A calmodulin-binding protein from Arabidopsis has an essential role in pollen germination
A calmodulin-binding protein from Arabidopsis has an essential role in pollen germination
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DOI：
10.1073/pnas.1734110100
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发表时间：
2003-09-02
影响因子：
11.1
通讯作者：
Reddy, ASN
中科院分区：
文献类型：
--
作者：
Golovkin, M;Reddy, ASN
钙调素（CaM）是一种普遍存在于真核生物中的多功能钙感受器，通过调节许多无关蛋白的活性/功能来介导钙离子的作用。虽然钙和CaM在花粉萌发和花粉管生长中起着至关重要的作用，但介导其作用的蛋白质尚未被鉴定。我们分离了三个密切相关的钙调素结合蛋白（NPG 1，（.）NPGR 1和NPGR 2）。NPG 1（无花粉萌发1）仅在花粉中表达，而NPG相关蛋白（NPGR 1和NPGR 2）在花粉和其他组织中表达。细菌表达的NPG 1绑定三个异构体的拟南芥钙调素在钙依赖性的方式。为了分析NPG 1的功能，我们进行了反向遗传学筛选，并分离出一个突变体，其中NPG 1被T-DNA插入破坏。NPG 1基因敲除突变体的分离和分子分析以及与雄性不育突变体的杂交表明，突变的NPG 1不通过雄配子体传递。敲除突变体中NPG 1的表达补充了突变体表型。通过光学显微镜分析敲除突变体的花粉发育显示正常的花粉发育。四倍体背景下的NPG 1突变体花粉证实了NPG 1是花粉发育的关键基因。然而，在四重背景中用来自突变体的花粉进行的萌发研究表明，携带突变体等位基因的花粉不萌发。我们的遗传学，组织学和花粉萌发的研究与敲除突变株系表明，NPG 1是不必要的小孢子和配子发生，但花粉萌发是必不可少的。
Calmodulin (CaM), a ubiquitous multifunctional calcium sensor in all eukaryotes, mediates calcium action by regulating the activity/function of many unrelated proteins. Although calcium and CaM are known to play a crucial role in pollen germination and pollen tube growth, the proteins that mediate their action have not been identified. We isolated three closely related CaM-binding proteins (NPG1,(.)NPGR1, and NPGR2) from Arabidopsis. NPG1 (No Pollen Germination 1) is expressed only in pollen, whereas the NPG-related proteins (NPGR1 and NPGR2) are expressed in pollen and other tissues. The bacterially expressed NPG1 bound three isoforms of Arabidopsis CaM in a calcium-dependent manner. To analyze the function of NPG1, we performed a reverse genetics screen and isolated a mutant in which NPG1 is disrupted by a T-DNA insertion. Segregation and molecular analyses of the NPG1 knockout mutant and a cross with a male sterile mutant indicate that the mutated NPG1 is not transmitted through the male gametophyte. Expression of NPG1 in the knockout mutant complemented the mutant phenotype. Analysis of pollen development in the knockout mutant by light microscopy showed normal pollen development. Pollen from NPG1 mutant in the quartet background has confirmed that NPG1 is dispensable for pollen development. However, germination studies with pollen from the mutant in the quartet background indicate that pollen carrying a mutant allele does not germinate. Our genetic, histological, and pollen germination studies with the knockout mutant line indicate that NPG1 is not necessary for microsporogenesis and gametogenesis but is essential for pollen germination.