Functional Analysis Of Tomato SlMADS48 Gene

The development of angiosperm floral organs is an extremely complex process,regulated by multiple internal genetic factors and external environmental signals.As one of the largest transcription factor family in plants,MADS-box transcription factors are extremely vital to the growth and development of organs in angiosperms,especially the development of floral organs.In this study,tomato was used as the experimental material,using bioinformatics,histocytology,yeast two-hybrid,RNA-seq and other strategies to explore the biological functions of the MADS-box transcription factor SlMADS48 in tomato.In addition,we preliminarily clarified the molecular mechanism of SlMADS48gene involved in regulating the development of tomato sepal.The key research findings were as followed:(1)The sequence analysis of the SlMADS48 gene showed that the full-length of c DNA was 966 bp,encoding a protein with 195 amino acids,containing a typical MADS-box region.In addition,the analysis of cis-acting elements showed that the promoter of SlMADS48 contained phytohormone response elements,light and stress response elements.(2)The analysis of the tissue expression profile showed that m RNA accumulation of SlMADS48 gene was highest in fruits 7 days after breaking(B+7),followed by stems and sepals,and the least in senescent leaves.The analysis of abiotic stress and hormone expression profile showed that this gene can respond to drought,high salinity and plant hormone auxin/gibberellin.(3)Using Agrobacterium LBA4404-mediated transformation,the SlMADS48overexpression vector was transformed into tomato AC⁺⁺,and three SlMADS48transgenic lines with higher overexpression level were obtained.(4)The growth rate of SlMADS48 overexpression plant seedlings was significantly faster than that of AC⁺⁺seedlings.q RT-PCR results showed that auxin-and gibberellin-related genes were significantly accumulated in the leaves of SlMADS48 overexpression plants.In addition,after treated by paclobutrazol(PAC)and 1-naphthylacetic acid(NAA),the growth rate of SlMADS48 overexpression plant seedlings were slower than that of AC⁺⁺,indicating that SlMADS48 overexpression plants were more sensitive to exogenous PAC and NAA.(5)The sepal size of SlMADS48 transgenic plants was larger than that of wild type.The results of histocytology analysis showed that the number of cells in the sepals of transgenic plants increased significantly than that of the wild type.Using q RT-PCR technology to detect and analyze related genes,the results showed that cell cycle,cell division and gibberellin-related genes were all significantly induced in sepals of the transgenic lines,indicating that the SlMADS48 gene probably affect cell cycle,division and gibberellin-related gene expression,regulating the process of cell proliferation and ultimately affecting the development of tomato sepals.RNA-seq results showed that overexpression of the SlMADS48 gene caused the expression of cell division-related,gibberellin-related and auxin-related genes in tomato sepals to be significantly induced,which is consistent with the q RT-PCR results of sepals.(6)Yeast two-hybrid experiments showed that SlMADS48 protein can interact with MC,JOINTLESS,Sl MBP21 and Sl FYFL proteins,which have been determined to regulate the development of sepals.It showed that the SlMADS48 regulated the development of sepals by interacting with the four proteins above possibly,leading to changes in the sepal size of transgenic plants.(7)SlMADS48 overexpression plants have weak drought tolerance,with the faster water loss rate,and lower relative water content.Moreover,the content of hydrogen peroxide and MDA was higher,and soluble sugar and free proline lower.It showed that the overexpression of SlMADS48 gene reduced the resistance of tomato plants to drought stress.In summary,we found that the MADS-box transcription factor SlMADS48 regulates the development of tomato sepals by affecting cell proliferation in tomato sepals possibly.Furthermore,we preliminarily clarified that the SlMADS48 protein may regulate the development of tomato sepals by interacting with four sepal development-related proteins(MC,JOINTLESS,Sl MBP21 and Sl FYFL).In addition,we analyzed that overexpression of SlMADS48 gene affects tomato plant seedling height by affecting the expression of auxin-and gibberellin-related genes probably,and explored that overexpression of SlMADS48 gene could reduce the drought resistance of plants by breaking the homeostasis of plant cells.