Abstract
Sex determination of flowers is a critical component of reproductive biology in plants. The plant hormone ethylene promotes female flower formation in mulberry. Illumina RNA-Seq technology was used for de novo transcriptome assembly to compare the gene expression profiling from six early floral bud stages of mulberry flowers. A set of 87,719 unigenes were generated from 6 different stages in flower bud development. Differential gene expression analysis revealed that the transcriptomes from flower buds of the two early stages (FB2 and FB3) were very different from the remaining. There were 317 genes upregulated in FB2 to FB1. For FB3 and FB4, there were 122 genes upregulated and 328 genes downregulated. We also analyzed several genes and gene families that were previously shown to be involved in floral bud development. Quite a few AP2/ERF genes showed higher expression in both FB2 and FB3. Most of MADS genes showed a gradually increasing expression profile. A gene that belonged to the family of the ethylene biosynthesis genes, 1-aminocyclopropane-1-carboxylate synthase (ACS), showed a rapid rise in FB2, followed by a slow decline. Two genes (MaDREB-A1-1b and MaDREB-A1-1a) were upregulated by ethephon treatment, suggesting a response to ethylene signal during early development of floral bud. The co-expression patterns of genes were also analyzed, along with associated genes previously identified to be important in floral initiation and development. Our dataset provides a rich resource for the analysis of mulberry early floral development.
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References
Aceto S, Gaudio L (2011) The MADS and the beauty: genes involved in the development of orchid flowers. Curr Genomics 12:342
Ahn JH et al (2006) A divergent external loop confers antagonistic activity on floral regulators FT and TFL1. EMBO J 25:605–614
Amasino R (2010) Seasonal and developmental timing of flowering. Plant J 61:1001–1013
Angenent GC, Colombo L (1996) Molecular control of ovule development. Trends Plant Sci 1:228–232
Blázquez MA, Green R, Nilsson O, Sussman MR, Weigel D (1998) Gibberellins promote flowering of Arabidopsis by activating the LEAFY promoter. Plant Cell 10:791–800
Boualem A et al (2008) A conserved mutation in an ethylene biosynthesis enzyme leads to andromonoecy in melons. Science 321:836–838. doi:10.1126/science.1159023
Bowman JL, Smyth DR, Meyerowitz EM (2012) The ABC model of flower development: then and now. Development 139:4095–4098
Conesa A, Götz S, García-Gómez JM, Terol J, Talón M, Robles M (2005) Blast2GO: a universal tool for annotation, visualization and analysis in functional genomics research. Bioinformatics 21:3674–3676
Diggle PK, Stilio VSD, Gschwend AR, Golenberg EM, Moore RC, Russell JRW et al (2011) Multiple developmental processes underlie sex differentiation in angiosperms. Trends Genet 27(9):368–376
Franks RG, Wang C, Levin JZ, Liu Z (2002) SEUSS, a member of a novel family of plant regulatory proteins, represses floral homeotic gene expression with LEUNIG. Development 129:253–263
Grabherr MG et al (2011) Full-length transcriptome assembly from RNA-seq data without a reference genome. Nat Biotechnol 29:644–652
Gu Q, Ferrándiz C, Yanofsky MF, Martienssen R (1998) The FRUITFULL MADS-box gene mediates cell differentiation during Arabidopsis fruit development. Development 125:1509–1517
Guo H, Ecker JR (2003) Plant responses to ethylene gas are mediated by SCFEBF1/EBF2-dependent proteolysis of EIN3 transcription factor. Cell 115:667–677
Hao Y-J et al (2003) DNA damage in the early primordial anther is closely correlated with stamen arrest in the female flower of cucumber (Cucumis sativus L.). Planta 217:888–895
Hepworth SR, Valverde F, Ravenscroft D, Mouradov A, Coupland G (2002) Antagonistic regulation of flowering-time gene SOC1 by CONSTANS and FLC via separate promoter motifs. EMBO J 21:4327–4337
Hepworth SR, Klenz JE, Haughn GW (2006) UFO in the Arabidopsis inflorescence apex is required for floral-meristem identity and bract suppression. Planta 223:769–778. doi:10.1007/s00425-005-0138-3
Kim S, Choi K, Park C, Hwang HJ, Lee I (2006) SUPPRESSOR OF FRIGIDA4, encoding a C2H2-type zinc finger protein, represses flowering by transcriptional activation of Arabidopsis FLOWERING LOCUS C. Plant Cell 18:2985–2998. doi:10.1105/tpc.106.045179
Klintenäs M, Pin PA, Benlloch R, Ingvarsson PK, Nilsson O (2012) Analysis of conifer flowering locus T/terminal flower1-like genes provides evidence for dramatic biochemical evolution in the angiosperm FT lineage. New Phytol 196:1260–1273
