C-terminal frameshift mutations generate viable knockout mutants with developmental defects for three essential protein kinases.

C 端移码突变产生具有三种必需蛋白激酶发育缺陷的可行敲除突变体。

• 发表时间: 2024 Jun
• 来源期刊: aBIOTECH
• DOI: 10.1007/s42994-024-00165-5
• PMID: 38974866
• 文章类型: Journal Article
• 作者列表: Zhang Y,Cui MM,Ke RN,Chen YD,Xie K

关键词: C-terminus; CRISPR/Cas9; Genome editing; Lethal mutant; Protein kinases

Abstract

Loss-of-function mutants are fundamental resources for gene function studies. However, it is difficult to generate viable and heritable knockout mutants for essential genes. Here, we show that targeted editing of the C-terminal sequence of the embryo lethal gene MITOGEN-ACTIVATED PROTEIN KINASES 1 (OsMPK1) results in weak mutants. This C-terminal-edited osmpk1 mutants displayed severe developmental defects and altered disease resistance but generated tens of viable seeds that inherited the mutations. Using the same C-terminal editing approach, we also obtained viable mutants for a wall-associated protein kinase (Os07g0493200) and a leucine-rich repeat receptor-like protein kinase (Os01g0239700), while the null mutations of these genes were lethal. These data suggest that protein kinase activity could be reduced by introducing frameshift mutations adjacent to the C-terminus, which could generate valuable resources for gene function studies and tune protein kinase activity for signaling pathway engineering.
UNASSIGNED: The online version contains supplementary material available at 10.1007/s42994-024-00165-5.

摘要

功能缺失突变体是基因功能研究的基础资源。然而,很难为必需基因产生可行和可遗传的敲除突变体。这里,我们表明，胚胎致死基因有丝分裂原激活的蛋白激酶1（OsMPK1）的C端序列的靶向编辑导致弱突变体。这种C端编辑的osmpk1突变体表现出严重的发育缺陷和改变的抗病性，但产生了数十个遗传突变的可行种子。使用相同的C端编辑方法，我们还获得了壁相关蛋白激酶（Os07g0493200）和富含亮氨酸重复受体样蛋白激酶（Os01g0239700）的可行突变体，而这些基因的无效突变是致命的。这些数据表明，蛋白激酶活性可以通过在C端附近引入移码突变来降低，这可以为基因功能研究提供有价值的资源，并为信号通路工程调节蛋白激酶活性。
■在线版本包含补充材料，可在10.1007/s42994-024-00165-5获得。