Tian Chunyan, Rehman Abdul, Wang Xiaoyang, Wang Zhenzhen, Li Hongge, Ma Jun, Du Xiongming, Peng Zhen, He Shoupu
Zhengzhou Research Base, State Key Laboratory of Cotton Bio-breeding and Integrated Utilization, School of Agricultural Sciences, Zhengzhou University, Zhengzhou 450001, China.
State Key Laboratory of Cotton Bio-breeding and Integrated Utilization, Institute of Cotton Research, Chinese Academy of Agricultural Sciences (ICR, CAAS), Anyang, Henan 455000, China.
Gene. 2025 May 15;949:149372. doi: 10.1016/j.gene.2025.149372. Epub 2025 Feb 27.
The productivity and quality of cotton are significantly compromised by salt stress. In this study, the full length of encoding region and genomic DNA sequences of GhLEA_5A/D (Gh_A10G166600 and Gh_D10G188300), which belong to the late embryogenesis abundant gene family in allotetraploid upland cotton (Gossypium hirsutum L.) and semi-wild cotton (Gossypium purpurascens), were isolated and their salt tolerance was experimentally confirmed. Analysis of sequence alignments and phylogenetic trees indicated a significant level of homology between GhLEA-5A and GhLEA-5D. Additionally, a conserved protein motif was consistently identified across these sequences. The transcriptome data analysis showed that the expression level of GhLEA-5A/D was substantially enhanced in the leaves of salt-tolerant G. purpurascens accessions compared to salt-sensitive materials. In the real-time quantitative reverse transcription PCR (qRT-PCR) assays, notable expression levels of the GhLEA-5D gene were detected in salt-tolerant upland cotton materials following exposure to salt stress at 3 and 12-hour time points. The suppression of GhLEA-5A/D transcription via Virus-induced Gene Silencing (VIGS) technology significantly exacerbates salt sensitivity in cotton. This is evidenced by the nearly 50 % increase in malondialdehyde (MDA) content alongside a 60 % reduction in peroxidase (POD) levels in salt-treated plants when compared to the control group. The overexpression of the GhLEA-5A/D gene conferred enhanced salt tolerance in Arabidopsis, resulting in a 25 % increase in root length, a 30 % improvement in survival rate, a 15 % increase in water retention, and a 15 % boost in photosynthetic efficiency. The chlorophyll fluorescence parameters, enzyme activities, diaminobenzine, and nitroblue tetrazolium staining suggested that GhLEA-5A/D likely exhibited a positive regulatory role for cotton responding to salt stress. Furthermore, we identified 76 candidate proteins that potentially interact with GhLEA-5 in the yeast two-hybrid screening library. These results provide a theoretical basis for studying the mechanism of cotton salt tolerance and offer new resources for improving cotton salt tolerance genes.
盐胁迫严重影响棉花的产量和品质。在本研究中，从异源四倍体陆地棉（Gossypium hirsutum L.）和半野生棉（Gossypium purpurascens）中分离出属于晚期胚胎发生丰富基因家族的GhLEA_5A/D（Gh_A10G166600和Gh_D10G188300）的编码区全长和基因组DNA序列，并通过实验证实了它们的耐盐性。序列比对和系统发育树分析表明，GhLEA-5A和GhLEA-5D之间具有显著的同源性。此外，在这些序列中一致鉴定出一个保守的蛋白质基序。转录组数据分析表明，与盐敏感材料相比，耐盐的紫色棉材料叶片中GhLEA-5A/D的表达水平显著提高。在实时定量逆转录PCR（qRT-PCR）分析中，在耐盐陆地棉材料于3小时和12小时时间点暴露于盐胁迫后，检测到GhLEA-5D基因有显著的表达水平。通过病毒诱导基因沉默（VIGS）技术抑制GhLEA-5A/D转录会显著加剧棉花的盐敏感性。与对照组相比，盐处理植株中丙二醛（MDA）含量增加近50%，而过氧化物酶（POD）水平降低60%，这证明了这一点。GhLEA-5A/D基因的过表达赋予拟南芥增强的耐盐性，使根长增加25%，存活率提高30%，保水率增加15%，光合效率提高15%。叶绿素荧光参数、酶活性、二氨基联苯胺和氮蓝四唑染色表明，GhLEA-5A/D可能对棉花响应盐胁迫发挥正调控作用。此外，我们在酵母双杂交筛选文库中鉴定出76个可能与GhLEA-5相互作用的候选蛋白。这些结果为研究棉花耐盐机制提供了理论依据，并为改良棉花耐盐基因提供了新的资源。
