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Calmodulin Domain Protein Kinase PiCDPK1 Regulates Pollen Tube Growth Polarity through Interaction with RhoGDI.

ABSTRACT: The pollen-specific calcium-dependent protein kinase PiCDPK1 of Petunia inflata has previously been shown to regulate polarity in tip growth in pollen tubes. Here we report the identification of a Rho Guanine Dissociation Inhibitor (PiRhoGDI1) as a PiCDPK1 interacting protein. We demonstrate that PiRhoGDI1 and PiCDPK1 interact in a yeast 2-hybrid assay, as well as in an in vitro pull-down assay, and that PiRhoGDI1 is phosphorylated by PiCDPK1 in vitro. We further demonstrate the PiRhoGDI1 is capable of rescuing the loss of growth polarity phenotype caused by over-expressing PiCDPK1 in vivo using stable transgenic plants. We confirmed that PiRhoGDI1 interacts with a pollen-expressed ROP GTPase isoform consistent with the established role of RhoGDIs in negatively regulating GTPases through their membrane removal and locking them in an inactive cytosolic complex. ROP is a central regulator of polarity in tip growth, upstream of Ca2+, and PiCDPK1 over-expression has been previously reported to lead to dramatic elevation of cytosolic Ca2+ through a positive feedback loop. The discovery that PiCDPK1 impacts ROP regulation via PiRhoGDI1 suggests that PiCDPK1 acts as RhoGDI displacement factor and leads us to propose a model which we hypothesize regulates the rapid recycling of ROP GTPase at the pollen tube tip.

SUBMITTER: Scheible N

PROVIDER: S-EPMC8838988 | biostudies-literature | 2022 Jan

REPOSITORIES: biostudies-literature

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Calmodulin Domain Protein Kinase PiCDPK1 Regulates Pollen Tube Growth Polarity through Interaction with RhoGDI.

Scheible Nolan N Yoon Gyeong Mee GM McCubbin Andrew G AG

Plants (Basel, Switzerland) 20220119 3

The pollen-specific calcium-dependent protein kinase PiCDPK1 of <i>Petunia inflata</i> has previously been shown to regulate polarity in tip growth in pollen tubes. Here we report the identification of a Rho Guanine Dissociation Inhibitor (PiRhoGDI1) as a PiCDPK1 interacting protein. We demonstrate that PiRhoGDI1 and PiCDPK1 interact in a yeast 2-hybrid assay, as well as in an in vitro pull-down assay, and that PiRhoGDI1 is phosphorylated by PiCDPK1 in vitro. We further demonstrate the PiRhoGDI1 ...[more]

PMID: 35161234

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Project description:Background and aimsAn unusual form of pollen tube growth was observed for several Conospermum species (family Proteaceae). The rate of pollen tube growth, the number of tubes to emerge and the ultrastructure of these tubes are given here.MethodsPollen was germinated in vitro in different sucrose concentrations and in the presence of calcium channel blockers, and tube emergence and growth were recorded on a VCR. Measurements were taken of the number of tubes to emerge and rate of tube emergence. Pollen behaviour in vivo was also observed. The ultrastructure of germinated and ungerminated pollen was observed using TEM.ResultsAfter 10 s to 3 min in germination medium, up to three pollen tubes emerged and grew at rates of up to 55 micro m s(-1); the rate then slowed to around 2 micro m s(-1), 30 s after the initial growth spurt. Tubes were observed to grow in pulses, and the pulsed growth continued in the presence of calcium channel blockers. Optimal sugar concentration for pollen germination was 300 g L(-1), in which up to 81 % of pollen grains showed fast germination. Germination and emergence of multiple tubes were observed in sucrose concentrations of 100-800 g L(-1). The vegetative and generative nuclei moved into one of the tubes. Multiple tubes from a single grain were observed on the stigma. Under light microscopy, the cytoplasm in the tube showed a clear region at the tip. The ultrastructure of C. amoenum pollen showed a bilayered exine, with the intine being very thick at the pores, and elsewhere having large intrusions into the plasma membrane. The cytoplasm was dense with vesicles packed with inner tube cell wall material. Golgi apparatus producing secretory vesicles, and mitochondria were found throughout the tube. The tube wall was bilayered; both layers being fibrous and loosely packed.ConclusionsIt is proposed that, for Conospermum, initial pollen tube wall constituents are manufactured and stored prior to pollen germination, and that tube extension occurs as described in the literature for other species, but at an exceptionally fast rate.

Dataset Information

Calmodulin Domain Protein Kinase PiCDPK1 Regulates Pollen Tube Growth Polarity through Interaction with RhoGDI.

Publications

Calmodulin Domain Protein Kinase PiCDPK1 Regulates Pollen Tube Growth Polarity through Interaction with RhoGDI.

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