Abstract
Key message
The novel ZmR1CQ01 allele for maize anthocyanin synthesis was identified, and the biological function and regulatory molecular mechanisms of three ZmR1 alleles were unveiled.
Abstract
Anthocyanins in maize are valuable to human health. The R1 gene family is one of the important regulatory genes for the tissue-specific distribution of anthocyanins. R1 gene allelic variations are abundant and its biological function and regulatory molecular mechanisms are not fully understood. By exploiting genetic mapping and transgenic verification, we found that anthocyanin pigmentation in maize leaf midrib was controlled by ZmR1 on chromosome 10. Allelism test of maize zmr1 EMS mutants confirmed that anthocyanin pigmentation in leaf sheath was also controlled by ZmR1. ZmR1CQ01 was a novel ZmR1 allelic variation obtained from purple maize. Its overexpression caused the whole maize plant to turn purple. ZmR1B73 allele confers anthocyanin accumulation in near ground leaf sheath rather than in leaf midribs. The mRNA expression level of ZmR1B73 was low in leaf midribs, resulting in no anthocyanin accumulation. ZmR1B73 overexpression promoted anthocyanin accumulation in leaf midribs. Loss of exon 5 resulted in ZmR1ZN3 allele function destruction and no anthocyanin accumulation in leaf midrib and leaf sheath. DNA affinity purification sequencing revealed 1010 genes targeted by ZmR1CQ01, including the bz2 in anthocyanin synthesis pathway. RNA-seq analysis showed 55 genes targeted by ZmR1CQ01 changed the expression level significantly, and the expression of genes encoding key enzymes in flavonoid and phenylpropanoid biosynthesis pathways were significantly up-regulated. ZmR1 functional molecular marker was developed. These results revealed the effects of transcriptional regulation and sequence variation on ZmR1 function and identified the genes targeted by ZmR1CQ01 at the genome-wide level.
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Data availability
The BSR-seq raw data were deposited in the NCBI Sequence Read Archive (SRA) database (accession number: PRJNA657510). The RNA-seq raw data of leaf midribs of CQ01 and B73 were deposited in the NCBI SRA database (accession number: PRJNA657510). The RNA-seq raw data of leaves of wild-type and ZmR1CQ01-overexpressing transgenic Jing724 were deposited in the NCBI SRA database (accession number: PRJNA792089). The DAP-seq raw data were deposited in the NCBI SRA database [accession number: PRJNA792070 (SAMN26185953)]. The DNA and transcript sequences of ZmR1 in different maize accessions have been deposited in the NCBI GenBank database under the following accession numbers: ON256208 (ZmR1B73), ON256209 (ZmR1CQ01), ON256210 (ZmR1ZN3).
References
Aoki H, Kuze N, Kato Y (2002) Anthocyanin isolated from purple corn (Zea mays L.). Foods Food Ingred J Jpn 199:41–45
Bolger AM, Lohse M, Usadel B (2014) Trimmomatic: a flexible trimmer for Illumina sequence data. Bioinformatics 30:2114–2120
Cone KC, Burr FA, Burr B (1987) Molecular analysis of the maize anthocyanin regulatory locus C1. Proc Natl Acad Sci 83:9631–9635
Cone KC, Cocciolone SM, Burr FA, Burr B (1993) Maize anthocyanin regulatory gene pl 1s a duplicate of c1 that functions in the plant. Plant Cell 5:1795–1805
Consonni G, Viotti A, Dellaporta SL, Tonelli C (1992) cDNA nucleotide sequence of Sn, a regulatory gene in maize. Nucleic Acids Res 20:373
Dellaporta SL, Greenblatt I, Kermicle JL, Hicks JB, Wessler SR (1988) Molecular cloning of the maize R-nj allele by transposon tagging with Ac. Chromosome Struct Funct 263–282
