Apicomplexan parasites exhibit a unique form of substrate-dependent motility, gliding motility, which is essential during their invasion of host cells and during their spread between host cells. This process is dependent on actin filaments and myosin that are both located between the plasma membrane and two underlying membranes of the inner membrane complex. We have identified a protein complex in the apicomplexan parasite Toxoplasma gondii that contains the class XIV myosin required for gliding motility, TgMyoA, its associated light chain, TgMLC1, and two novel proteins, TgGAP45 and TgGAP50. We have localized this complex to the inner membrane complex of Toxoplasma, where it is anchored in the membrane by TgGAP50, an integral membrane glycoprotein. Assembly of the protein complex is spatially controlled and occurs in two stages. These results provide the first molecular description of an integral membrane protein as a specific receptor for a myosin motor, and further our understanding of the motile apparatus underlying gliding motility in apicomplexan parasites.

Toxoplasma gondii motility is powered by the myosin XIV motor complex, which consists of the myosin XIV heavy chain (MyoA), the myosin light chain (MLC1), GAP45, and GAP50, the membrane anchor of the complex. MyoA, MLC1, and GAP45 are initially assembled into a soluble complex, which then associates with GAP50, an integral membrane protein of the parasite inner membrane complex. While all proteins in the myosin XIV motor complex are essential for parasite survival, the specific role of GAP45 remains unclear. We demonstrate here that final assembly of the motor complex is controlled by phosphorylation of GAP45. This protein is phosphorylated on multiple residues, and by using mass spectroscopy, we have identified two of these, Ser 163

Membrane skeletons play an important role in the maintenance of cell shape and integrity in many cell types. In the protozoan parasite Toxoplasma gondii this function is performed by the subpellicular network, a resilient structure composed of tightly interwoven 10-nm filaments. We report here that this network is assembled at an early stage in the development of daughter parasites. The networks of immature and mature parasites differ dramatically with respect to their stability. Although in immature parasites the network is completely solubilized by detergent, the network in mature parasites is entirely detergent-resistant. Conversion of the detergent-labile to the detergent-resistant network occurs late in daughter cell development and appears to be coupled to proteolytic processing of the carboxyl terminus of TgIMC1, the major subunit of the network filaments. A single cysteine residue in the TgIMC1 carboxyl terminus was found to be essential for this processing event. The dramatic change in resistance to detergent extraction probably reflects an overall change in structural stability of the subpellicular network that accompanies maturation of daughter parasites and allows a switch from an assembly-competent but loose structure to one that is rigid and offers mechanical strength to the mature parasite.Membrane skeletons, composed of the plasma membrane and its organized underlying coat, are found in many cell types and are essential for mechanical strength and the maintenance of cell shape. Two widely divergent examples are the spectrinbased membrane skeletal system of the erythrocyte and the articulin-based system in the unicellular Euglena. In the erythrocyte, spectrin along with its associated proteins such as ankyrin form a meshwork underlying the plasma membrane (1). In the Euglena, two articulin proteins interact stoichiometrically to form a membrane skeletal system consisting of 40 interdigitating strips (2-4). Along with the articulins of the Euglena, a number of unique cytoskeletal systems have been described in protists, including the giardins (5, 6), assemblins (7), and tetrins (8) as well as the components of the subpellicular network, a novel cytoskeletal structure recently identified in the protozoan parasite Toxoplasma gondii (9).T. gondii is an obligate intracellular parasite that infects a wide range of nucleated cells. Human infection with the para-

A bystander effect typically refers to the death, altered growth or damage of cells that have not directly received chemotherapy or irradiation. Cancer cells derived from solid tumors readily undergo senescence in response to chemotherapeutic agents, prompting us to test for the existence of a senescence bystander effect. MCF-7 breast cancer cells were acutely exposed to Adriamycin to trigger senescence. Naïve MCF-7 cells, when cultured in conditioned media from senescent breast cancer cells, growth arrested despite mitogenic stimulation and exhibited SA-β-galactosidase activity, an enlarged cell size and stable upregulation of p21 WAF1 protein, collectively indicating a senescent state. In contrast, HCT-116 colon cancer cells, which also undergo p53-mediated senescence in response to acute AdR, did not undergo growth inhibition or senescence when cultured with conditioned media from senescent HCT-116 cells. Reciprocal experiments indicated that naïve HCT-116 cells, like MCF-7 cells, are susceptible to the growth inhibitory effects of a breast cancer-derived mediator, which is independent of residual drug in conditioned media. Our study reveals a novel action of Adriamycin, which may contribute to its potent anti-breast cancer activity and lead to the discovery of additional therapeutic targets for the exploitation of a senescence bystander effect.

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