Submitted to: Proceedings of the National Academy of Sciences
Publication Type: Peer reviewed journal
Publication Acceptance Date: 9/28/2007
Publication Date: 11/20/2007
Publication URL: www.pnas.org/content/104/47/18830.full.pdf+html?sid=40265c3e-6ff7-4381-ba0d-a2791bc98377
Citation: Zhang, Y., McCormick, S.M. 2007. A distinct mechanism regulating a pollen-specific guanine nucleotide exchange factor for the small GTPase Rop in Arabidopsis thaliana. Proceedings of the National Academy of Sciences. 104(47):18830-18835. Interpretive Summary: Pollen tube growth is partly regulated by receptor kinases. One protein that links receptor kinases to the small GTPase called Rop is a protein called Rop-GEF, for guanine exchange factor. We studied how Rop-GEF is regulated during pollen tube growth, by making gene constructs fused to a green fluorescent protein and introducing them into tobacco pollen.
Technical Abstract: Rop/Rac small GTPases are central to diverse developmental and cellular activities in plants, playing an especially important role in polar growth of pollen tubes. Although it is established that a class of plant-specific RopGEFs promotes the activity of Rop/Rac through the catalytic PRONE (Plant specific Rop nucleotide exchanger) domain, not much is known about how RopGEF function is controlled to allow a spatiotemporally regulated Rop activity. To understand such a process in pollen, we did functional analysis with a pollen-specific RopGEF, AtRopGEF12. Overexpression of AtRopGEF12 had minimal phenotypic effects, whereas overexpression of a C-terminally truncated version disturbed tube growth, suggesting that the C-terminus was inhibitory to GEF function. In contrast to non-pollen-expressed RopGEFs, pollen-expressed RopGEFs have conserved C-termini. A phospho-mimicking mutation at an invariant serine within the C-terminus of AtRopGEF12 resulted in loss of the Cterminal inhibition, suggesting that phosphorylation regulates GEF activity in vivo. The PRONE domain of AtRopGEF12 (PRONE12) was not sufficient to induce isotropic tube growth whereas co-overexpression of AtRopGEF12 with AtPRK2a, an Arabidopsis homolog of the tomato pollen receptor kinase LePRK2, caused isotropic growth reminiscent of that seen upon over-expression of a constitutively active (CA) Rop. Co-overexpression of AtPRK2a with an N-terminally truncated AtRopGEF12 did not induce isotropic growth, indicating a positive role for the Nterminal domain. Our results suggest a mechanism by which the non-catalytic domains of pollen-specific/enriched RopGEFs regulate PRONE function, leading to polarized pollen tube growth.