| Tomato is a typical respiratory leap fleshy berry and is an important model plant for studying fruit development.The formation of locular gel is an important developmental process and a typical feature of tomato and other fleshy berries.Tomato locular gel is the second most abundant tissue in the fruit after the pericarp,accounting for about 23% of the fresh mass of the fruit.Programmed cell death is a fundamental physiological mechanism for the active removal of senescent necrotic cells in plants,and plays an important role in maintaining normal growth and development and physiological metabolism.For plants,PCD is prevalent during plant development and is a cell death process that occurs at specific times and in specific regions of plant development,and is the final stage of plant cell differentiation.In this thesis,the role of the programmed cell death gene SlLFG2 in tomato was investigated in the context of Micro-Tom tomato,with the following main results.(1)qPCR results showed that SlLFG2 gene was expressed in all tissues of tomato,with higher expression levels in locular gel.GUS promoter staining showed that SlLFG2 gene was expressed in locular gel at all stages of tomato fruit.SlLFG2 gene transgenic lines had the following phenotypes.First,development of tomato locular gel was accelerated after overexpression of the SlLFG2 gene.Inhibition of SlLFG2 gene expression delayed the development of locular gel.Second,overexpression of the SlLFG2 gene accelerated the elongation of plant roots and stems.The elongation of plant roots and stems was significantly inhibited by inhibiting the expression of SlLFG2 gene.(2)The transcriptome changes of SlLFG2 transgenic lines were analyzed by RNA-Seq.RNA-Seq identified 5179 differentially expressed genes between wild type and overexpressing lines and 3438 differentially expressed genes between wild type and interfering lines,of which 2002 genes were differentially expressed in different lines compared with the wild type.Through the construction of gene co-expression networks,SlATG8 f and SlERF5 were screened for possible interactions with SlLFG2 protein,where the SlATG8 f gene was associated with autophagic activity and the SlERF5 gene was associated with ethylene response.Membrane yeast two-hybrid,bimolecular fluorescence complementation(BIFC)and immunoprecipitation(Co-IP)experiments verified that SlLFG2 proteins can interact with SlATG8 f and SlERF5 proteins.(3)Three genes,SlLFG2,SlATG8 f and SlERF5,are involved in programmed cell death,autophagy and ethylene response processes,respectively.These three physiological processes were examined in SlLFG2 transgenic lines.It was verified that SlLFG2 gene inhibited programmed cell death and affected autophagic activity and ethylene response in plants.Two knockout lines of SlATG8 f and three interfering lines of SlERF5 were obtained,and the effects of SlATG8 f and SlERF5 genes on programmed cell death,autophagy and ethylene response processes were clarified.(4)Gibberellin promotes plant cell elongation and stem elongation.Protein interactions between SlLFG2 and the gibberellin oxidase gene SlGA2 were verified.The interfering lines of SlLFG2 showed a deficiency in responsiveness to gibberellin.Dark treatment experiments revealed that light signals affected the function of SlLFG2 in response to gibberellin.SlLFG2 could affect the plant response to gibberellin by binding to SlGA2 and suppressing its expression.In summary,SlLFG2 interacts with SlATG8 f and SlERF5 proteins to affect PCD,cell autophagy,and ethylene response,thus regulating the formation of tomato locular gel.SlLFG2 interacts with SlGA2 to affect gibberellin metabolic process,thus regulating plant growth process.Our results demonstrate that PCD,cell autophagy,hormone response and metabolic processes interact with each other to influence multiple processes in tomato growth and development.Our study provides new insights to resolve the interactions of multiple physiological processes involved in plant growth and development. |
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