Krizek BA, Fletcher JC (2005) Molecular mechanisms of flower development: an armchair guide. Nat Rev Genet 6:688–698
Langfelder P, Horvath S (2008) WGCNA: an R package for weighted correlation network analysis. BMC bioinformatics 9:559
Lee J, Lee I (2010) Regulation and function of SOC1, a flowering pathway integrator. J Exp Bot 61:2247–2254
Lee J, Oh M, Park H, Lee I (2008) SOC1 translocated to the nucleus by interaction with AGL24 directly regulates leafy. Plant J 55:832–843. doi:10.1111/j.1365-313X.2008.03552.x
Li B, Dewey CN (2011) RSEM: accurate transcript quantification from RNA-seq data with or without a reference genome. BMC bioinformatics 12:1
Li Z et al (2009) Molecular isolation of the M gene suggests that a conserved-residue conversion induces the formation of bisexual flowers in cucumber plants. Genetics 182:1381–1385. doi:10.1534/genetics.109.104737
Liu Z, Mara C (2010) Regulatory mechanisms for floral homeotic gene expression. In: Seminars in cell & developmental biology, vol 1. Elsevier, p 80–86
Liu C et al (2008) Direct interaction of AGL24 and SOC1 integrates flowering signals in Arabidopsis. Development 135:1481–1491
Masiero S, Colombo L, Grini PE, Schnittger A, Kater MM (2011) The emerging importance of type I MADS box transcription factors for plant reproduction. Plant Cell 23:865–872
Michaels S, Amasino R (2000) Memories of winter: vernalization and the competence to flower. Plant Cell Environ 23:1145–1153
Michaels SD, Ditta G, Gustafson-Brown C, Pelaz S, Yanofsky M, Amasino RM (2003) AGL24 acts as a promoter of flowering in Arabidopsis and is positively regulated by vernalization. Plant J 33:867–874
Moriya Y, Itoh M, Okuda S, Yoshizawa AC, Kanehisa M (2007) KAAS: an automatic genome annotation and pathway reconstruction server. Nucleic Acids Res 35:W182–W185
Nakano T, Suzuki K, Fujimura T, Shinshi H (2006) Genome-wide analysis of the ERF gene family in Arabidopsis and rice. Plant Physiol 140:411–432
Nam J, Ma H, Nei M (2003) Antiquity and evolution of the MADS-box gene family controlling flower development in plants. Mol Biol Evol 20:1435–1447
Ohme-Takagi M, Shinshi H (1995) Ethylene-inducible DNA binding proteins that interact with an ethylene-responsive element. Plant Cell 7:173–182
Parcy F (2005) Flowering: a time for integration. Int J Dev Biol 49:585
Pařenicová L et al (2003) Molecular and phylogenetic analyses of the complete MADS-box transcription factor family in Arabidopsis. Plant Cell 15:1538–1551
Pelaz S, Ditta GS, Baumann E, Wisman E, Yanofsky MF (2000) B and C floral organ identity functions require SEPALLATA MADS-box genes. Nature 405:200–203
Pelaz S, Gustafson-Brown C, Kohalmi SE, Crosby WL, Yanofsky MF (2001) APETALA1 and SEPALLATA3 interact to promote flower development. Plant J 26:385–394
Pin P, Nilsson O (2012) The multifaceted roles of FLOWERING LOCUS T in plant development. Plant Cell Environ 35:1742–1755
Posé D, Yant L, Schmid M (2012) The end of innocence: flowering networks explode in complexity. Curr Opin Plant Biol 15:45–50. doi:10.1016/j.pbi.2011.09.002
Potuschak T, Lechner E, Parmentier Y, Yanagisawa S, Grava S, Koncz C, Genschik P (2003) EIN3-dependent regulation of plant ethylene hormone signaling by two Arabidopsis F box proteins: EBF1 and EBF2. Cell 115:679–689
Rasmussen M, Karypis G (2004) gcluto: an interactive clustering, visualization, and analysis system. UMN-CS TR-04 21
Rounsley SD, Ditta GS, Yanofsky MF (1995) Diverse roles for MADS box genes in Arabidopsis development. Plant Cell 7:1259–1269
Rowell P, Miller DG (1971) Induction of male sterility in wheat with 2-chloroethylphosphonic acid. (ethrel). Crop Sci 11:629–631
Rutitzky M, Ghiglione HO, Cura JA, Casal JJ, Yanovsky MJ (2009) Comparative genomic analysis of light-regulated transcripts in the Solanaceae. BMC Genomics 10:60. doi:10.1186/1471-2164-10-60
Saito T, Takahashi H (1987) Role of leaves in ethylene-induced femaleness of cucumber plants. J Jpn Soc Hortic Sci
Saito S et al (2007) Correlation between development of female flower buds and expression of the CS-ACS2 gene in cucumber plants. J Exp Bot 58:2897–2907. doi:10.1093/jxb/erm141
Salathia N, Davis SJ, Lynn JR, Michaels SD, Amasino RM, Millar AJ (2006) FLOWERING LOCUS C-dependent and -independent regulation of the circadian clock by the autonomous and vernalization pathways. BMC Plant Biol 6:10. doi:10.1186/1471-2229-6-10
