Biochemical And Morphological Analysis Of Cytoskeletal Proteins And Trail Secretion During Gliding Motility In The Marine Benthic Diatom, Amphora Coffeaeformis

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Doctor of Philosophy (Ph.D.)


Cell Biology and Anatomy


Gliding motility was investigated in the motile marine diatom, Amphora coffeaeformis (Agardh) Kutz. Early attempts to identify cytoskeletal organelles by ultrastructural means proved fruitless. Vesicles, possibly associated with motility and secretory in nature, were found in the area above the raphe canal. Scanning electron microscopy revealed the presence of extracellular material extending from the raphe canal to the substratum in cultures allowed to settle for only five minutes. An assay for motility was designed to measure per cent motile cells and motility rate, using a video recording apparatus attached to a light microscope. Various anti-cytoskeletal drugs were used to measure inhibition of both parameters. Anti-microfilament drugs found effective in inhibiting motility after two hours incubation were cytochalasins D and E. Podophyllotoxin and vinblastine were the only anti-microtubule drugs found effective under these conditions. Similarly, an assay for secretion of soluble polymer (> 1800 daltons) was devised to measure secretion when motility was inhibited. Monensin, a sodium-specific ionophore which has been shown to stimulate secretion, both inhibited motility and stimulated secretion as measured by the assay, as did cytochalasin E and vinblastine. The inhibition of motility was reversible in all cases. Colchicine had no detectable effect on motility, but enhanced secretion. Attempts were made to isolate an actin-like protein from whole-cell homogenates using ('35)S-labelled cells and either myosin-affinity extraction, copolymerizing labeled protein with exogenously added rabbit muscle G-actin, or immunoblot analysis with rabbit antiactin serum. All attempts were unsuccessful. It is concluded that cytoskeletal proteins are involved with gliding motility, and secretion of extracellular polymer is not positively correlated with motility in Amphora coffeaeformis. The failure to identify actin unequivocally may reflect the inability of actin from this organism to react with associated proteins of mammalian origin.


Biophysics, Medical

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