Flachowsky H, Szankowski I, Fischer TC, Richter K, Peil A, Hofer M, Dorschel C, Schmoock S, Gau AE, Halbwirth H, Hanke MV (2010) Transgenic apple plants overexpressing the Lc gene of maize show an altered growth habit and increased resistance to apple scab and fire blight. Planta 231:623–635
Heinz S, Benner C, Spann N, Bertolino E, Lin YC, Laslo P, Cheng JX, Murre C, Singh H, Glass GK (2010) Simple combinations of lineage-determining transcription factors prime cis-regulatory elements required for macrophage and B cell identities. Mol Cell 38:576–589
Hernandez JM, Feller A, Morohashi K, Frame K, Grotewold E (2007) The basic helix–loop– helix domain of maize R links transcriptional regulation and histone modifications by recruitment of an EMSY-related factor. In: Proceedings of the National Academy of Sciences USA 104:17222–17227
Hosoda K, Eruden B, Matsuyama H, Shioya S (2012) Effect of anthocyanin-rich corn silage on digestibility, milk production and plasma enzyme activities in lactating dairy cows. Anim Sci J 83:453–459
Hu C, Li Q, Shen X, Quan S, Lin H, Duan L, Wang Y, Luo Q, Qu G, Han Q, Lu Y, Zhang D, Yuan Z, Shi J (2016a) Characterization of factors underlying the metabolic shifts in developing kernels of colored maize. Sci Rep 6:35479
Hu DG, Sun CH, Zhang QY, An JP, Hao YJ (2016b) Glucose sensor MdHXK1 phosphorylates and stabilizes MdbHLH3 to promote anthocyanin biosynthesis in apple. PLoS Genet 12:e1006273
Jing P, Noriega V, Schwartz SJ, Giusti MM (2007) Effects of growing conditions on purple corncob (Zea mays L.) anthocyanins. J Agric Food Chem 55:8625–8629
Kim D, Langmead B, Salzberg SL (2015) HISAT: a fast spliced aligner with low memory requirements. Nat Methods 12:357–360
Kong Q, Pattanaik S, Feller A, Werkman JR, Chai C, Wang Y, Grotewold E, Yuan L (2012) Regulatory switch enforced by basic helix-loop-helix and ACT-domain mediated dimerizations of the maize transcription factor R. In: Proceedings of the National Academy of Sciences USA 109:E2091–E2097
Li H, Flachowsky H, Fischer TC, Hanke MV, Forkmann G, Treutter D, Schwab W, Hoffmann T, Szankowski I (2007) Maize Lc transcription factor enhances biosynthesis of anthocyanins, distinct proanthocyanidins and phenylpropanoids in apple (Malus domestica Borkh.). Planta 226:1243–1254
Li P, Du C, Zhang Y, Yin S, Zhang E, Fang H, Lin D, Xu C, Yang Z (2018) Combined bulked segregant sequencing and traditional linkage analysis for identification of candidate gene for purple leaf sheath in maize. PLoS ONE 13:e0190670
Liu S, Yeh CT, Tang HM, Nettleton D, Schnable PS (2012) Gene mapping via bulked segregant RNA-seq (BSR-seq). PLoS ONE 7:e36406
Lopes-Da-Silva F, Pascual-Teresa SD, Rivas-Gonzalo J, Santos-Buelga C (2002) Identification of anthocyanin pigments in strawberry (cv Camarosa) by LC using DAD and ESI-MS detection. Eur Food Res Technol 214:248–253
Love MI, Huber W, Anders S (2014) Moderated estimation of fold change and dispersion for rna-seq data with deseq2 moderated estimation of fold change and dispersion for RNA-seq data with DESeq2. Genome Biol 15:550
Ludwig SR, Habera LF, Dellaporta SL, Wessler SR (1989) Lc, a member of the maize R gene family responsible for tissue-specific anthocyanin production, encodes a protein similar to transcriptional activators and contains the myc-homology region. In: Proceedings of the National Academy of Sciences USA 86:7092–7096
Luo M, Shi Y, Yang Y, Zhao Y, Zhao J (2020) Sequence polymorphism of the waxy gene in waxy maize accessions and characterization of a new waxy allele. Sci Rep 10:15851
