Monomeric G-actin is uniformly distributed in pollen tubes and is rapidly redistributed via cytoplasmic streaming during pollen tube growth

Dynamic assembly and disassembly of the actin cytoskeleton has been implicated in the regulation of pollen germination and subsequent tube growth. It is widely accepted that actin filaments are arrayed into distinct structures within different regions of the pollen tube. Maintenance of the equilibrium between monomeric globular actin (G-actin) and filamentous actin (F-actin) is crucial for actin assembly and array construction, and the local concentration of G-actin thus directly impacts actin assembly. The localization and dynamics of G-actin in the pollen tube, however, remain to be determined conclusively. To address this question, we created a series of fusion proteins between green fluorescent protein (GFP) and the Arabidopsis reproductive actin ACT11. Expression of a fusion protein with GFP inserted after methionine at position 49 within the DNase I-binding loop of ACT11 (GFP(Met49)-ACT11) rescued the phenotypes in act11 mutants. Consistent with the notion that the majority of actin is in its monomeric form, GFP(Met49)-ACT11 and GFP fusion proteins of four other reproductive actins generated with the same strategy do not obviously label filamentous structures. In further support of the functionality of these fusion proteins, we found that they can be incorporated into filamentous structures in jasplakinolide (Jasp)-treated pollen tubes. Careful observations showed that G-actin is distributed uniformly in the pollen tube and is rapidly redistributed via cytoplasmic streaming during pollen tube growth. Our study suggests that G-actin is readily available in the cytoplasm to support continuous actin polymerization during rapid pollen tube growth. Significance Statement This study has created a functionally appropriate GFP-actin and demonstrates that G-actin is distributed evenly in the pollen tube and is redistributed via cytoplasmic streaming during pollen tube growth.