GS5 encoding a serine carboxypeptidase-like protein positively regulates grain size and weight through the regulation of grain width and filling and is helpful in improving cereal yields. Grain width variation determined by GS5 is associated with cell number and size, but the actual underlying mechanism is still unclear. Two orthologs of GS5, TtGS5-3A-G and TtGS5-3G-G, were cloned from the Triticum timopheevi accession no. CWI17006. To identify the proteins that interacted with TtGS5-3A-G and TtGS5-3G-G in premature grains, we performed pull-down assays followed by liquid chromatography-mass spectrometry/mass spectrometry analysis. The analyses revealed 18 proteins were present in both the TtGS5-3A-G and TtGS5-3G-G interactomes. Among five candidates selected, only Annexin D1 interacted with both TtGS5-3A-G and TtGS5-3G-G in yeast. Annexin D1, TtGS5-3A-G, and TtGS5-3G-G were located on the cytoplasmic membranes of Arabidopsis protoplasts and onion epidermal cells, and interactions between Annexin D1 and TtGS5-3A-G, as well as TtGS5-3G-G, were shown by bimolecular fluorescence complementation assays. Annexin D1 was expressed widely in different tissues, and it co-expressed with TtGS5-3A-G/TtGS5-3G-G at the grain enlargement phase. These results indicated that Annexin D1 interacted with TtGS5-3A-G and TtGS5-3G-G in premature grains. Together with the structural similarities of Annexin D1 to known fiber elongation factors, we proposed that TtGS5 might regulate the cell size by interacting with Annexin D1. The results provide significant new information for understanding the roles that GS5 plays in regulating grain size, which may be useful in improving crop yields.

中文翻译：

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小麦小麦种子中丝氨酸羧肽酶样蛋白TtGS5和膜联蛋白D1的相互作用。

编码丝氨酸羧肽酶样蛋白的GS5通过调节谷粒宽度和填充来正向调节谷粒大小和重量，并有助于提高谷物产量。由GS5确定的晶粒宽度变化与细胞数量和大小有关，但实际的潜在机制仍不清楚。两个同源基因GS5，TtGS5-3A-G和TtGS5 - 3G-G ，从克隆的小麦timopheevi登记号 CWI17006。为了鉴定在早熟谷物中与TtGS5-3A-G和TtGS5-3G-G相互作用的蛋白质，我们进行了下拉分析，然后进行液相色谱-质谱/质谱分析。分析显示，TtGS5-3A-G和TtGS5-3G-G相互作用基因组中均存在18种蛋白质。在选定的五种候选物中，只有膜联蛋白D1与酵母中的TtGS5-3A-G和TtGS5-3G-G相互作用。Annexin D1，TtGS5-3A-G和TtGS5-3G-G位于拟南芥的细胞质膜上通过双分子荧光互补分析显示了原生质体和洋葱表皮细胞，以及膜联蛋白D1和TtGS5-3A-G以及TtGS5-3G-G之间的相互作用。膜联蛋白D1在不同的组织中广泛表达，并在晶粒增大阶段与TtGS5-3A-G / TtGS5-3G-G共表达。这些结果表明膜联蛋白D1与早熟谷物中的TtGS5-3A-G和TtGS5-3G-G相互作用。连同膜联蛋白D1与已知纤维伸长因子的结构相似性，我们提出TtGS5可能通过与膜联蛋白D1相互作用来调节细胞大小。结果为理解GS5在调节籽粒大小中的作用提供了重要的新信息，这可能对提高农作物的产量有用。