向日葵GH3基因家族鉴定及其在花发育中的功能分析

doi:10.13560/j.cnki.biotech.bull.1985.2025-0560

摘要/Abstract

摘要：

目的探究向日葵（Helianthus annuus L.）HaGH3家族成员的理化特征和表达分析，以及其在向日葵花发育中的作用，为向日葵品种选育提供参考。方法基于向日葵基因组和转录组数据，利用生物信息学方法筛选鉴定HaGH3家族成员，并对蛋白理化性质、染色体定位、系统进化关系、保守基序、顺式作用元件和组织表达特征等进行分析。利用RT-qPCR分析HaGH3家族成员在野生型向日葵及其花分生组织转变与花器官发生缺陷突变体cb1中St2‒St8时期的表达差异，并对其在St5和St6时期不同花器官中的表达情况进行分析。结果向日葵HaGH3基因家族包含19个成员（HaGH3.1‒HaGH3.19），分散分布在12条染色体上，分属于2个亚家族，同一亚家族的成员具有相同的蛋白保守结构域。转录组数据分析显示，HaGH3.1、HaGH3.2、HaGH3.3、HaGH3.4、HaGH3.5、HaGH3.8、HaGH3.11、HaGH3.12、HaGH3.13、HaGH3.14和HaGH3.18等11个基因在向日葵花发育特定时期高水平表达。基于RT-qPCR的表达分析，显示HaGH3.3、HaGH3.4、HaGH3.8、HaGH3.13和HaGH3.14在WT的St2时期表达水平达到峰值，且其表达水平在WT和cb1中存在显著差异；HaGH3.1、HaGH3.2、HaGH3.5、HaGH3.12和HaGH3.18在WT花发育后期（St5‒St6时期）高水平表达，且具有较强的花器官特异性。顺式作用元件分析显示，11个花发育相关HaGH3基因启动子区域内含有大量生长素和赤霉素响应元件和MIKC_MADS识别元件。进一步分析结果显示，HaGH3.8、HaGH3.14和HaGH3.18受到ARF介导的生长素信号调节，HaGH3.1和HaGH3.8受到MYB和bZIP等赤霉素信号相关转录因子的调控，HaGH3.2、HaGH3.8、HaGH3.13、HaGH3.14和HaGH3.18受到花同源异型转录因子MIKC_MADS的调控。结论向日葵HaGH3.1和HaGH3.3等11个成员在向日葵花生长发育的调控中发挥重要作用，其中HaGH3.1、HaGH3.2、HaGH3.8、HaGH3.13、HaGH3.14和HaGH3.18在调控花发育过程中，受到生长素或赤霉素信号相关转录因子以及MIKC_MADS成员的调控。

关键词: 向日葵, 花发育, cb1突变体, GH3基因家族, 表达水平, 顺式作用元件, 转录因子, 转录调控

Abstract:

