TY - JOUR
T1 - Two proteins isolated from sea urchin sperm flagella
T2 - Structural components common to the stable microtubules of axonemes and centrioles
AU - Hinchcliffe, Edward H.
AU - Linck, Richard W.
PY - 1998
Y1 - 1998
N2 - Biochemical fractionation of axonemal microtubules yields the protofilament ribbon (pf-ribbon), an insoluble structure of 3-4 longitudinal protofilaments composed primarily of α/β tubulin, tektins A, B and C, and two previously uncharacterized polypeptides of 77 kDa and 83 kDa. We have isolated the 77/83 kDa polypeptides (termed Sp77 and Sp83) from sperm flagella of the sea urchin Stronglyocentrotus pupuratus and raised polyclonal antibodies against them. Sp77 and Sp83 copurify exclusively with the pf-ribbon. Both the anti-Sp77 and anti-Sp83 antibodies detected the nine outer doublets and the basal bodies of sea urchin sperm by immunofluorescence microscopy. In addition, the anti-Sp83 antibody, but not the anti-Sp77 antibody, detected a single 83 kDa polypeptide on immunoblots of unfertilized sea urchin egg cytoplasm, and a single polypeptide of 80 kDa on blots of isolated mitotic spindles from Chinese hamster ovary (CHO) cells. Previous studies have shown that tektins are present in the basal bodies and centrosomes/centrioles of cells ranging from clam to human. We found that anti-Sp83 decorates the spindle poles in sea urchin zygotes, and the interphase centrosome and spindle poles in CHO cells. In CHO cells arrested in S phase with aphidicolin, anti-Sp83 detects multiple centrosomes. The staining of the centrosome was not disrupted by prolonged nocodazole treatment, suggesting that the 80 kDa polypeptide is associated with the centrioles themselves. Our observations demonstrate that, like tektins, Sp77 and Sp83 are structural proteins associated with stable doublet microtubules, and may be components of basal bodies and centrioles of sea urchins and mammalian cells.
AB - Biochemical fractionation of axonemal microtubules yields the protofilament ribbon (pf-ribbon), an insoluble structure of 3-4 longitudinal protofilaments composed primarily of α/β tubulin, tektins A, B and C, and two previously uncharacterized polypeptides of 77 kDa and 83 kDa. We have isolated the 77/83 kDa polypeptides (termed Sp77 and Sp83) from sperm flagella of the sea urchin Stronglyocentrotus pupuratus and raised polyclonal antibodies against them. Sp77 and Sp83 copurify exclusively with the pf-ribbon. Both the anti-Sp77 and anti-Sp83 antibodies detected the nine outer doublets and the basal bodies of sea urchin sperm by immunofluorescence microscopy. In addition, the anti-Sp83 antibody, but not the anti-Sp77 antibody, detected a single 83 kDa polypeptide on immunoblots of unfertilized sea urchin egg cytoplasm, and a single polypeptide of 80 kDa on blots of isolated mitotic spindles from Chinese hamster ovary (CHO) cells. Previous studies have shown that tektins are present in the basal bodies and centrosomes/centrioles of cells ranging from clam to human. We found that anti-Sp83 decorates the spindle poles in sea urchin zygotes, and the interphase centrosome and spindle poles in CHO cells. In CHO cells arrested in S phase with aphidicolin, anti-Sp83 detects multiple centrosomes. The staining of the centrosome was not disrupted by prolonged nocodazole treatment, suggesting that the 80 kDa polypeptide is associated with the centrioles themselves. Our observations demonstrate that, like tektins, Sp77 and Sp83 are structural proteins associated with stable doublet microtubules, and may be components of basal bodies and centrioles of sea urchins and mammalian cells.
KW - Basal body
KW - Centrosome
KW - Microtubule associated protein
KW - Spindle pole
KW - Tektin
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U2 - 10.1242/jcs.111.5.585
DO - 10.1242/jcs.111.5.585
M3 - Article
C2 - 9454732
AN - SCOPUS:0031913583
SN - 0021-9533
VL - 111
SP - 585
EP - 595
JO - Journal of cell science
JF - Journal of cell science
IS - 5
ER -