Quantitative analysis and modeling of katanin function in flagellar length control.
Bottom Line: Previous work demonstrated that Chlamydomonas cytoplasm contains a pool of flagellar precursor proteins sufficient to assemble a half-length flagellum and that assembly of full-length flagella requires synthesis of additional precursors to augment the preexisting pool.We used quantitative analysis of length distributions to identify candidate genes controlling pool regeneration and found that a mutation in the p80 regulatory subunit of katanin, encoded by the PF15 gene in Chlamydomonas, alters flagellar length by changing the kinetics of precursor pool utilization.We tested this model using a stochastic simulation that confirms that cytoplasmic microtubules can compete with flagella for a limited tubulin pool, showing that alteration of cytoplasmic microtubule severing could be sufficient to explain the effect of the pf15 mutations on flagellar length.
Affiliation: Department of Biochemistry and Biophysics, University of California, San Francisco, San Francisco, CA 94158.Show MeSH
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Mentions: With the combined model of cytoplasmic microtubule dynamics coupled with flagellar length dynamics in hand, we can simulate how both microtubule populations fluctuate in length over time in simulated regeneration experiments in which the flagellum is initialized at zero length and allowed to regenerate to a new steady-state length (Figure 4C). When many such simulations are combined, we find that decreasing the katanin-mediated severing of cytoplasmic microtubules produces a short steady-state flagellar length (Figure 5). With the particular parameters of this simulation, ∼10-fold decrease in katanin activity would produce ∼2-fold decrease in average flagellar length, similar to that seen in pf15 mutants. Obviously, given the highly simplified nature of our model, it is not possible to directly infer the level of katanin activity change that would be necessary in a real cell, but the model does support the possibility that alteration in severing activity can in principle be sufficient to produce a change in flagellar length.
Affiliation: Department of Biochemistry and Biophysics, University of California, San Francisco, San Francisco, CA 94158.