Objective Physicochemical property and expression feature of HaGH3 genes were analyzed to explore their functions in the floral development in sunflower (Helianthus annuus L.), and discover some molecular clues for sunflower breeding. Method Bioinformatics methods were adapted to identify the HaGH3 candidates based on the genome and transcriptome data of sunflower, and to analyze their physicochemical properties, chromosome localization, phylogenetic relationship, conserved motif, cis-acting elements, and tissue-expressing pattern. Subsequently, RT-qPCR was used to analyze the expression differences of HaGH3 genes between WT sunflower and cb1 (crazy broccoli 1) mutant in the phases of reproductive development (from St2 to St8), and to measure the expressions of HaGH3 genes in different flower organs of WT sunflower at St5 and St6 phases. Result The 19 members of HaGH3 gene family were in the sunflower genome, and named as HaGH3.1 to HaGH3.19, located on 12 chromosomes and categorized into two distinct subfamilies, and all members of the HaGH3 genes belonging to one subfamily possessed the same conserved domains. Transcription analysis indicated that 11 members presented the high expressions at the specific phase of flower development respectively, including HaGH3.1, HaGH3.2, HaGH3.3, HaGH3.4, HaGH3.5, HaGH3.8, HaGH3.11, HaGH3.12, HaGH3.13, HaGH3.14, and HaGH3.18. RT-qPCR analysis showed that the expressions of HaGH3.3, HaGH3.4, HaGH3.8, HaGH3.13, and HaGH3.14 reached their peak values in the St2 of WT, with significant differences between WT and cb1. HaGH3.1, HaGH3.2, HaGH3.5, HaGH3.12, and HaGH3.18 were highly expressed during St5 and St6 in WT, demonstrating obviously floral organs specificity. Cis-acting element analysis showed that a multitude of hormone response elements and MIKC_MADS recognition elements existed in the promoter regions of these 11 HaGH3 genes associated with floral development. Further analysis showed that HaGH3.8, HaGH3.14 and HaGH3.18 were regulated by ARF-mediated auxin signaling, HaGH3.1 and HaGH3.8 were regulated by gibberellin signal-related transcription factors such as MYB and bZIP, and HaGH3.2, HaGH3.8, HaGH3.13, HaGH3.14 and HaGH3.18 were regulated by the transcription factor MIKC_MADS. Conclusion Eleven members including HaGH3.1 and HaGH3.3 play important roles in regulating the growth and development of sunflower inflorescences. Among them, HaGH3.1, HaGH3.2, HaGH3.8, HaGH 3.13,HaGH 3.14 and HaGH 3.18 were regulated by auxin or gibberellin signal-related transcription factors and MADS-box members during flower development.

Key words: sunflower, flower development, cb1 mutant, GH3 gene family, expression level, cis-acting element, transcription factor, transcriptional regulation

龙林茜, 曾银萍, 王茜, 邓玉萍, 葛敏茜, 陈彦灼, 李鑫娟, 杨军, 邹建. 向日葵GH3基因家族鉴定及其在花发育中的功能分析[J]. 生物技术通报, 2026, 42(1): 125-138.

LONG Lin-xi, ZENG Yin-ping, WANG Qian, DENG Yu-ping, GE Min-qian, CHEN Yan-zhuo, LI Xin-juan, YANG Jun, ZOU Jian. Identification of Sunflower GH3 Gene Family and Analysis of Their Function in Flower Development[J]. Biotechnology Bulletin, 2026, 42(1): 125-138.

图/表 7

图1 HaGH3 基因家族成员的染色体定位（A）及系统进化分析（B）

Fig. 1 Chromosome localization (A) and phylogenetic analysis (B) of HaGH3 gene family members

图2 HaGH3基因结构和蛋白保守基序分布A：HaGH3基因编码的蛋白保守基序；B：保守结构域图；C：基因结构图

Fig. 2 Gene structure and protein conserved motif distribution of HaGH3A: Conserved motif of protein encoded by HaGH3 gene. B: Conserved domain diagram. C: Gene structure map

图3 HaGH3 家族基因在WT向日葵不同组织的表达分析St2：花原基起始期；St3：花器官开始形成期；St4：减数分裂期；St5：花粉成熟期；St6：小花开放期；St7：果实灌浆期；St8：种子饱满期；下同

Fig. 3 Expression analysis of HaGH3 family genes in different tissues of WT sunflowerSt2: Initiation phase of flower primordium. St3: Phase of flower organs starting formation. St4: Meiosis phase. St5: Phase of pollen maturity. St6: Floret opening phase. St7: Fruit filling phase. St8: Seed-full phase. The same below

图4 HaGH3基因在野生型（WT）向日葵和cb1突变体中的差异表达分析A：WT向日葵和cb1突变体的花盘；B：WT向日葵和cb1突变体花序纵剖图；C：HaGH3基因在WT花和cb1突变体类花序组织（St2‒St8）的相对表达水平分析。数据为平均值±SD（n=3）；*P<0.05，**P<0.01，下同

