未折叠蛋白反应途径中的黄曲霉 hacA 基因是宿主诱导基因沉默的候选靶标

IF 4.2 2区 生物学 Q2 MICROBIOLOGY
Perng-Kuang Chang
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引用次数: 0

摘要

真菌 HacA/Hac1 转录因子在调节未折叠蛋白反应(UPR)中发挥着至关重要的作用。UPR 帮助细胞维持内质网(ER)蛋白质的平衡,这对生长、发育和毒力至关重要。黄曲霉的 hacA 基因编码一个富含碱性和酸性氨基酸(Bsc)的结构域和一个碱性亮氨酸拉链(bZip)结构域,并具有一个非常规内含子(Nt20)。本研究利用 CRISPR/Cas9 对 Bsc 编码、bZip 编码和 Nt20 序列进行了剖析,以阐明基因型与表型之间的关系。在 Bsc 和 bZip 实验集中,所有观察到的编码序列突变都在框内,这表明框外突变是致死的。Nt20 实验集中转化子的存活率很低,约为 7%。内含子中的突变主要包括框架外插入和缺失。除了类似野生型的分生孢子形态外,突变体还表现出不同的菌落形态,包括硬核、混合(分生孢子和硬核)和菌丝形态。使用二硫苏糖醇进行的ER压力测试表明,硬核突变体和菌丝突变体比分生孢子突变体敏感得多。此外,菌丝突变体不能产生黄曲霉毒素,但仍能产生曲霉酸和曲霉酸。针对包括 Bsc 和 bZip 的区域进行的 RNAi 实验表明,转化体的存活率普遍下降,少数转化体出现表型变化。hacA 基因在 DNA 和转录本水平上的缺陷影响了黄曲霉的存活、生长和发育。因此,该基因有可能成为未来宿主诱导基因沉默策略的目标,以控制感染并减少农作物中的黄曲霉毒素污染。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
The Aspergillus flavus hacA Gene in the Unfolded Protein Response Pathway Is a Candidate Target for Host-Induced Gene Silencing.

Fungal HacA/Hac1 transcription factors play a crucial role in regulating the unfolded protein response (UPR). The UPR helps cells to maintain endoplasmic reticulum (ER) protein homeostasis, which is critical for growth, development, and virulence. The Aspergillus flavus hacA gene encodes a domain rich in basic and acidic amino acids (Bsc) and a basic leucine zipper (bZip) domain, and features a non-conventional intron (Nt20). In this study, CRISPR/Cas9 was utilized to dissect the Bsc-coding, bZip-coding, and Nt20 sequences to elucidate the relationship between genotype and phenotype. In the Bsc and bZip experimental sets, all observed mutations in both coding sequences were in frame, suggesting that out-of-frame mutations are lethal. The survival rate of transformants in the Nt20 experiment set was low, at approximately 7%. Mutations in the intron primarily consisted of out-of-frame insertions and deletions. In addition to the wild-type-like conidial morphology, the mutants exhibited varied colony morphologies, including sclerotial, mixed (conidial and sclerotial), and mycelial morphologies. An ER stress test using dithiothreitol revealed that the sclerotial and mycelial mutants were much more sensitive than the conidial mutants. Additionally, the mycelial mutants were unable to produce aflatoxin but still produced aspergillic acid and kojic acid. RNAi experiments targeting the region encompassing Bsc and bZip indicated that transformant survival rates generally decreased, with a small number of transformants displaying phenotypic changes. Defects in the hacA gene at the DNA and transcript levels affected the survival, growth, and development of A. flavus. Thus, this gene may serve as a promising target for future host-induced gene-silencing strategies aimed at controlling infection and reducing aflatoxin contamination in crops.

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来源期刊
Journal of Fungi
Journal of Fungi Medicine-Microbiology (medical)
CiteScore
6.70
自引率
14.90%
发文量
1151
审稿时长
11 weeks
期刊介绍: Journal of Fungi (ISSN 2309-608X) is an international, peer-reviewed scientific open access journal that provides an advanced forum for studies related to pathogenic fungi, fungal biology, and all other aspects of fungal research. The journal publishes reviews, regular research papers, and communications in quarterly issues. Our aim is to encourage scientists to publish their experimental and theoretical results in as much detail as possible. Therefore, there is no restriction on paper length. Full experimental details must be provided so that the results can be reproduced.
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