过量表达小麦TaCYP78A5增加花器官的大小

doi:10.3864/j.issn.0578-1752.2023.09.002

摘要/Abstract

摘要：

【目的】利用表达模式分析、转基因过表达和细胞学观察等策略，解析TaCYP78A5调控花器官大小的功能和机制，为作物遗传改良提供基因资源和理论基础。【方法】根据EnsemblPlants基因组数据库中不同物种CYP78A家族成员的序列信息，对小麦TaCYP78A5和其他物种中的同源基因进行序列比对和进化分析；利用生物信息学分析小麦TaCYP78A5的基因和蛋白结构，以及不同器官的表达模式；通过在拟南芥中组成型过表达和生殖器官局部特异性过表达TaCYP78A5的策略，明确TaCYP78A5具有调控花器官大小的功能；利用显微镜观察不同转基因拟南芥花器官的细胞学特征，解析TaCYP78A5调控花器官大小的细胞学机制；利用小麦转基因过表达策略，明确TaCYP78A5调控小麦穗部大小等其他穗部性状的功能；利用323份小麦品种的单倍型数据与穗部表型数据进行关联分析，探析不同小麦品种TaCYP78A5表达量的高低对穗部大小等其他穗部性状的影响。【结果】小麦TaCYP78A5与拟南芥AtCYP78A5的基因和蛋白序列相似性较低，但基因和蛋白结构相似性较高。小麦TaCYP78A5和拟南芥AtCYP78A5均在多个器官广泛表达，但在花器官中表达量最高。相较于野生型，在拟南芥中组成型过表达TaCYP78A5能导致花器官增大，花瓣面积显著增加13.5%—35.4%；而且仅在胚珠局部特异性过表达TaCYP78A5也足以导致拟南芥花器官增大，花瓣面积显著增加9%—22.1%；相反，拟南芥cyp78a5突变体的花器官则显著小于野生型，花瓣面积显著减少27%。在拟南芥中组成型过表达TaCYP78A5导致花瓣表皮细胞大小较野生型显著增加49%—54%，细胞数目较野生型显著减少11%—19%，局部特异性过表达TaCYP78A5也导致花瓣表皮细胞大小较野生型显著增加20%—49%，细胞数目较野生型显著减少8%—24%。在小麦中组成型过表达TaCYP78A5导致小麦穗长增加7.9%—8.9%，颖壳面积增大9.6%—14.7%，穗粒数增加12.4%—23.8%，小穗数和小穗粒数呈现不同程度的变化。单倍型分析结果显示，在323份小麦品种中，具有较高TaCYP78A5-A表达量的小麦品种比具有较低TaCYP78A5-A表达量的小麦品种的穗长更长，小穗粒数更多，小穗数更少，而穗粒数无显著差异。【结论】TaCYP78A5以一种非细胞自制的模式促进花器官生长，小麦和拟南芥过量表达该基因均能导致花器官的增大。

关键词: 小麦, TaCYP78A5, 过量表达, 花器官大小, 细胞扩张

Abstract:

【Objective】The function and mechanism of TaCYP78A5 regulating flower organ size was preliminarily analyzed by means of expression pattern analysis, transgenic overexpression and cytological observation. The results provide genetic resources and theoretical basis for crop genetic improvement. 【Method】According to the sequence information of CYP78A family members of different species in EnsemblePlants genome database, sequence alignment and evolutionary analysis were carried out for the homologous genes of TaCYP78A5 in wheat and other species. The gene and protein structures and the expression patterns of different organs of wheat TaCYP78A5 were analyzed by bioinformatics. Through the strategy of constitutive overexpression and local specific overexpression in reproductive organs of TaCYP78A5 in Arabidopsis, it is clear that TaCYP78A5 has the function of regulating flower organ size. The cytological characteristics of flower organs of different transgenic Arabidopsis were observed under microscope, and the cytological mechanism of TaCYP78A5 regulating flower organ size was analyzed. The function of TaCYP78A5 in regulating wheat spike size and other spike traits was clarified by using the strategy of transgenic overexpression in wheat. The correlation analysis of haplotype data and spike phenotype data of 323 wheat accessions was used to explore the effect of TaCYP78A5 expression on spike size and other spike traits of different wheat accessions. 【Result】The gene and protein sequence similarity of wheat TaCYP78A5 and Arabidopsis AtCYP78A5 is low, but the gene and protein structure similarity is high. Wheat TaCYP78A5 and Arabidopsis AtCYP78A5 are widely expressed in many organs, but highly expressed in flower organs. Compared with wild type, the constitutive overexpression of TaCYP78A5 in Arabidopsis could lead to the enlargement of flower organs and a significant increase in petal area of 13.5%-35.4%. Moreover, the specific overexpression of TaCYP78A5 only in the ovule was enough to cause the enlargement of the flower organ of Arabidopsis, and the petal area increased significantly by 9%-22.1%. On the contrary, the flower organ of Arabidopsis cyp78a5 mutant was significantly smaller than that of wild type, and the petal area was significantly reduced by 27%. The constitutive overexpression of TaCYP78A5 in Arabidopsis resulted in a significant increase in the size of petal epidermal cells by 49%-54% compared with wild type, and a significant decrease in the number of cells by 11%-19% compared with wild type. Locally specific overexpression of TaCYP78A5 in Arabidopsis also resulted in a significant increase in the size of petal epidermal cells by 20%-49% compared with wild type, and a significant decrease in the number of cells by 8%-24% compared with wild type. The constitutive overexpression of TaCYP78A5 in wheat resulted in the increase of wheat spike length by 7.9%-8.9%, glume area by 9.6%-14.7%, and grain number per spike by 12.4%-23.8%. The spikelet number per spike and grain number per spikelet showed different degrees of change. The results of haplotype analysis showed that among 323 wheat accessions, wheat accessions with higher TaCYP78A5-A expression level had longer spike length, more grains per spikelet and fewer spikelets per spike than wheat accessions with lower TaCYP78A5-A expression level, but there was no significant difference in grain number per spike. 【Conclusion】TaCYP78A5 promoted the growth of flower organs in a non cellular self-made mode. The overexpression of TaCYP78A5 in wheat and Arabidopsis could lead to the enlargement of flower organs.

Key words: wheat, TaCYP78A5, overexpression, flower organ size, cell expansion

彭海霞, 卡得艳, 张天星, 周梦蝶, 吴林楠, 辛转霞, 赵惠贤, 马猛. 过量表达小麦TaCYP78A5增加花器官的大小[J]. 中国农业科学, 2023, 56(9): 1633-1645.

PENG HaiXia, KA DeYan, ZHANG TianXing, ZHOU MengDie, WU LinNan, XIN ZhuanXia, ZHAO HuiXian, MA Meng. Overexpression of Wheat TaCYP78A5 Increases Flower Organ Size[J]. Scientia Agricultura Sinica, 2023, 56(9): 1633-1645.

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