PT  - JOURNAL ARTICLE
AU  - Brittany Jack
AU  - David M. Mueller
AU  - Ann C. Fee
AU  - Ashley Tetlow
AU  - Prachee Avasthi
TI  - Partially redundant actin genes in <em>Chlamydomonas</em> control flagellum-directed traffic and transition zone organization
AID  - 10.1101/227553
DP  - 2018 Jan 01
TA  - bioRxiv
PG  - 227553
4099  - http://biorxiv.org/content/early/2018/12/03/227553.short
4100  - http://biorxiv.org/content/early/2018/12/03/227553.full
AB  - 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 using Latrunculin B (LatB) treatment on the nap1 mutant background, we find flagellar growth from newly synthesized limiting flagellar proteins is actin-dependent. Upon total actin disruption during flagellar assembly, transmission electron microscopy identified an accumulation of Golgi-adjacent vesicles, suggesting impaired vesicular trafficking may be the mechanism by which actin supports flagellar growth from new flagellar proteins. We also find there is a mislocalization of a key transition zone gating and ciliopathy 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 transport of Golgi-adjacent vesicles and flagellar incorporation of newly synthesized proteins.