水稻谷氨酸脱羧酶4（GAD4）中钙调蛋白结合结构域的截短导致γ-氨基丁酸积累，并赋予水稻幼苗非生物胁迫耐受性。

Truncation of the calmodulin binding domain in rice glutamate decarboxylase 4 (GAD4) leads to accumulation of γ-aminobutyric acid and confers abiotic stress tolerance in rice seedlings.

作者信息

Akter Nadia, Kulsum Ummey, Moniruzzaman Mohammad, Yasuda Norito, Akama Kazuhito

机构信息

Graduate School of Natural Science and Technology, Shimane University, 1060 Nishikawatsu, Matsue, Shimane 690-8504 Japan.

Genetic Resources and Seed Division, Bangladesh Rice Research Institute, Gazipur, 1701 Bangladesh.

出版信息

Mol Breed. 2024 Feb 29;44(3):21. doi: 10.1007/s11032-024-01460-1. eCollection 2024 Mar.

DOI:10.1007/s11032-024-01460-1

PMID:38435472

原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC10904699/

Abstract

UNLABELLED

GABA (Gamma-aminobutyric acid) is a non-protein amino acid widely known as major inhibitory neurotransmitter. It is synthesized from glutamate via the enzyme glutamate decarboxylase (GAD). GAD is ubiquitous in all organisms, but only plant GAD has ability to bind Ca/calmodulin (CaM). This kind of binding suppresses the auto-inhibition of Ca/calmodulin binding domain (CaMBD) when the active site of GAD is unfolded resulting in stimulated GAD activity. is one of the five genes in rice genome. It was confirmed that has ability to bind to Ca/CaM. Moreover, it exhibits strongest expression against several stress conditions among the five genes. In this study, CRISPR/Cas9-mediated genome editing was performed to trim the coding region of CaMBD from the gene, to remove its autoinhibitory function. DNA sequence analysis of the genome edited rice plants revealed the truncation of CaMBD (216 bp). Genome edited line (#14-1) produced 11.26 mg GABA/100 g grain, which is almost nine-fold in comparison to wild type. Short deletion in the coding region for CaMBD yielded in mutant (#14-6) with lower GABA content than wild type counterpart. Abiotic stresses like salinity, flooding and drought significantly enhanced GABA accumulation in #14-1 at various time points compared to wild-type and #14-6 under the same stress conditions. Moreover, upregulated mRNA expression in vegetative tissues seems correlated with the stress-responsiveness of when exposed to the above-mentioned stresses. Stress tolerance of genome edited lines was evidenced by the higher survival rate indicating the gene may induce tolerance against abiotic stresses in rice. This is the first report on abiotic stress tolerance in rice modulated by endogenous GABA.

SUPPLEMENTARY INFORMATION

The online version contains supplementary material available at 10.1007/s11032-024-01460-1.

摘要

未标记

γ-氨基丁酸（GABA）是一种非蛋白质氨基酸，广为人知的主要抑制性神经递质。它由谷氨酸通过谷氨酸脱羧酶（GAD）合成。GAD在所有生物体中普遍存在，但只有植物GAD具有结合钙/钙调蛋白（CaM）的能力。当GAD的活性位点展开时，这种结合会抑制钙/钙调蛋白结合域（CaMBD）的自身抑制，从而导致GAD活性受到刺激。OsGAD2是水稻基因组中的五个GAD基因之一。已证实OsGAD2具有与钙/钙调蛋白结合的能力。此外，在五个OsGAD基因中，它在几种胁迫条件下表现出最强的表达。在本研究中，进行了CRISPR/Cas9介导的基因组编辑，以从OsGAD2基因中去除CaMBD的编码区域，消除其自身抑制功能。对基因组编辑水稻植株的DNA序列分析揭示了CaMBD（216bp）的截短。基因组编辑系（#14-1）产生11.26mg GABA/100g谷物，与野生型相比几乎高出九倍。CaMBD编码区域的短缺失产生了突变体（#14-6），其GABA含量低于野生型对应物。与相同胁迫条件下的野生型和#14-6相比，盐度、水淹和干旱等非生物胁迫在不同时间点显著增强了#14-1中的GABA积累。此外，营养组织中上调的mRNA表达似乎与OsGAD2在暴露于上述胁迫时的胁迫反应性相关。OsGAD2基因组编辑系的胁迫耐受性通过较高的存活率得到证明，表明该基因可能诱导水稻对非生物胁迫的耐受性。这是关于内源性GABA调节水稻非生物胁迫耐受性的首次报道。