A Calcium-Dependent Protein Kinase Interacts with and Activates A Calcium Channel to Regulate Pollen Tube Growth Liming Zhou, Wenzhi Lan, Yuanqing Jiang,

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Presentation transcript:

A Calcium-Dependent Protein Kinase Interacts with and Activates A Calcium Channel to Regulate Pollen Tube Growth Liming Zhou, Wenzhi Lan, Yuanqing Jiang, Wei Fang, Sheng Luan Molecular Plant Volume 7, Issue 2, Pages 369-376 (February 2014) DOI: 10.1093/mp/sst125 Copyright © 2014 The Authors. All rights reserved. Terms and Conditions

Figure 1 CPK32 Overexpression Induces Excessive Ca2+ Accumulation in the Tip of Pollen Tubes. Ratiometric Ca2+ imaging of pollen tubes was performed using the calcium-sensitive dye Indo-1-dextran. Calcium concentrations have been pseudo-color-coded according to the inset scale. Molecular Plant 2014 7, 369-376DOI: (10.1093/mp/sst125) Copyright © 2014 The Authors. All rights reserved. Terms and Conditions

Figure 2 CPK32 Interacts with CNGC18. (A, B) Yeast two-hybrid assay of CACPK32 (or CACPK34) with C-terminal soluble domain of CNGC members. The pGADT7 and pGBKT7 vectors were used as negative control. (C) Overexpression phenotype and subcellular localization for CNGC18. (D) Overexpression of CNGC18 disturbs tip-focused Ca2+ gradient visualized by ratiometric Ca2+ imaging. (E, F) Quantitative analysis of pollen tube phenotypes induced by co-expression of LAT52::CNGC18 with LAT52::GFP. Asterisks represent a significant difference from LAT52::GFP control (p < 0.05; t-test); error bars indicate SD. Data were collected from 50–60 tubes per experiment. Molecular Plant 2014 7, 369-376DOI: (10.1093/mp/sst125) Copyright © 2014 The Authors. All rights reserved. Terms and Conditions

Figure 3 FRET Analysis of CACPK32 Interaction with CNGC18. Tobacco pollen tubes co-expressing CNGC18–YFP and CACPK32–CFP or CACPK34–CFP were examined with FRET procedure as described in the ‘Methods’ section. The intensity of FRET signals is displayed in pseudo-color mode (red, highest signal). Molecular Plant 2014 7, 369-376DOI: (10.1093/mp/sst125) Copyright © 2014 The Authors. All rights reserved. Terms and Conditions

Figure 4 Phenotypes Analysis of Pollen Tubes Co-Expressing CNGC18 with CPK32–GFP. (A) Representative confocal images of pollen tubes transiently expressing CPK32–GFP, CNGC18–GFP, or co-expressing CNGC18 with CPK32–GFP. (B) Quantitative analysis of the co-expression phenotype. Data were collected from 50–60 tubes per experiment. Error bars indicate SD. Molecular Plant 2014 7, 369-376DOI: (10.1093/mp/sst125) Copyright © 2014 The Authors. All rights reserved. Terms and Conditions

Figure 5 CPK32 Activates CNGC18 in the Xenopus Oocytes. (A) Typical current traces recorded from the oocytes injected with water (Control), CACPK32, CNGC18, or CNGC18 plus CACPK32 (CNGC18+CACPK32). The oocytes were perfused with bath solution containing 10mM CaCl2, 0.1mM dibutyryl-cAMP, 185mM mannitol, and 10mM Mes-Tris (pH 7.4). Dotted lines represent the zero current level. (B) Effects of CACPK32 and CACPK34 on the currents generated by CNGC18 at –150 mV. The pooled current values were at 1.8 s of each voltage-clamp episode and presented as means ± SE, with n = 5 in each case. Molecular Plant 2014 7, 369-376DOI: (10.1093/mp/sst125) Copyright © 2014 The Authors. All rights reserved. Terms and Conditions