Eukaryot Cell. 2008 Dec 1. [Epub ahead of print]
GAP45 PHOSPHORYLATION CONTROLS ASSEMBLY OF THE TOXOPLASMA MYOSIN XIV COMPLEX
Gilk SD, Gaskins E, Ward GE, Beckers CJ
Department of Cell and Developmental Biology, University of North Carolina, Chapel Hill, NC 27599; and Department of Microbiology and Molecular Genetics, University of Vermont, Burlington, VT 05404.
Toxoplasma gondii motility is powered by the myosin XIV motor complex which consists of myosin XIV heavy chain (MyoA), 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, using mass spectroscopy, we have identified two of these, Ser(163) and Ser(167). The importance of these phosphorylation events was determined by mutation of Ser(163) and Ser(167) to Glu and Ala residues to mimic phosphorylated and non-phosphorylated residues, respectively. Mutation of Ser(163) and Ser(167) to either Ala or Glu residues does not affect targeting of GAP45 to the inner membrane complex or its association with MyoA and MLC1. Mutation of Ser(163) and Ser(167) into Ala residues also does not affect assembly of the mutant GAP45 into the myosin motor complex. Mutation of Ser(163) and Ser(167) to Glu residues, however, prevents association of the MyoA-MLC1-GAP45 complex with GAP50. These observations indicate that phosphorylation of Ser(163) and Ser(167) in GAP45 controls the final step in assembly of the myosin XIV motor complex.
PMID: 19047362 [PubMed - as supplied by publisher]