1. Pollen Tube -- the model system for the study of cell polarity control and tip growth Speaker: Chunhui Cai

2. Introduction Following germination on the surface of the stigma, pollen tubes are directed through the stigma surface, grow within the transmitting track, emerge from the transmitting tract and finally are targeted toward the micropyle to deliver sperms to the ovule.

3. Introduction Pollen tubes provide a simple model system for studying cell growth and cell polarity control. Pollen tubes extend exclusively at the cell apex via an extreme form of polar growth, known as tip growth, producing uniformly shaped cylindrical cells.

4. Introduction Pollen tube growth requires spatiotemporal regulation of many cellular functions, -- ion fluxes, organization and dynamics of cytoskeletal elements, vesicular trafficking, exocytosis, endocytosis, and wall synthesis, assembly and remodelling

5. Introduction An important issue: tip growth domain (PM domain) which leads to localized extension of the plasma membrane and cell walls. Both internal signals and external guidance signals may regulate the tip growth.

6. Introduction Questions of interest: –How the tip growth domain is established in pollen tubes; –How internal tip growth signals and external growth guidance signals spatially regulate the tip growth domain; –How the tip growth domain signals to the machinery for exocytosis and vesicle targeting

8. Rop GTPase Rop is a plant-specific subfamily of the Rho family of G protein that include Cdc42, Rac and Rho subfamilies from animals and fungi. (Rho GTPase signaling has been shown to regulate a wide variety of cellular processes, including reorganization and dynamics of F-actin, microtubule organization, gene transcription, RNA processing, cell cycle progression, and activation of specific enzymes such as NADPH oxidase and glucan synthase)

9. A global study of Rop signaling network in Arabidopsis

10. Arabidopsis: 11 ROPs, divided into four groups based on their amino acid sequence similarities. Members within the same groups appear to be functionally redundant; different groups are distinct. ROP1 regulates pollen germination and pollen tube growth. The existence of a tip-localized ROP1- dependent signaling pathway activates the pollen germination and pollen tube growth and the active ROP1 signaling complex defines a PM domain for growth.

11. ROP1 localization and/or its localized activation indeed define the tip growth domain The active ROP1 promoted ROP1 recruitment to the apical PM region, forming a ROP activation/recruitment positive feedback loop, which provides a mechanism for the rapid reproduction of tip growth site. The positive feedback loop is activated locally, amplified laterally and inhibited globally.

12. Rop1 signaling network Rop1 activates two downstream pathways through direct targets: RIC3 and RIC4. –RIC4 promotes F-actin assembly, whereas RIC3 activate Ca2+ signaling that leads to F- actin disassembly. This Rop-dependent dynamics of tip F-actin is important for polar growth.

13. RIC3 and RIC4 RIC3 and RIC4 are two distinct ROP1 target proteins RIC4 promotes the assembly of apical F-actin RIC3 regulates and promotes the tip-localized influx of intracellular Ca2+, which subsequently modulates for the formation of the tip-focused Ca2+ gradient. RIC3 and RIC4 counteract to control actin dynamics and apical growth A balance between RIC3 and RIC4 is critical for efficient tip growth RIC3 acts through Ca2+ to promote the disassembly of RIC4-dependent F-actin RIC4 acts through F-actin to counteract the RIC3 pathway.

15. The mechanism by which RIC4 promotes actin assembly and the structural nature of RIC4-mediated F-actin remain to be investigated. RIC4-actin pathway counteract the RIC- Ca2+ pathway; RIC4-mediated actin assembly appears to inhibit the accumulation of Ca2+ at the tip by inhibiting a Ca2+ permeable PM channel.

16. Phases

17. RIC3 in the cytosol and RIC4 in the PM. Because RIC3 and RIC4 both contain a CRIB motif and are expected to bind the same effector domain on ROP1, it is important to have distinct kinetics during their interaction with ROP1 so that they would not compete with each other in binding to ROP1, but would co-ordinate with each other to transmit ROP1 signals. These two ROP signaling targets coordinately oscillate with the same period but in the different phases during the growth of pollen tubes