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First published online 5 May 2004
doi: 10.1242/jcs.01082

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Analysis of microtubule sliding patterns in Chlamydomonas flagellar axonemes reveals dynein activity on specific doublet microtubules

Department of Biological Sciences, Dartmouth College, Hanover, NH 03755, USA

View larger version (48K): [in a new window]   Fig. 1. (A) Electron micrograph and accompanying diagram of axonemal transverse section following the induction of microtubule sliding. The axoneme is oriented as viewed, proximal to distal and based on the doublet (Db) numbering system of Hoops and Witman (Hoops and Witman, 1983). Db3 has slid away from the axoneme leaving doublet 2 on the dynein exposed edge of the active area (arrow). (B) Diagram of the six possible sliding patterns observed following ATP-induced microtubule sliding. The associated central apparatuses are not meant to imply that only transverse sections with a specific central apparatus orientation are included in the analysis. The numbers indicate the number of doublet microtubules that remain associated with the central apparatus. (C) Flagella as viewed looking towards the cell body. The arrows indicate the direction of the beat plane during the effective stroke.

View larger version (42K): [in a new window]   Fig. 2. (A) Distributions of the microtubule sliding patterns (Pattern) and the doublet present at the dynein-exposed edge of the active area (Doublet Number) following the induction of microtubule sliding in low Ca2+ buffer. For sliding pattern data, A54-e18: 3 trials, n=110. 137c: 3 trials, n=145. For doublet number data: A54-e18: 3 trials, n=63. 137c: 3 trials, n=33. (B) Distributions of the microtubule sliding patterns (Pattern) and the doublet present at the dynein-exposed edge of the active area (Doublet Number) following the induction of microtubule sliding in high Ca2+ buffer. For sliding pattern data: A54-e18: 3 trials, n=171; 137c: 3 trials, n=90. For doublet number data: A54-e18: 3 trials, n=81; 137c: 3 trials, n=27.

View larger version (41K): [in a new window]   Fig. 3. Distributions of the microtubule sliding patterns (Pattern) and the doublet present at the dynein-exposed edge of the active area (Doublet Number) following the induction of microtubule sliding in low (dark bars) and high (light bars) Ca2+ buffer. For microtubule sliding pattern data: cpc1 low Ca2+: 3 trials, n=143; high Ca2+: 3 trials, n=84; pf6 low Ca2+: 3 trials, n=125; high Ca2+: 4 trials, n=90; pf14 low Ca2+: 4 trials, n=105; high Ca2+: 4 trials, n=95; pf17 low Ca2+: 3 trials, n=64; high Ca2+: 3 trials, n=50. For doublet number data: cpc1 low Ca2+: 3 trials, n=46; high Ca2+: 3 trials, n=42; pf6 low Ca2+: 3 trials, n=65; high Ca2+: 3 trials, n=19; pf14 low Ca2+: 4 trials, n=52; high Ca2+: 4 trials, n=32; pf17 low Ca2+: 3 trials, n=20; high Ca2+: 3 trials, n=13.

View larger version (28K): [in a new window]   Fig. 4. Distributions of the microtubule sliding patterns (Pattern) and the doublet present at the dynein-exposed edge of the active area (Doublet Number) following the induction of microtubule sliding in low (dark bars) and high (light bars) Ca2+ buffer. For microtubule sliding pattern data: ida1 low Ca2+: 3 trials, n=148; high Ca2+: 3 trials, n=128; pf3 low Ca2+: 2 trials, n=55; high Ca2+: 2 trials, n=56; oda1 low Ca2+: 3 trials, n=164; high Ca2+: 3 trials, n=58; pf28 low Ca2+: 3 trials, n=113; high Ca2+: 4 trials, n=97; pf30pf28 low Ca2+: 4 trials, n=107; high Ca2+: 4 trials, n=67. For doublet number data: ida1 low Ca2+: 3 trials, n=115; high Ca2+: 3 trials, n=70; pf3 low Ca2+: 2 trials, n=26; high Ca2+: 2 trials, n=13.

View larger version (23K): [in a new window]   Fig. 5. Principal components analysis for microtubule sliding pattern data. The first principal component (PC1) explained 70.9% of the variation among microtubule sliding patterns for axonemes isolated from wild-type and mutant strains in both low and high Ca2+ buffer. Larger PC1 values have higher frequencies of sliding patterns P7 and P8 and lower frequencies of patterns P3-P6. The second principal component, PC2, explained 13.9% of the variation; larger values of PC2 have higher frequencies of sliding pattern P6 and lower frequencies of sliding pattern P4.

View larger version (23K): [in a new window]   Fig. 6. Principal components analysis for data derived from the analysis of doublet number present on the dynein exposed edge of the active area. The first principal component (PC1) explained 69.5% of the variation; larger values for PC1 had higher frequencies of Db2 and lower frequencies of Db3 on the dynein exposed edge of the active area. The second principal component (PC2) explained 18.6% of the variation, with larger values for PC2 having larger frequencies of Db1 and lower frequencies of Db3 on the dynein exposed edge.