The Plasmodium Class XIV Myosin, MyoB, Has a Distinct Subcellular Location in Invasive and Motile Stages of the Malaria Parasite and an Unusual Light Chain

Myosin B (MyoB) is one of the two short class XIV myosins encoded in the Plasmodium genome. Class XIV myosins are characterized by a catalytic "head," a modified "neck," and the absence of a "tail" region. Myosin A (MyoA), the other class XIV myosin in Plasmodium, has been established as a component of the glideosome complex important in motility and cell invasion, but MyoB is not well characterized. We analyzed the properties of MyoB using three parasite species as follows: Plasmodium falciparum, Plasmodium berghei, and Plasmodium knowlesi. MyoB is expressed in all invasive stages (merozoites, ookinetes, and sporozoites) of the life cycle, and the protein is found in a discrete apical location in these polarized cells. In P. falciparum, MyoB is synthesized very late in schizogony/merogony, and its location in merozoites is distinct from, and anterior to, that of a range of known proteins present in the rhoptries, rhoptry neck or micronemes. Unlike MyoA, MyoB is not associated with glideosome complex proteins, including the MyoA light chain, myosin A tail domain-interacting protein (MTIP). A unique MyoB light chain (MLC-B) was identified that contains a calmodulin-like domain at the C terminus and an extended N-terminal region. MLC-B localizes to the same extreme apical pole in the cell as MyoB, and the two proteins form a complex. We propose that MLC-B is a MyoB-specific light chain, and for the short class XIV myosins that lack a tail region, the atypical myosin light chains may fulfill that role.

Characterization of a shorter recombinant polypeptide chain of bone morphogenetic protein 2 on osteoblast behaviour

BACKGROUND Recombinant bone morphogenetic protein two (rhBMP2) has been utilised for a variety of clinical applications in orthopaedic surgery and dental procedures. Despite its widespread use, concerns have been raised regarding its short half-life and transient bioactivity in vivo. Recent investigation aimed at developing rhBMP2 synthesized from a shorter polypeptide chain (108 amino acids) has been undertaken. METHODS The osteopromotive properties of BMP2 were investigated on cell behaviour. Five concentrations of rhBMP2_108 including 10, 50, 100, 200 and 500 ng/ml were compared to a commercially available rhBMP2 (100 ng/ml). Each of the working concentrations of rhBMP2_108 were investigated on MC3T3-E1 osteoblasts for their ability to induce osteoblast recruitment, proliferation and differentiation as assessed by alkaline phosphatase (ALP) staining, alizarin red staining, and real-time PCR for genes encoding ALP, osteocalcin (OCN), collagen-1 (COL-1) and Runx2. RESULTS The results demonstrate that all concentrations of rhBMP2_108 significantly improved cell recruitment and proliferation of osteoblasts at 5 days post seeding. Furthermore, rhBMP2_108 had the most pronounced effects on osteoblast differentiation. It was found that rhBMP2_108 had over a four fold significant increase in ALP activity at seven and 14 days post-seeding and the concentrations ranging from 50 to 200 ng/ml demonstrated the most pronounced effects. Analysis of real-time PCR for genes encoding ALP, OCN, COL-1 and Runx2 further confirmed dose-dependant increases at 14 days post-seeding. Furthermore, alizarin red staining demonstrated a concentration dependant increase in staining at 14 days. CONCLUSION The results from the present study demonstrate that this shorter polypeptide chain of rhBMP2_108 is equally as bioactive as commercially available rhBMP2 for the recruitment of progenitor cells by facilitating their differentiation towards the osteoblast lineage. Future in vivo study are necessary to investigate its bioactivity.