.

The insulin-like growth factors (IGFs) are essential for development; bioavailable IGF is tightly regulated by six related IGF-binding proteins (IGFBPs). Igfbp5 is the most conserved and is developmentally up-regulated in key lineages and pathologies; in vitro studies suggest that IGFBP-5 functions independently of IGF interaction. Genetic ablation of individual Igfbps has yielded limited phenotypes because of substantial compensation by remaining family members. Therefore, to reveal Igfbp5 actions in vivo, we generated lines of transgenic mice that ubiquitously overexpressed Igfbp5 from early development. Significantly increased neonatal mortality, reduced female fertility, whole-body growth inhibition, and retarded muscle development were observed in Igfbp5-overexpressing mice. The magnitude of the response in individual transgenic lines was positively correlated with Igfbp5 expression. Circulating IGFBP-5 concentrations increased a maximum of only 4-fold, total and free IGF-I concentrations increased up to 2-fold, and IGFBP-5 was detected in high Mr complexes; however, no detectable decrease in the proportion of free IGF-I was observed. Thus, despite only modest changes in IGF and IGFBP concentrations, the Igfbp5-overexpressing mice displayed a phenotype more extreme than that observed for other Igfbp genetic models. Although growth retardation was obvious prenatally, maximal inhibition occurred postnatally before the onset of growth hormone-dependent growth, regardless of Igfbp5 expression level, revealing a period of sensitivity to IGFBP-5 during this important stage of tissue programming.T he insulin-like growth factors (IGF-I and -II) are essential for growth and development (1). Six high-affinity IGF-binding proteins (IGFBP-1 to IGFBP-6; refs. 2 and 3) strictly orchestrate IGF action. Despite their considerable sequence homology, each exhibits a discrete expression pattern and possesses an individual subset of motifs, signifying that although IGFBPs have common actions, they may also have unique properties.IGFBP-5 is the most conserved of the IGFBPs (4) and has been highlighted as a focal regulatory factor during the development of several key cell lineages, e.g., myoblasts (5) and neural cells (6, 7). In mice, Igfbp5 is expressed in the embryo from early development, principally in the myotomal component of the somites and developing central nervous system (8). Postnatally, serum IGFBP-5, in common with IGFBP-3, forms a ternary complex with IGF-I or IGF-II and the acid-labile subunit (9). Igfbp5 is up-regulated in the aggressive pediatric cancer, rhabdomyosarcoma (10), in the progression of prostate cancers to androgen independence (11), and in smooth muscle-derived uterine leiomyoma (12), indicating a function in neoplasia.IGFBP-5 initially binds IGFs with high affinity, principally by an N-terminal motif (13), and inhibits IGF activity by preventing IGF interaction with the type 1 receptor. It is further subject to regulated posttranslational modifications (3) to induce conformational changes that dec...

Our understanding of the basis of severe disease in malaria is incomplete. It is clear that pathology is in part related to the pro-inflammatory nature of the host response but a number of other factors are also thought to be involved, including the interaction between infected erythrocytes and endothelium. This is a complex system involving several host receptors and a major parasite-derived variant antigen (PfEMP1) expressed on the surface of the infected erythrocyte membrane. Previous studies have suggested a role for ICAM-1 in the pathology of cerebral malaria, although these have been inconclusive. In this study we have examined the cytoadherence patterns of 101 patient isolates from varying clinical syndromes to CD36 and ICAM-1, and have used variant ICAM-1 proteins to further characterise this adhesive phenotype. Our results show that increased binding to CD36 is associated with uncomplicated malaria while ICAM-1 adhesion is raised in parasites from cerebral malaria cases.

Sequestration of Plasmodium falciparum‐infected erythrocytes (IE) within the brain microvasculature is a hallmark of cerebral malaria (CM). Using a microchannel flow adhesion assay with TNF‐activated primary human microvascular endothelial cells, we demonstrate that IE isolated from Malawian paediatric CM cases showed increased binding to brain microvascular endothelial cells compared to IE from uncomplicated malaria (UM) cases. Further, UM isolates showed significantly greater adhesion to dermal than to brain microvascular endothelial cells. The major mediator of parasite adhesion is P. falciparum erythrocyte membrane protein 1, encoded by var genes. Higher levels of var gene transcripts predicted to bind host endothelial protein C receptor (EPCR) and ICAM‐1 were detected in CM isolates. These data provide further evidence for differential tissue binding in severe and uncomplicated malaria syndromes, and give additional support to the hypothesis that CM pathology is based on increased cytoadherence of IE in the brain microvasculature.