Santner A, Estelle M (2007) The JAZ proteins link jasmonate perception with transcriptional changes. Plant Cell 19:3839–3842. doi:10.1105/tpc.107.056960
Shang J, Song P, Ma B, Qi X, Zeng Q, Xiang Z, He N (2014) Identification of the mulberry genes involved in ethylene biosynthesis and signaling pathways and the expression of MaERF-B2-1 and MaERF-B2-2 in the response to flooding stress. Funct Integr Genomics 14:767–777
Shoukang L, Zhiqiang L, Yunxiang W (1993) Effect of gibberellin and ethrel on the sex of mulberry flower. Bulletin of Sericulture (China)
Tamura K, Peterson D, Peterson N, Stecher G, Nei M, Kumar S (2011) MEGA5: Molecular Evolutionary Genetics Analysis using maximum likelihood, evolutionary distance, and maximum parsimony methods. Mol Biol Evol 28:2731–2739
Theißen G (2001) Development of floral organ identity: stories from the MADS house. Curr Opin Plant Biol 4:75–85
Thomas TD (2004) In vitro modification of sex expression in mulberry (Morus alba) by ethrel and silver nitrate. Plant Cell Tissue Organ Cult 77:277–281
Thompson BE, Hake S (2009) Translational biology: from Arabidopsis flowers to grass inflorescence architecture. Plant Physiol 149:38–45
Tilly JJ, Allen DW, Jack T (1998) The CArG boxes in the promoter of the Arabidopsis floral organ identity gene APETALA3 mediate diverse regulatory effects. Development 125:1647–1657
Trapnell C et al (2010) Transcript assembly and quantification by RNA-seq reveals unannotated transcripts and isoform switching during cell differentiation. Nat Biotechnol 28:511–515
Trebitsh T, Staub JE, O’Neill SD (1997) Identification of a 1-aminocyclopropane-1-carboxylic acid synthase gene linked to the female (F) locus that enhances female sex expression in cucumber. Plant Physiol 113:987–995
Untergasser A, Cutcutache I, Koressaar T, Ye J, Faircloth BC, Remm M, Rozen SG (2012) Primer3—new capabilities and interfaces. Nucleic Acids Res 40:e115. doi:10.1093/nar/gks596
Wang J-W, Czech B, Weigel D (2009) miR156-regulated SPL transcription factors define an endogenous flowering pathway in Arabidopsis thaliana. Cell 138:738–749
Wang L, Feng Z, Wang X, Wang X, Zhang X (2010) DEGseq: an R package for identifying differentially expressed genes from RNA-seq data. Bioinformatics 26:136–138. doi:10.1093/bioinformatics/btp612
Weigel D, Alvarez J, Smyth DR, Yanofsky MF, Meyerowitz EM (1992) LEAFY controls floral meristem identity in Arabidopsis. Cell 69:843–859
Wellmer F, Riechmann JL (2010) Gene networks controlling the initiation of flower development. Trends Genet 26:519–527
Wigge PA, Kim MC, Jaeger KE, Busch W, Schmid M, Lohmann JU, Weigel D (2005) Integration of spatial and temporal information during floral induction in Arabidopsis. Science 309:1056–1059
Willits MG, Giovanni M, Prata RT, Kramer CM, De LV, Steffens JC et al (2004) Biofermentation of modified flavonoids: an example of in vivo diversification of secondary metabolites. Phytochemistry 65(1):31
Xie C et al (2011) KOBAS 2.0: a web server for annotation and identification of enriched pathways and diseases. Nucleic Acids Res 39:W316–W322. doi:10.1093/nar/gkr483
Yamasaki K et al (2005) Solution structure of the major DNA-binding domain of Arabidopsis thaliana ethylene-insensitive3-like3. J Mol Biol 348:253–264
Yin T, Quinn JA (1995) Tests of a mechanistic model of one hormone regulating both sexes in Cucumis sativus (Cucurbitaceae). Am J Bot:1537–1546
Zafari M, Malekzadeh A, Jahani R (2015) Effect of plant hormone auxin in flower development. Agriculture Science Developments 2:27–30
Zhang Y, Fan W, Kinkema M, Li X, Dong X (1999) Interaction of NPR1 with basic leucine zipper protein transcription factors that bind sequences required for salicylic acid induction of the PR-1 gene. P Natl Acad Sci Usa 96:6523–6528
Zhang L, Li Z, Quan R, Li G, Wang R, Huang R (2011) An ap2 domain-containing gene, ese1, targeted by the ethylene signaling component ein3 is important for the salt response in arabidopsis. Plant Physiol 157(2):854–865
Zik M (2002) Global identification of target genes regulated by APETALA3 and PISTILLATA floral homeotic gene action. Plant Cell 15:207–222. doi:10.1105/tpc.006353
Acknowledgements
This project was funded by the research grants from the National Hi-Tech Research and Development Program of China (No. 2013AA100605-3), Natural Science Foundation of China (No. 31572323), China Postdoctoral Science Foundation funded projects (No. 2013 M540694 and No. 2014 T70845), and the “111” Project (B12006).
