Kato Yoshinobu, Tadokoro Shun, Ishida Shota, Niidome Maki, Kimura Yuka, Takayama Seiji, Fujii Sota
Graduate School of Agricultural and Life Sciences, The University of Tokyo, 1-1-1 Yayoi, Bukyo-ku, Tokyo 113-8657, Japan.
Japan Science and Technology Agency, Precursory Research for Embryonic Science and Technology, 4-1-8 Honcho, Kawaguchi, Saitama 332-0012, Japan.
Plant Cell Physiol. 2025 Jul 24;66(6):926-939. doi: 10.1093/pcp/pcaf039.
In plants, there is an active prezygotic interspecific-incompatibility mechanism to prevent unfavorable hybrids between two species. We previously reported that an uncharacterized protein with four-transmembrane domains, named as Stigmatic Privacy 1 (SPRI1), is responsible for rejecting heterospecific pollen grains in Arabidopsis thaliana. However, the lack of notable functional domains in SPRI1 has limited our understanding of its biochemical properties. In this study, we conducted a functional analysis of the SPRI1 protein through point-mutational experiments and biochemical analysis. We explored the molecular regulatory mechanisms of SPRI1 and the relationships with its function. Alanine- and glycine-scanning experiments together with the evolutional analysis showed that the structural integrity of the C-terminal regions of the extracellular domain of this protein is important for its function. In addition, we found two cysteines (C67 and C80) within the extracellular domain that may be involved in the formation of intermolecular disulfide bonds. These cysteine residues are required for the stabilization of the SPR1 protein. Furthermore, SPRI1 may form homo-multimers and is present as part of a ∼300 kDa complex. Our present study indicates that SPRI1 forms large protein machinery for the rejection of hetero-specific pollen in stigmatic papilla cells.
在植物中，存在一种活跃的合子前种间不亲和机制，以防止两个物种之间产生不良杂种。我们之前报道过，一种具有四个跨膜结构域的未表征蛋白，命名为柱头隐私1（SPRI1），负责在拟南芥中排斥异种花粉粒。然而，SPRI1中缺乏显著的功能结构域限制了我们对其生化特性的理解。在本研究中，我们通过点突变实验和生化分析对SPRI1蛋白进行了功能分析。我们探索了SPRI1的分子调控机制及其与功能的关系。丙氨酸和甘氨酸扫描实验以及进化分析表明，该蛋白胞外结构域C端区域的结构完整性对其功能很重要。此外，我们在胞外结构域中发现了两个半胱氨酸（C67和C80），它们可能参与分子间二硫键的形成。这些半胱氨酸残基是SPRI1蛋白稳定所必需的。此外，SPRI1可能形成同多聚体，并作为约300 kDa复合物的一部分存在。我们目前的研究表明，SPRI1在柱头乳头细胞中形成大型蛋白机制以排斥异种花粉。
