Nowak Katarzyna, Gaj Małgorzata D
Department of Genetics, University of Silesia, Jagiellonska 28, 40-032 Katowice, Poland.
Department of Genetics, University of Silesia, Jagiellonska 28, 40-032 Katowice, Poland.
J Plant Physiol. 2016 Apr 1;193:119-26. doi: 10.1016/j.jplph.2016.02.012. Epub 2016 Mar 2.
The bHLH109 gene of the bHLH family was identified among the transcription factor encoding genes that were differentially expressed in an embryogenic culture of Arabidopsis. A strong activation of bHLH109 expression was found to be associated with somatic embryogenesis (SE) induction. Several pieces of evidence suggested the involvement of bHLH109 in SE, including the high stimulation of the gene expression in SE-induced explants, which contrasts to the drastically lower level of the gene transcripts in the non-embryogenic callus and in tissue that is induced towards shoot regeneration via organogenesis. Moreover, in contrast to the overexpression of bHLH109, which has been indicated to enhance SE induction in a culture, the bhlh109 knock-out mutation was found to impair the embryogenic potential of explants. In order to identify the genes interacting with the bHLH109, the candidate co-expressed genes were identified in a yeast one hybrid assay. The in vitro regulatory interactions that were identified were verified through mutant and expression analysis. The results suggest that in SE bHLH109 acts as an activator of ECP63, a member of the LEA (LATE EMBRYOGENESIS ABUNDANT) family. Among the potential regulators of bHLH109, three candidates (At5g61620, bZIP4 and bZIP43) were indicated to possibly control bHLH109. The functions of all of the genes that are assumed to interact with bHLH109 are annotated to stress responses. Collectively, the results of the study provide new evidence that cell responses to stress that is imposed under in vitro conditions underlies the promotion of SE. bHLH109 may play a central role in the stress-related mechanism of SE induction via an increased accumulation of the LEA protein (ECP63), which results in the enhanced tolerance of the cells to stress.
在拟南芥胚性培养中差异表达的转录因子编码基因中，鉴定出了bHLH家族的bHLH109基因。研究发现，bHLH109表达的强烈激活与体细胞胚胎发生（SE）诱导有关。多项证据表明bHLH109参与了SE过程，包括在SE诱导的外植体中该基因表达受到高度刺激，这与非胚性愈伤组织和通过器官发生诱导芽再生的组织中该基因转录本水平极低形成对比。此外，与已表明能增强培养中SE诱导的bHLH109过表达相反，发现bhlh109基因敲除突变会损害外植体的胚性潜能。为了鉴定与bHLH109相互作用的基因，在酵母单杂交试验中鉴定了候选共表达基因。通过突变体和表达分析验证了所鉴定的体外调控相互作用。结果表明，在SE过程中，bHLH109作为LEA（胚胎后期丰富蛋白）家族成员ECP63的激活因子发挥作用。在bHLH109的潜在调控因子中，有三个候选基因（At5g61620、bZIP4和bZIP43）被指出可能调控bHLH109。所有假定与bHLH109相互作用的基因的功能都与应激反应相关。总体而言，该研究结果提供了新的证据，即细胞对体外条件下施加的应激的反应是促进SE的基础。bHLH109可能通过增加LEA蛋白（ECP63）的积累在SE诱导的应激相关机制中发挥核心作用，这导致细胞对应激的耐受性增强。
