Studies On The Microtubular Organelle And Protein Components Of Two Kinds Of Hypotrichous Ciliate

Cytoskeleton is the protein fibre network in eukaryotic cell and becomes a focus of attention in cell biology research. In general the cytoskeleton which contains cytomatrix, membrane skeleton, cytoplasm skeleton and nuclear skeleton, forms network system running through the whole cell. The cytoskeleton plays an important role in keeping the shape of cell and holding the order of cell inner structure, also closely relates with cell movement, material transportation, energy conversion, information transmission, cell division, gene expression, cell differentiation, and other life activities. Currently the cytoskeleton research has rapidly advanced from the morphology observation to the molecular level. The protozoa ciliate which consists of a single cell, exercise all features of life activities. So it is much more complex than the one cell from multi-cell organisms. It has important significance to investigate the cytoskeleton of protozoa, understand the function of the cell and reveal the evolution relationship between different genus by using hypotrichous ciliate as the materials. Different groups of ciliates have different cytoskeleton arrangement and differentiation. The hyportrichous ciliate has complex microtubule structure, including the cilia and the ciliature-skeleton, cortical microtubule skeleton and other cytoplasm microtubule skeleton. These skeleton have an important role in maintenance the cells form and function. Allotricha curdsi and Oxytricha are advanced species in hyportrichous ciliates. In this paper, we use these two kinds of ciliate as materials to detect their cilia-structure differentiation in the term of morphology, morphogenesis and cyst through the method of Flutax direct fluorescent-labeled and anti-αtubulin antibody. We found that the cortical microtubule skeleton is different in the different lifecycle. This revealed the law of its morphogenesis, tubulin differentiation and positioning. And we also use the biochemical extraction and SDS-PAGE to analyze its protein components. The results are as follows:1. Morphology and morphogenesis of Allotricha curdsiThe microtubular organelles in the hyportrichous ciliate Allotricha curdsi were analyzed with fluorescence labeling of FLUTAX and anti-αtubulin antibody. These microtubular organelles consisted of adoral zone of membranelles (AZM), undulating membranes (UM), frontal-ventral-transverse cirri (FVTC), left and right marginal cirri (L- & RMC), dorsal kineties (DK) and the base-associated microtubules of the ciliatures. The base-associated microtubules in the ventral cortex of ciliature comprised membranelle brackets and its base-associated microtubules, UM cytoskeleton, fibrillar cirrral basket (FCB) located at the base of FVTC, anterior longitudinal microtubules (ALM), posterior longitudinal microtubules (PLM) and transverse microtubules (TM) situated at the base of FVTC, LMC, and RMC. Sending from the base of ciliatures and extending to different directions in the cortex, these microtubules formed a complex three-dimensional microtubular net underneath of cortex. In addition, the arrangement of transverse microtubules (TM) at the base of the LMC and RMC displayed a non-mirror symmetrical image, and anterior longitudinal microtubules (ALM) at the base of the TC extend forwards and just did not converge together. During the term of morphogenesis, the oral primordium of the opisthe would originated underneath the cell ellicle, protruding from the cell pellicle to form corresponding ciliature primordium, which then developes into new ciliature through the process of differentiation. The development of ventral ciliature consists the collapse of AZM, UM, FVTC. new ciliature originated in the given cortex of cell groups and moves form certain ciliature primordium, and finally becomes new ciliature.2. The consistence of microtubule cytoskeleton and morphogenesis in Oxytricha sp.Microtubular organelles of protozoa Oxytricha n.sp. was analyzed with anti-α-tubulin antibody and FLUTAX. The results show that the microtubular organelles consisted of adoral zone of membranelles(AZM), undulating membranes(UM), frontal-ventral-transverse cirri(FVTC), left-and-right-marginal cirri(L-RMC), dorsal kinetics(DK) and their base-associated microtubules. The ciliature microtubule and its base-associated microtubule which oriented in such cortical form: AZM-FVTC-LRMC, plays an important role in supporting the movement of ciliary organelles and sustaining the dorsoventral differentiation. During the term of binary fission, two sets of FVTC primordia occurs beside the old and the new AZM, then form certain structures of opisthe and proter. Some old FC and TC finally disappeared after the new structure formed.3. Analysis on the protein components of Allotricha curdsi and OxytrichaThe differences of cytoskeleton protein components between Allotricha curdsi and Oxytricha was investigated using biochemical extraction and electrophoresis. The results are as follows: the cytoskeleton protein components of two kinds of ciliate express between 43kDa to 66kDa. This showed thatα-tubulin is conservative in microtubule cytoskeleton of ciliate. However the insoluble protein has some differences between two kinds of ciliate.