Immunocytochemical studies with antibodies to three proteins prominent in the isolated microtubule cytoskeleton of a trypanosomatid.
The cytoskeleton of Crithidia fasciculata consists of a corset of parallel microtubules enclosing the cell body and closely underlying the plasma membrane. Distinct sets of crosslinks appear to connect tubules to each other and to membrane. Our objective is to determine the composition of these crosslinks and to elucidate the basis of this spectacular example of membrane-microtubule interaction. We purified three proteins (designated COP-33, -41, and -61 by their subunit Mr), which were consistently abundant in highly purified cytoskeletons. All three bound strongly to microtubules in vitro, and the first two induced bundles through periodic crosslinking. Polyclonal antibodies against each have been used to try to localize these proteins in thin sections of cells or whole mounts of cytoskeletons. Antibodies to COP-41 bound specifically to glycosomes, organelles that encapsulate many glycolytic enzymes in these protozoa, and COP-41 has been identified as glyceraldehyde 3-P dehydrogenase.
Laboratory of Cell Biology, National Heart, Lung, and Blood Institute, Bethesda, Maryland.
G T
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Bramblett
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Kambadur
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Flavin
D000818:Animals; D000913:Antibodies, Protozoan / Q000276:immunology*; D005591:Chemical Fractionation; D003421:Crithidia / Q000032:analysis* / Q000648:ultrastructure; D003432:Cross-Linking Reagents; D003599:Cytoskeleton / Q000276:immunology*; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases / Q000032:analysis / Q000378:metabolism; D007150:Immunohistochemistry; D008868:Microtubule Proteins / Q000032:analysis* / Q000276:immunology; D008869:Microtubule-Associated Proteins / Q000276:immunology / Q000302:isolation & purification; D008870:Microtubules / Q000032:analysis*; D014404:Tubulin / Q000302:isolation & purification / Q000378:metabolism
D000913:Antibodies, Protozoan; D003432:Cross-Linking Reagents; D008868:Microtubule Proteins; D008869:Microtubule-Associated Proteins; D014404:Tubulin; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases