Transcriptome-wide Effects Of Loss Of CYCLOPHILIN71 Function On Gene Expression During Seedlings And Floral Buds Development In Arabidopsis

In Arabidopsis,AtCYP71 is a multidomain cyclophilin that contains a typical peptidyl prolyl cis-trans isomerase(PPIase)catalytic domain in the C-terminal region and four WD40 repeats in the N-terminal region.In one side,previous studies reported that AtCYP71 expresses in almost all plant tissues,and that AtCYP71 physically interacts with histone H3,like heterochromatin protein 1(LHP1),and chromatin assembly factor-1(FAS1),respectively,and is involved in histone H3K27 methylation,multiple chromatin remodeling process,and then functions in gene repression and pleiotropic development including meristem activity,leaf,root,and flower development.In other side,previous studies also reported that many non-coding RNA(ncRNA)play crucial roles in plant development via regulating the expression of related genes,especially transcriptional factors.For example,microRNAs(miRNAs),a class of 20-24 nucleotides noncoding RNAs,participate in many aspects of development in plant,including leaf,root and flower development and organogenesis.However,up to date,studies on the functions of ncRNAs mediated by AtCYP71 in growth and development have been rarely reported.In this study,we analysed the transcriptome-wide gene expression in wild type and cyp71 mutant during seedling and floral buds development in Arabidopsis by high-throughput sequencing and bioinformatics,and discussed whether AtCYP71 functions in seedlings and floral buds development via potentially mediating ncRNAs.In this study,the total RNA were extracted from 10-day-old seedlings of wild type and cyp71 mutant,and then two sRNA libraries were constructed and sequenced via high through-put sequencing platform,respectively.A total of 7,559,382 and 7,497,107 clean reads were obtained from the two libraries,respectively.In total,253 known miRNAs belonging to 118 miRNA families were identified.Among these,96 differentially expressed known miRNAs were identified,in which 30 miRNAs were upregulated and 66 miRNAs were downregulated.Via degradome analysis,337 transcripts targeted by 95 known miRNAs were detected.Notably,functional analysis exhibited that several target genes(including NAC1,SCL6&SCL23,TOE2,AGL16,and AFB1&TIR1)of miRNAs differentially expressed,function in meristem,lateral organ development.Overall,these results suggested that loss of AtCYP71 function directly or indirectly contribute to alter the expression of some key miRNAs and their target genes,and then might result in some defect phenotypes of organogenesis and development in Arabidopsis cyp71 seedling.Flower development is an important trait for spermatophytes.To survey the expression pattern of related genes encoding proteins and ncRNAs during floral bud development affected by loss of AtCYP71 function,we performed the whole transcriptome study on floral buds(younger than 12th stage)by combining strand-specific RNA-sequencing,sRNA-sequencing and degradome sequencing.Compared with wild type,we identified 132 upregulated and 562 downregulated differentially expressed genes(DEGs)encoding proteins in cyp71 mutant.Further functional analysis revealed that some significantly downregulated DEGs,such as AP3,ASK1 and FLOR1,were involved in petals and stamen development.Notably,other significantly downregulated DEGs,including GRP18,GRP 19,GRP20,EXL4,EXL6 etc,were regulated by DELLA protein(RGA)and were also involved in stamen development.Besides that,we also detected 36 IncRNAs that significantly differentially expressed in cyp71 mutant,compared with wild type.Additionally,12 differentially expressed lncRNAs were predicted to have 20 cis-regulated target genes,among which there were some pollen specific expressed genes such as ASK1,EXL4 and EXL6.Via sRNA high throughput sequencing,a total of 297 known miRNAs and 82 novel miRNAs were detected.Among them,74 miRNAs were differentially expressed between WT and cyp71 mutant.Of these,45 miRNAs(39 known and 6 novel)were obviously downregulated in the floral bud of cyp71 mutant.Additionally,843 transcripts targeted by 166 miRNAs were detected through degradome sequencing and analysis.Moreover,we found 113 transcripts targeted by 32 differentially expressed miRNAs among 843 transcripts,in which several genes,such as SPL2,SPL9,SPL15,ARF8 and ARF17,play key roles in flower development.In sum,these present studies demonstrated that loss of AtCYP71 function lead to significantly differential expression of many genes encoding proteins and ncRNAs,respectively,in floral buds and seedlings,and suggested that AtCYP71 functions in gene activation and regulates the sepal,stamen,pollen development besides of gene repression during vegetative growth stage,and might cross talk with GA and JA signal pathways to regulated stamen development.