Proteins that control the geometry of microtubules at the ends of cilia.

Cilia, essential motile and sensory organelles, have several compartments: the basal body, transition zone, and the middle and distal axoneme segments. The distal segment accommodates key functions, including cilium assembly and sensory activities. While the middle segment contains doublet microtubules (incomplete B-tubules fused to complete A-tubules), the distal segment contains only A-tubule extensions, and its existence requires coordination of microtubule length at the nanometer scale.

FAP206 is a microtubule-docking adapter for ciliary radial spoke 2 and dynein c.

Radial spokes are conserved macromolecular complexes that are essential for ciliary motility. A triplet of three radial spokes, RS1, RS2, and RS3, repeats every 96 nm along the doublet microtubules. Each spoke has a distinct base that docks to the doublet and is linked to different inner dynein arms.

Discovery and functional evaluation of ciliary proteins in Tetrahymena thermophila.

The ciliate Tetrahymena thermophila is an excellent model system for the discovery and functional studies of ciliary proteins. The power of the model is based on the ease with which cilia can be purified in large quantities for fractionation and proteomic identification, and the ability to knock out any gene by homologous DNA recombination. Here, we include methods used by our laboratories for isolation and fractionation of cilia, in vivo tagging and localization of ciliary proteins, and the evaluation of ciliary mutants.

A role for the membrane in regulating Chlamydomonas flagellar length.

Flagellar assembly requires coordination between the assembly of axonemal proteins and the assembly of the flagellar membrane and membrane proteins. Fully grown steady-state Chlamydomonas flagella release flagellar vesicles from their tips and failure to resupply membrane should affect flagellar length. To study vesicle release, plasma and flagellar membrane surface proteins were vectorially pulse-labeled and flagella and vesicles were analyzed for biotinylated proteins.

Recording and analyzing IFT in Chlamydomonas flagella.

The transport of materials to and from the cell body and tips of eukaryotic flagella and cilia is carried out by a process called intraflagellar transport, or IFT. This process is essential for the assembly and maintenance of cilia and flagella: in the absence of IFT, cilia cannot assemble and, if IFT is arrested in ciliated cells, the cilia disassemble. The major IFT complex proteins and the major motor proteins, kinesin-2 and osm-3 (which transport particles from the cell body to ciliary tips) and cytoplasmic dynein 1b (which transports particles from ciliary tips to the cell body) have been identified.

FLI-1 Flightless-1 and LET-60 Ras control germ line morphogenesis in C. elegans.

Background: In the C. elegans germ line, syncytial germ line nuclei are arranged at the cortex of the germ line as they exit mitosis and enter meiosis, forming a nucleus-free core of germ line cytoplasm called the rachis. Molecular mechanisms of rachis formation and germ line organization are not well understood.

Paclitaxel-induced microtubule stabilization causes mitotic block and apoptotic-like cell death in a paclitaxel-sensitive strain of Saccharomyces cerevisiae.

Wild-type Saccharomyces cerevisiae tubulin does not bind the anti-mitotic microtubule stabilizing agent paclitaxel. Previously, we introduced mutations into the S. cerevisiae gene for beta-tubulin that imparted paclitaxel binding to the protein, but the mutant strain was not sensitive to paclitaxel and other microtubule-stabilizing agents, due to the multiple ABC transporters in the membranes of budding yeast.

Intraflagellar transport (IFT) during assembly and disassembly of Chlamydomonas flagella.

Intraflagellar transport (IFT) of particles along flagellar microtubules is required for the assembly and maintenance of eukaryotic flagella and cilia. In Chlamydomonas, anterograde and retrograde particles viewed by light microscopy average 0.12-microm and 0.

Defective flagellar assembly and length regulation in LF3 null mutants in Chlamydomonas.

Four long-flagella (LF) genes are important for flagellar length control in Chlamydomonas reinhardtii. Here, we characterize two new null lf3 mutants whose phenotypes are different from previously identified lf3 mutants. These null mutants have unequal-length flagella that assemble more slowly than wild-type flagella, though their flagella can also reach abnormally long lengths.

Disulfide bond formation promotes the cis- and trans-dimerization of the E-cadherin-derived first repeat.

Cadherin is a cell adhesion molecule crucial for epithelial and endothelial cell monolayer integrity. The previously solved x-ray crystallographic structure of the E-CAD12 cis-dimer displayed an unpaired Cys(9), which protruded away from the Cys(9) on the other protomer. To investigate the possible biological function of Cys(9) within the first repeat (the E-cadherin-derived N-terminal repeat), E-CAD1 was overexpressed and secreted into the periplasmic space of Escherichia coli cells.