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Understanding gene expression kinetics and the underlying physiological mechanisms in stress combinations is a challenge for the purpose of stress resistance breeding. The novelty of this study is correlating the physiological mechanisms with the expression of key target genes in tall fescue under a combination of various salinity and osmotic stress treatments. Four drought- and salt-responsive genes belonging to different crucial pathways evaluated included one transcription factor FabZIP69, one for the cytosolic polyamine synthetase FaADC1, one for ABA signaling FaCYP707A1, and another one for the specific Na + /H + plasma membrane antiporter FaSOS1 involve in osmotic homeostasis. FaSOS1, FaCYP707A1, and FabZIP69 were induced early at 6 h after NaCl treatment, while FaSOS1 and FaCYP707A1 were transcribed gradually after exposure to PEG. However, stress interactions showed a significantly increased expression in all genes. Expression of these genes was positively correlated to Pro, SSs, IL, DPPH, and antioxidant enzyme activity and negatively correlated with RWC, total Chl, and MSI. Chemical analyses showed that tall fescue plants exposed to the combination of stresses exhibited increased quantity of reactive oxygen species (H 2 O 2 ), EL and DPPH, and higher levels of antioxidant enzyme activities (CAT, and SOD), Pro, and SSs content, compared with control seedlings. Under dual-stress conditions, the expression of FabZIP69 was effective in controlling the expression of FaSOS1 and FaADC1 genes differently., (© 2023. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Somatic embryogenesis (SE) depends on a variety of developmental pathways that are influenced by several environmental factors. Therefore, it is important to understand the relationship between environmental and genetic factors by identifying the gene networks involved in SE through gene set enrichment analysis (GSEA). For determination of SE effective transcription factors, upstream sequences of core-enriched genes were analyzed. The results indicated that response to hormones is one of the biological pathways activated by the enriched TFs at all stages of somatic embryogenesis and about half of the hormonal pathways were enriched. On the fifth day after 2,4-Dichlorophenoxyacetic acid (2,4-D) treatment, the activity of hormone-affecting genes reached its maximum. At this time, more transcription factors regulated the enriched genes compared to the other stages of somatic embryogenesis. MYBs, AT-HOOKs, and HSFs are the main families of transcription factors which affect core-enriched genes during SE. CCA1 , PRR7 , and TOC1 and their related genes at the center of protein-protein interaction of SE-key transcription factors, involved in the regulation of the circadian clock. Gene expression analysis of CCA1 , PRR7 , and TOC1 revealed that the genes involved in circadian clock reached their maximum activity when embryonic cells formed. Also, auxin response elements were identified at the upstream of SE-circadian clock transcription factors, indicating that they might mediate between auxin signaling and SE-related hormonal pathways as well as SE marker genes such as AGL15 , BBM , and LEC s. Based on these results, it is possible that the cellular circadian rhythm activates various developmental pathways under the influence of auxin signal transduction and their interactions determine the induction of somatic embryogenesis . According to the results of this study, modifying pathways affected by SE-related transcription factors such as circadian rhythm may result in cell reprogramming and increase somatic embryogenesis efficiency., Supplementary Information: The online version contains supplementary material available at 10.1007/s13205-023-03546-7., Competing Interests: Conflict of interestThe authors have no relevant financial or non-financial interests to disclose., (© King Abdulaziz City for Science and Technology 2023, Springer Nature or its licensor (e.g. a society or other partner) holds exclusive rights to this article under a publishing agreement with the author(s) or other rightsholder(s); author self-archiving of the accepted manuscript version of this article is solely governed by the terms of such publishing agreement and applicable law.)