No abstract

Actin cables in pollen tubes serve as molecular tracks for cytoplasmic streaming and organelle movement and are formed by actin bundling factors like villins and fimbrins. However, the precise mechanisms by which actin cables are generated and maintained remain largely unknown. Fimbrins comprise a family of five members in Arabidopsis thaliana. Here, we characterized a fimbrin isoform, Arabidopsis FIMBRIN5 (FIM5). Our results show that FIM5 is required for the organization of actin cytoskeleton in pollen grains and pollen tubes, and FIM5 loss-of-function associates with a delay of pollen germination and inhibition of pollen tube growth. FIM5 decorates actin filaments throughout pollen grains and tubes. Actin filaments become redistributed in fim5 pollen grains and disorganized in fim5 pollen tubes. Specifically, actin cables protrude into the extreme tips, and their longitudinal arrangement is disrupted in the shank of fim5 pollen tubes. Consequently, the pattern and velocity of cytoplasmic streaming were altered in fim5 pollen tubes. Additionally, loss of FIM5 function rendered pollen germination and tube growth hypersensitive to the actin-depolymerizing drug latrunculin B. In vitro biochemical analyses indicated that FIM5 exhibits actin bundling activity and stabilizes actin filaments. Thus, we propose that FIM5 regulates actin dynamics and organization during pollen germination and tube growth via stabilizing actin filaments and organizing them into higher-order structures.

Protein concentration gradients organize cells and tissues and commonly form through diffusion away from a local source of protein. Interestingly, during the asymmetric division of the zygote, the RNA-binding proteins MEX-5 and PIE-1 form opposing concentration gradients in the absence of a local source. In this study, we use near-total internal reflection fluorescence (TIRF) imaging and single-particle tracking to characterize the reaction/diffusion dynamics that maintain the MEX-5 and PIE-1 gradients. Our findings suggest that both proteins interconvert between fast-diffusing and slow-diffusing states on timescales that are much shorter (seconds) than the timescale of gradient formation (minutes). The kinetics of diffusion-state switching are strongly polarized along the anterior/posterior (A/P) axis by the PAR polarity system such that fast-diffusing MEX-5 and PIE-1 particles are approximately symmetrically distributed, whereas slow-diffusing particles are highly enriched in the anterior and posterior cytoplasm, respectively. Using mathematical modeling, we show that local differences in the kinetics of diffusion-state switching can rapidly generate stable concentration gradients over a broad range of spatial and temporal scales.