Partially redundant actin genes in Chlamydomonas control transition zone and flagellum-directed protein dynamics.

Flagella of the unicellular green alga Chlamydomonas reinhardtii are nearly identical to cilia of mammalian cells and provide an excellent model to study ciliogenesis. These biflagellated cells have two actin genes: one encoding a conventional actin (IDA5) and the other encoding a divergent novel actin-like protein (NAP1). Previously, we described a role for actin in the regulation of flagella-building intraflagellar transport machinery. Here, we probe how actin redundancy contributes to this process using a nap1 mutant Chlamydomonas strain. Disruption of a single actin allows normal or slower incorporation but complete flagellar assembly. However, when we disrupt both actins with Latrunculin B (LatB) treatment on the nap1 mutant background, we find that actins are necessary for flagellar growth from newly synthesized limiting flagellar proteins. In addition, total actin loss also showed reduced protein synthesis during regeneration. We find under actin depleted conditions, there is reduced incorporation of existing flagellar proteins as well as mislocalization of a key transition zone gating protein, NPHP-4. Extended (2 hour) treatment with LatB, a condition under which NAP1 is upregulated, restores NPHP-4 localization. This suggests NAP1 can perform the functions of conventional actin at the transition zone. Our experiments demonstrate that each stage of flagellar biogenesis requires redundant actin function to varying degrees, with an absolute requirement for these actins in incorporation of newly synthesized flagellar proteins.