Outer doublet heterogeneity reveals structural polarity related to beat direction in Chlamydomonas flagella.

Abstract

Analysis of serial cross-sections of the Chlamydomonas flagellum reveals several structural asymmetries in the axoneme. One doublet lacks the outer dynein arm, has a beaklike projection in its B-tubule, and bears a two-part bridge that extends from the A-tubule of this doublet to the B-tubule of the adjacent doublet. The two doublets directly opposite the doublet lacking the arm have beak-like projections in their B-tubules. These asymmetries always occur in the same doublets from section to section, indicating that certain doublets have consistent morphological specializations. These unique doublets give the axoneme an inherent structural polarity. All three specializations are present in the proximal portion of the axoneme; based on their frequency in random cross-sections of isolated axonemes, the twopart bridge and the beak-like projections are present in the proximal one quarter and one half of the axoneme, respectively, and the outer arm is absent from the one doublet >90% of the axoneme's length. The outer arm-less doublet of each flagellum faces the other flagellum, indicating that each axoneme has the same rotational orientation relative to the direction of its effective stroke. This strongly suggests that the direction of the effective stroke is controlled by a structural component within the axoneme. The striated fibers are associated with specific triplets in a manner suggesting that they play a role in setting up or maintaining the 180 ° rotational symmetry of the two flagella.In forward swimming Chlamydomonas, the two fagella beat with an effective stroke in approximately the same plane, but in opposite directions (17,32,34). Although the fine structure (6, ll, 14, 30, 32, 39, 40), biochemistry (10, 15, 21, 27, 28, 31, 40), and waveform (3,5,17,32,34) of the Chlamydomonas flagellum have been extensively investigated, little information is available on the mechanism by which the direction of the effective stroke is established. The isolated flagellar apparatuses of Chlamydomonas appear to retain the same beat pattern as in intact cells (16,17), so the direction of beat must be controlled by a structural component within the flagellar apparatus. However, it is not known whether this component is internal or external to the axoneme. Structural differences have been observed in some of the outer doublets of Chlamydomonas (15,38), but it has not been determined whether these asymmetries always occur in the same doublets along the axoneme, nor whether they are oriented in a way that is related to the plane of beat. The central pair of Chlamydomonas rotates during flagellar movement (19,20), so it can not give polarity to the axoneme.To determine whether the axonemes of Chlamydomonas have an intrinsic polarity related to the plane of beat, we made a detailed examination of the fine structure of the flagellar axoneme in serial thin sections having a known orientation relative to the two basal bodies. The results indicate that: (a) there are consistent morphological differences between several ou...

Analysis of microtubule sliding patterns inChlamydomonasflagellar axonemes reveals dynein activity on specific doublet microtubules

“…Structural analyses performed by Hoops and Witman (Hoops and Witman, 1983) revealed several structural asymmetries in Chlamydomonas axonemes. Most relevant to our study, a single doublet microtubule lacks outer dynein arms along the entire length of the axoneme (Hoops and Witman, 1983). This microtubule was designated doublet number one (Db1 , Fig.…”

“…The sliding pattern histograms were constructed by calculating the mean of the percentage contribution of each pattern from all trials. Second, for data regarding the identity of the outer doublet on the dynein exposed edge of the active area, doublet number was determined according to the criteria of Hoops and Witman (Hoops and Witman, 1983). The key identifying structure was the lack of an outer dynein arm on doublet one and the unusual cross-bridge between doublets one and two (Fig.…”

“…Using the same functional approach combined with structural markers that distinguish the outer doublet microtubules (Hoops and Witman, 1983) we have now assessed whether dynein-driven microtubule sliding occurs between specific subsets of doublet microtubules. In addition, we have assessed whether changes in Ca 2+ concentration modulate this activity.…”

Tubal transport of gametes and embryos: a review of physiology and pathophysiology

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Oda5p, a Novel Axonemal Protein Required for Assembly of the Outer Dynein Arm and an Associated Adenylate Kinase

“…Whole cells and isolated axonemes were processed as described previously (Hoops and Witman, 1983). Samples were embedded in a mixture of LX112/ Araldite 502 epoxy resin and sectioned at 50 -70 nm.…”