钙依赖蛋白激酶CPK14在花粉管生长中的功能分析

doi:10.13560/j.cnki.biotech.bull.1985.2019-0219

生物技术通报 ›› 2019, Vol. 35 ›› Issue (6): 55-61.doi: 10.13560/j.cnki.biotech.bull.1985.2019-0219

钙依赖蛋白激酶CPK14在花粉管生长中的功能分析

周利明, 卢鑫蕊, 马声葳, 房玮

华北理工大学生命科学学院,唐山 063210

收稿日期:2019-03-12 出版日期:2019-06-26 发布日期:2019-07-08
作者简介:周利明,男,博士,研究方向：植物细胞信号传导研究;E-mail：zhouliming363@163.com
基金资助:
国家自然科学基金项目（31400168）,河北省自然科学基金（C2014209216,C2018209112）,国家重点实验室开放课题（SKLECRA2013OFP15）

Functional Analysis of Calcium-dependent Protein Kinase CPK14 in Pollen Tube Growth

ZHOU Li-ming, LU Xin-rui, MA Sheng-wei, FANG Wei

College of Life Sciences,North China University of Science and Technology,Tangshan 063210

Received:2019-03-12 Published:2019-06-26 Online:2019-07-08

摘要/Abstract

摘要： 旨在揭示钙依赖蛋白激酶CPK14在花粉管极性生长中的生物学功能。利用基因芯片数据绘制花粉高量表达CPK的表达图谱。去除CPK14位于C端的自抑制区及Ca²⁺结合区,保留完整的激酶区及N端可变区,构建组成激活型（CA）的CPK14（CA-CPK14）。利用基因枪技术在烟草花粉中进行瞬时表达,比较分析全长CPK14、CA-CPK14及GFP对照之间的花粉管表型差异。通过CA-CPK14与GFP-RIC4ΔC（活性ROP1的标记物）共同表达试验,分析CA-CPK14对ROP1活性的影响。结果显示,CPK家族中存在8个花粉高量表达成员（其中包括CPK14）,它们在花粉中高量表达而在其他组织中表达较少。CPK14过量表达可以造成花粉管顶端的膨大,而持续激活且不受Ca²⁺调控的CA-CPK14则抑制花粉管萌发及生长。ROP1是花粉管生长的调控关键因子,CA-CPK14的表达可以抑制活性ROP1的标记物（GFP-RIC4ΔC）在花粉管顶端的积累与分布,说明CPK14在ROP1所介导的花粉管生长调控机制中发挥一定的作用。

关键词: 花粉管, 极性生长, 钙依赖蛋白激酶, 钙信号途径

Abstract: The purpose of the study is to reveal the biological function of CPK14（calcium-dependent protein kinase 14）in pollen tube polar growth. Based on the available microarray database,the expression heat-map of pollen-specific CPK was depicted. The constitutively active CPK14（CA-CPK14）was constructed by removing the auto-inhibitor and Ca²⁺ binding domain at the C-terminal and retaining the complete kinase and N-terminal variable regions. The resulting construct（full-length CPK14,CA-CPK14 or GFP alone）was transiently expressed in tobacco pollen tubes via particle bombardment,and the overexpression phenotypes of transformed tubes were compared and analyzed. The effect of CA-CPK14 on the activity of ROP1 was analyzed by co-expression experiment of CA-CPK14 and GFP-RIC4ΔC（a marker of active ROP1）. The results showed that eight pollen-specific CPKs（including CPK14）were identified,and they were highly expressed in mature pollen grains and less expressed in other tissues. CPK14 overexpression caused a swelling tip of pollen tubes,while CA-CPK14（continuous activation and lack of Ca²⁺ regulation）inhibited the germination and growth of pollen tube. ROP1 was a key regulator of pollen tube growth. The expression of CA-CPK14 inhibited the accumulation and distribution of active ROP1 maker（GFP-RIC4ΔC）at the top of pollen tube. These results suggest that CPK14 may play a role in the ROP1-mediated regulation mechanism of pollen tube growth.

Key words: pollen tube, polar growth, calcium-dependent protein kinase, Ca²⁺ signaling pathway

周利明, 卢鑫蕊, 马声葳, 房玮. 钙依赖蛋白激酶CPK14在花粉管生长中的功能分析[J]. 生物技术通报, 2019, 35(6): 55-61.

ZHOU Li-ming, LU Xin-rui, MA Sheng-wei, FANG Wei. Functional Analysis of Calcium-dependent Protein Kinase CPK14 in Pollen Tube Growth[J]. Biotechnology Bulletin, 2019, 35(6): 55-61.

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钙依赖蛋白激酶CPK14在花粉管生长中的功能分析

Functional Analysis of Calcium-dependent Protein Kinase CPK14 in Pollen Tube Growth

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