Author’s contribution statement
NH and JS conceived and designed the research. JS prepared the samples and analyzed the data. JS and NH conducted the experiments and wrote the paper. JL and ZX contributed new reagents and analytical tools. NH and JL revised the manuscript. All authors read and approved the manuscript.
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Communicated by M. Wirthensohn
Data archiving statement
The datasets reported in this paper have been deposited as raw reads in the Genbank SRA database (accession number SRR2868676). The BioProject ID related to this paper is PRJNA272494.
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Fig. S1
Expression correlations between flower buds. Abscissa: the value of sample1 was calculated using log10 (FPKM + 1). Ordinate: the value of sample2 was calculated using log10 (FPKM + 1). R 2, Pearson correlation coefficient. FB, flower bud (GIF 32 kb)
Fig. S2
Venn diagram depicting the overlap of DEG genes between two flower buds. The DEGs include both up and down-regulated genes in flower buds. A, the DEGs of between two adjacent time point floral buds. B, the DEGs of between the other time points against FB1. (GIF 16 kb)
Fig. S3
KEGG enriched in DEGs of between two adjacent time point floral buds. Benjamini-Hochberg-adjusted P values (q-value <0.05) are shown. (GIF 12 kb)
Fig. S4
FB1-FB2 DEGs KEGG enrichment in plant hormone signal transduction. Red: up-regulated genes, green: down-regulated genes (GIF 26 kb)
Fig. S5
FB3-FB4 DEGs KEGG enrichment in plant hormone signal transduction. Red: up-regulated genes, green: down-regulated genes (GIF 26 kb)
Fig. S6
Expression patterns of hormone-related genes. For each hormone, its related genes are divided into two functional categories: (1) synthesis-degradation, (2) signal transduction. The FPKM values were used. All the values were converted by log10 (FPKM + 1) and normalized by scale function of R program. (GIF 333 kb)
Fig. S7
The expression of floral initiation genes. A, The current known components and relationships of floral initiation in Arabidopsis. B, Expression patterns of putative mulberry orthologs of A. thaliana floral initiation. The FPKM of floral initiation genes were used. These genes were identified by BLAST method. (GIF 77 kb)
Fig. S8
Co-expression patterns with specific genes. Co-expression analysis of the transcript used the WGCNA package of R. Calculate the correlation between each two genes. According to the correlation, the genes were divided into different expression patterns. The correlation between expression pattern and the expression of gene marked in each small box. P1–27, highly similar expression patterns. AP2/ERF family genes were mainly expressed in the FB2 and FB3. The expression of MADS-boxes genes were gradual increased during floral bud development. (GIF 215 kb)
Fig. S9
KEGG analysis of P19 and P24 module genes. Rich_factor, input number/background number; q value, Benjamini-Hochberg-adjusted P values. (GIF 43 kb)
Fig. S10
Co-expression network comprehensive analysis of P24. The yellow ellipse represent the gene have more than 40 links. The red ellipse represent the genes have between 20 and 40 links. The green ellipse represent the genes have between 10 and 20 links. (GIF 68 kb)
Fig. S11
Verification of differentially expressed genes by qRT-PCR. Relative levels of gene expression by qRT-PCR were normalized against the mulberry ribosomal protein gene Morus024083. Data are represented as mean ± standard error of three replicates. (GIF 23 kb)
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Shang, J., Liang, J., Xiang, Z. et al. Anatomical and transcriptional dynamics of early floral development of mulberry (Morus alba). Tree Genetics & Genomes 13, 40 (2017). https://doi.org/10.1007/s11295-017-1122-3
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DOI: https://doi.org/10.1007/s11295-017-1122-3