Luo MJ, Zhang YX, Li JN, Zhang PP, Chen K, Song W, Wang XQ, Yang JX, Lu XD, Lu BS, Zhao YX, Zhao JR (2021) Molecular dissection of maize seedling salt tolerance using a genome-wide association analysis method. Plant Biotechnol J 19:1937–1951
Machanick P, Bailey TL (2011) MEME-ChIP: motif analysis of large DNA datasets. Bioinformatics 27:1696–1897
Matsui K, Umemura Y, Ohme-Takagi M (2008) AtMYBL2, a protein with a single MYB domain, acts as a negative regulator of anthocyanin biosynthesis in Arabidopsis. Plant J 55:954–967
Mckenna A, Hanna M, Banks E, Sivachenko A, Cibulskis K, Kernytsky A, Garimella K, Altshuler D, Gabriel S, Daly M, DePristo MA (2010) The Genome Analysis Toolkit: A MapReduce framework for analyzing next-generation DNA sequencing data. Genome Res 20:1297–1303
Mendel G (1866) Versuche über Pflanzenhybriden. Verhandlungen des naturforschenden Vereines in Brünn 4:3–47
Oshima M, Taniguchi Y, Akasaka M, Abe K, Ichikawa H, Tabei Y, Tanaka J (2019) Development of a visible marker trait based on leaf sheath-specific anthocyanin pigmentation applicable to various genotypes in rice. Breed Sci 69:244–254
Patra B, Pattanaik S, Yuan L (2013) Ubiquitin protein ligase 3 mediates the proteasomal degradation of GLABROUS 3 and ENHANCER OF GLABROUS 3, regulators of trichome development and flavonoid biosynthesis in Arabidopsis. Plant J 74:435–447
Paulsmeyer MN, Brown PJ, Juvik JA (2018) Discovery of anthocyanin acyltransferase1 (AAT1) in maize using genotyping-by-sequencing (GBS). G3 (Bethesda) 8:3669–3678
Petroni K, Pilu R, Tonelli C (2014) Anthocyanins in corn: a wealth of genes for human health. Planta 240:901–911
Procissi A, Dolfini S, Ronchi A, Tonelli C (1997) Light-dependent spatial and temporal expression of regulatory genes in developing maize seeds. Plant Cell 9:1547–1557
Riaz B, Chen H, Wang J, Du L, Wang K, Ye X (2019) Overexpression of maize ZmC1 and ZmR transcription factors in wheat regulates anthocyanin biosynthesis in a tissue-specific manner. Int J Mol Sci 20:5806
Sharma M, Cortes-Cruz M, Ahern KR, McMullen M, Brutnell TP, Chopra S (2011) Identification of the pr1 gene product completes the anthocyanin biosynthesis pathway of maize. Genetics 188:69–79
Shindo M, Kasai T, Abe A, Kondo Y (2007) Effects of dietary administration of plant-derived anthocyanin-rich colors to spontaneously hypertensive rats. J Nutr Sci Vitaminol 53:90–93
Sun X, Zhang Q, Zhou H (2021) Anthocyanins: from biosynthesis regulation to crop improvement. Botany Letters 168:546–557
Tang B, Luo M, Zhang Y, Guo H, Li J, Song W, Zhang R, Feng Z, Kong M, Li H, Cao Z, Lu X, Li D, Zhang J, Wang R, Wang Y, Chen Z, Zhao Y, Zhao J (2021) Natural variations in the P-type ATPase heavy metal transporter gene ZmHMA3 control cadmium accumulation in maize grains. J Exp Bot 72:6230–6246
Tena N, Martín J, Asuerol AG (2020) State of the art of anthocyanins: antioxidant activity, sources, bioavailability, and therapeutic effect in human health. Antioxidants (basel) 9:451
Tonelli C, Consonni G, Dolfini SF, Dellaporta SL, Viotti A, Gavazzi G (1991) Genetic and molecular analysis of Sn, a light-inducible, tissue specific regulatory gene in maize. Mol Gen Genet 225:401–410
Tonelli C, Dolfini S, Ronchi A, Consonni G, Gavazzi G (1994) Light inducibility and tissue specificity of the R gene family in maize. Genetica 94:225–234
Vasimuddin M, Misra S, Li H, Aluru S (2019) Efficient architecture-aware acceleration of BWA-MEM for multicore systems. In: IEEE International Parallel and Distributed Processing Symposium (IPDPS):314–324
