A crustacean neuronal cytoskeletal protein with characteristics of neurofilaments and microtubule-associated proteins.
The purpose of this study was to characterize further a unique protein that is a component of the cytoskeleton of crayfish neurons. This protein, referred to as P600, is unique because it is unusually large (Mr greater than 600 kD), and because it has characteristics in common with both mammalian microtubule-associated proteins and neurofilaments. Immunohistochemical techniques have shown that P600 colocalizes with microtubules and is a component of the fibrous side-arms that extend from microtubules (Weaver and Viancour, Brain Res. 544:49, 1991). We have developed a method for obtaining purified P600 by using gel filtration techniques. When viewed by negative staining electron microscopy, P600 obtained by that method produced 11 nm-wide beaded filaments. The number of filaments was strictly related to the P600 concentration in a column fraction. A small amount of P600 consistently copurified with taxol-stabilized microtubules. The proportion copurifying with microtubules was increased by using apyrase to deplete ATP, or by using a nonhydrolyzable ATP analogue to compete with ATP. Immunogold labeling localized P600 near the ends of a subset of the fibrous side-arms extending from endogenous axonal microtubules. Several polyclonal antibodies against mammalian microtubule-associated proteins were tested for P600 labeling on immunoblots, and positive labeling was obtained with an antiserum directed against a region of microtubule-associated protein 1B that has microtubule binding activity. Epitope homology between P600, mammalian microtubule-associated protein 1B, and the mammalian mid-molecular weight neurofilament subunit is discussed in the context of possible evolutionary relationships among these cytoskeletal proteins.
Weaver
D J
DJ
University of Maryland, Baltimore 21228.
Viancour
T A
TA
D000818:Animals; D003400:Astacoidea / Q000378:metabolism*; D002850:Chromatography, Gel; D003599:Cytoskeleton / Q000378:metabolism* / Q000648:ultrastructure; D004591:Electrophoresis, Polyacrylamide Gel; D010450:Endopeptidases; D006023:Glycoproteins / Q000032:analysis; D006868:Hydrolysis; D015151:Immunoblotting; D007150:Immunohistochemistry; D008854:Microscopy, Electron; D008869:Microtubule-Associated Proteins / Q000737:chemistry / Q000378:metabolism*; D009419:Nerve Tissue Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D016900:Neurofilament Proteins / Q000737:chemistry / Q000378:metabolism*; D009474:Neurons / Q000378:metabolism* / Q000648:ultrastructure; D010766:Phosphorylation; D051381:Rats; D011916:Rats, Inbred F344; D017207:Rats, Sprague-Dawley
D006023:Glycoproteins; D008869:Microtubule-Associated Proteins; D009419:Nerve Tissue Proteins; D016900:Neurofilament Proteins; D010450:Endopeptidases