Immunological dissimilarity in protein component (dynein 1) between outer and inner arms within sea urchin sperm axonemes

Abstract

The 0.5 M KCI-treatment solubilizes the outer arms from sea urchin sperm axonemes. -30% of A-polypeptide, corresponding to dynein 1 in SDS-polyacrylamide gel, was solubilized by this treatment (as SEA-dynein 1) . Electron microscopic observation indicated that the extracted axonemes lacked the outer arms in various degrees. The SEA-dynein 1 was purified and an antiserum against it was prepared in rabbits. The specificity of antiserum to dynein 1 was determined by immunoelectrophoresis and Ouchterlony's double-diffusion test . The anti-dynein 1 serum inhibited ATPase activity of purified SEA-dynein 1 by 95%. By the indirect peroxidase-conjugated antibody method, the loci of SEA-dynein 1 within the intact, salt-extracted and mechanically disrupted axonemes were determined to be the outer arms : deposition of electron-dense materials which represents their localization was detected at the distal ends of the outer arms, in the case of intact axonemes . The 5-6 cross-bridge was hardly decorated . No decoration was seen in the salt-extracted axonemes lacking all the outer arms . In disrupted axonemes, which consist of single to several peripheral doublets, electron-dense materials were deposited only at the outer arms .-73% of axonemal ATPase activity sensitive to antiserum was solubilized by repeated saltextractions . One-half of A-polypeptide (SEA-dynein 1 located at the outer arms) was contained in the pooled extracts . The extracted axonemes contained another half of A-polypeptide (SUAdynein 1 supposed to locate at the inner arms) and retained 31% of axonemal ATPase activity that was almost resistant to antiserum . Solubilized SUA-dynein 1 was immunologically the same as SEA-dynein 1 . This result indicates that in situ SUA-dynein 1 did not receive antidynein 1 antibodies, coinciding with the result obtained for salt-extracted axonemes lacking all the outer arms by the enzyme-antibody method mentioned above. These observations suggest the immunological dissimilarity in dynein 1 between outer and inner arms but do not tell us that the inner arms do not contain dynein 1 .

One of the Nine Doublet Microtubules of Eukaryotic Flagella Exhibits Unique and Partially Conserved Structures

“…Despite exhibiting the same 24-nm periodicities, the observed 3D structure of sea urchin o-SUB5-6 shows no indications that they are modified ODAs ( Figure 2 ; Movie S2 ). This feature is consistent with previous studies demonstrating that the o-SUB5-6 structures can not be decorated with antibodies against ODAs [69] , [70] . Previous studies have also observed that ODAs are almost completely missing from Chlamydomonas DMT1 [15] , [16] , [71] .…”

Association of anti-dynein-1 cross-reactive antigen with the mitotic spindle of mammalian cells.

“…In collaboration with Mohri and Ogawa, we found that the same antiserum also stained the spindle of rat-kangaroo PtK1 cells, as did another rabbit antidynein-1 prepared by Ogawa (18,19,31,32). Preliminary absorption experiments indicated that the fluorescence was due mainly to the presence of dynein-1 or a related antigen(s).…”

Displacement of gold marker in immunoelectron microscopy of human respiratory cilia

“…Other reports have demonstrated that anti-sea urchin sperm flagella dynein 1 antibodies recognize the outer dynein arms of sea urchin sperm flagella by preembedding immunoelectron microscopy (Mohri et al, 1979;Ogawa et al, 1977Ogawa et al, , 1982. Postembedding immunoelectron microscopy of cilia or flagella has not yet been successful.…”