Walker EL, Panavas T (2001) Structural features and methylation patterns associated with paramutation at the r1 locus of Zea mays. Genetics 159:1201–1215
Wu B, Chang H, Marini R, Chopra S, Reddivari L (2021) Characterization of maize near-isogenic lines with enhanced flavonoid expression to be used as tools in diet-health complexity. Front Plant Sci 11:619598
Wu TD, Reeder J, Lawrence M, Becker G, Brauer MJ (2016) GMAP and GSNAP for genomic sequence alignment: enhancements to speed, accuracy, and functionality. Methods Mol Biol 1418:283–334
Xie C, Mao X, Huang J, Ding Y, Wu J, Dong S, Kong L, Gao G, Li CY, Wei L (2011) KOBAS 2.0: a web server for annotation and identification of enriched pathways and diseases. Nucleic Acids Res 39:316–322
Yonemaru JI, Miki K, Choi S, Kiyosawa A, Goto K (2018) A genomic region harboring the Pl1 allele from the Peruvian cultivar JC072A confers purple cob on Japanese flint corn (Zea mays L.). Breed Sci 68:582–586
Young MD, Wakefield MJ, Smyth GK, Oshlack A (2010) Gene ontology analysis for RNA-seq: accounting for selection bias. Genome Biol 11:R14
Zhang B, Chopra D, Schrader A, Hulskamp M (2019) Evolutionary comparison of competitive protein-complex formation of MYB, bHLH, and WDR proteins in plants. J Exp Bot 70:3197–3209
Zhang B, Hulskamp M (2019) Evolutionary analysis of MBW Function by phenotypic rescue in Arabidopsis thaliana. Front Plant Sci 10:375
Zhang D, Wu S, An X, Xie K, Dong Z, Zhou Y, Xu L, Fang W, Liu S, Liu S, Zhu T, Li J, Rao L, Zhao J, Wan X (2018) Construction of a multicontrol sterility system for a maize male-sterile line and hybrid seed production based on the ZmMs7 gene encoding a PHD-finger transcription factor. Plant Biotechnol J 16:459–471
Zhang Y, Liu T, Meyer CA, Eeckhoute J, Johnson DS, Bernstein BE, Nusbaum C, Myers RM, Brown M, Li W, Liu XS (2008) Model-based analysis of ChIP-Seq (MACS). Genome Biol 9:R137
Zhu Q, Yu S, Zeng D, Liu H, Wang H, Yang Z, Xie X, Shen R, Tan J, Li H, Zhao X, Zhang Q, Chen Y, Guo J, Chen L, Liu YG (2017) Development of “purple endosperm rice” by engineering anthocyanin biosynthesis in the endosperm with a high-efficiency transgene stacking system. Mol Plant 10:918–929
Funding
This work was supported by the Special Program for Innovation of Beijing Academy of Agriculture and Forestry Sciences (KJCX201907-2), the Youth Research Fund of Beijing Academy of Agriculture and Forestry Sciences (QNJJ202028), the Construction of Collaborative Innovation Center of Beijing Academy of Agricultural and Forestry Sciences (Collaborative Innovation Center of Crop Phenomics, KJCX201917), the Major Scientific and Technological Achievements Cultivation Project of Beijing Academy of Agriculture and Forestry Sciences (MSTA202101), and the Post Expert of Crop Innovation Team of Beijing Industrial Technology System (PECI202201).
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LM, ZJ and WR designed the experiments. LM, LB, SY (Yaxing Shi), WZ, LH, SY (Yamin Shi) and YJ performed the experiments and collected data. LM, SW, ZY, LB, SY (Yaxing Shi), ZC, WY, LX, FY and XL analyzed the data. LM wrote the manuscript. All authors reviewed the manuscript.
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Luo, M., Lu, B., Shi, Y. et al. A newly characterized allele of ZmR1 increases anthocyanin content in whole maize plant and the regulation mechanism of different ZmR1 alleles. Theor Appl Genet 135, 3039–3055 (2022). https://doi.org/10.1007/s00122-022-04166-0
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DOI: https://doi.org/10.1007/s00122-022-04166-0