Fig. 4 Differential expression analysis of HaGH3 gene in WT sunflower and cb1 mutantA: Flower disc of WT sunflower and cb1 mutant. B: Longitudinal section of WT sunflower and cb1 mutant inflorescences. C: Relative expression of HaGH3 gene during inflorescence development of WT and cb1 mutants (St2‒St8). All data are mean ±SD (n=3). * P < 0.05, ** P < 0.01. The same below

图5 HaGH3家族在不同时期花器官中的表达水平A：向日葵St5时期管状花解剖图；B：向日葵St6时期管状花解剖图；C：HaGH3基因家族在St5时期花器官中表达水平；D：在St6时期花器官中表达水平；数据以平均值±SD表示（n=3）；不同小写字母表示组间差异具有统计学意义（P<0.05）

Fig. 5 Expressions of the HaGH3 family in flower organs at different phasesA: Anatomy of tubular flowers at St5 phase of sunflower. B: Anatomy of tubular flowers from the St6 phase of sunflower. C Expressions of sunflower GH3 gene family in flower organs at St5 phase. D: Expressions in flower organs at St6 phase. All data are mean ± SD (n=3). Groups marked with different lowercase letters indicate statistically significant differences (P < 0.05)

图6 HaGH3 基因启动子区顺式作用元件及其调控网络预测分析A：HaGH3基因顺式作用元件；B：HaGH3.8和HaGH3.14与ARF转录因子互作预测；C：HaGH3.8与MYB、bZIP转录因子互作预测；D：HaGH3.8、HaGH3.13和HaGH3.14基因与MIKC_MADS转录因子互作预测；E：HaGH3.18与ARF转录因子互作预测；F：HaGH3.1基因与MYB、bZIP转录因子互作预测；G：HaGH3.18基因与MIKC_MADS转录因子互作预测

Fig. 6 Prediction analysis of cis-acting elements and regulatory networks of the promoter region of HaGH3 geneA: Cis-acting element of the HaGH3 gene. B: Prediction of interaction between HaGH3.8 and HaGH3.14 with ARF transcription factors. C: Prediction of interaction between HaGH3.8 and MYB and bZIP transcription factors. D: Prediction of interaction between HaGH3.8, HaGH3.13 and HaGH3.14 genes with MIKC_MADS transcription factors. E: Prediction of interaction between HaGH3.18 and ARF transcription factors. F: Prediction of interaction between HaGH3.1 gene and MYB and bZIP transcription factors. G: Prediction of interaction between HaGH3.18 gene and MIKC_MADS transcription factors

图7 参与花发育调控HaGH3基因及其候选调控转录因子的相对表达水平A：HaGH3.8和HaGH3.14与其潜在上游转录调控因子ARF的相对表达水平；B：HaGH3.8与其潜在上游转录调控因子MYB、bZIP的相对表达水平；C：HaGH3.8、HaGH3.13和HaGH3.14与其潜在上游转录调控因子MIKC_MADS的相对表达水平；D：HaGH3.18与其潜在上游转录调控因子ARF的相对表达水平；E：HaGH3.1与其潜在上游转录调控因子MYB、bZIP的相对表达水平；F：HaGH3.18与其潜在上游转录调控因子MIKC_MADS的相对表达水平

Fig. 7 Relative expressions of HaGH3 gene and its candidate transcription factors involved in the regulation of flower developmentA: Relative expressions of HaGH3.8 and HaGH3.14 and their potential upstream transcriptional regulators ARF. B: Relative expressions of HaGH3.8 and their potential upstream transcriptional regulators MYB and bZIP. C: Relative expressions of HaGH3.8, HaGH3.13 and HaGH3.14 and their potential upstream transcriptional regulators MIKC_MADS. D: Relative expressions of HaGH3.18 and its potential upstream transcriptional regulators ARF. E: Relative expressions of HaGH3.1 and its potential upstream transcriptional regulators MYB and bZIP. F: Relative expressions of HaGH3.18 and its potential upstream transcriptional regulatorsMIKC_MADS

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