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Structural features of GmIRCHS, candidate of the I gene inhibiting seed coat pigmentation in soybean: implications for inducing endogenous RNA silencing of chalcone synthase genes

Plant Molecular Biology volume 64, pages 467–479 (2007)Cite this article

Abstract

Most commercial soybean varieties have yellow seeds due to loss of pigmentation in the seed coat. The I gene inhibits pigmentation over the entire seed coat, resulting in a uniform yellow color of mature harvested seeds. We previously demonstrated that the inhibition of seed coat pigmentation by the I gene results from post-transcriptional gene silencing (PTGS) of chalcone synthase (CHS) genes. Little is known about the structure of the I gene and the mechanism by which it induces PTGS of CHS genes. Here, we report a candidate of the I gene, GmIRCHS, which consists of a 5′-portion of a DnaJ-like gene containing a promoter region and a perfect inverted repeat (IR) of 1.1-kb truncated CHS3 sequences (5′-ΔCHS3 and 3′-ΔCHS3). RT-PCRs and RNase protection assay indicated the existence of the read-through product from 5′-ΔCHS3 to 3′-ΔCHS3 and the dsRNA region of ΔCHS3, suggesting that dsRNA of ΔCHS3 could be transcribed from GmIRCHS and could induce PTGS of CHS genes. Moreover, the IR structure of ΔCHS3 in GmIRCHS was lost in the soybean mutants in which I was changed to i, supporting the conclusion that GmIRCHS is the I gene.

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Abbreviations

CHS:

Chalcone synthase

dsRNA:

Double-stranded RNA

IR:

Inverted repeat

ORF:

Open reading frame

PTGS:

Post-transcriptional gene silencing

RACE:

Rapid amplification of cDNA ends

RDR:

RNA-directed RNA polymerase

RFLP:

Restriction fragment length polymorphism

RNAi:

RNA interference

RT:

Reverse transcription

siRNA:

Short interfering RNA

UTR:

Untranslated region

References

  • Akada S, Dube SK (1995) Organization of soybean chalcone synthase gene clusters and characterization of a new member of the family. Plant Mol Biol 29:189–199

    PubMed  Article  CAS  Google Scholar 

  • Akada S, Kung SD, Dube SK (1990) The nucleotide sequence of gene 3 of the soybean chalcone synthase multigene family. Nucleic Acids Res 18:5899

    PubMed  Article  CAS  Google Scholar 

  • Akada S, Kung SD, Dube SK (1993) Nucleotide sequence and putative regulatory elements of a nodule-development-specific member of the soybean (Glycine max) chalcone synthase multigene family, Gmchs7. Plant Physiol 102:321–323

    PubMed  Article  CAS  Google Scholar 

  • Baulcombe D (2004) RNA silencing in plants. Nature 431:356–363

    PubMed  Article  CAS  Google Scholar 

  • Bernard RL, Weiss MG (1973) Qualitative genetics. In: Caldwell BE (ed) Soybean: improvement, production, and uses. American Society of Agronomy, Madison, pp 117–154

    Google Scholar 

  • Cheetham ME, Caplan AJ (1998) Structure, function and evolution of DnaJ: conservation and adaptation of chaperone function. Cell Stress & Chaperones 3:28–36

    Article  CAS  Google Scholar 

  • Chuang CF, Meyerowitz EM (2000) Specific and heritable genetic interference by double-stranded RNA in Arabidopsis thaliana. Proc Natl Acad Sci USA 97:4985–4990

    PubMed  Article  CAS  Google Scholar 

  • Clough SJ, Tuteja JH, Li M, Marek LF, Shoemaker RC, Vodkin LO (2004) Features of a 103-kb gene-rich region in soybean include an inverted perfect repeat cluster of CHS genes comprising the I locus. Genome 47:819–831

    PubMed  Article  CAS  Google Scholar 

  • De Buck S, Jacobs A, Van Montagu M, Depicker A (1999) The DNA sequences of T-DNA junctions suggest that complex T-DNA loci are formed by a recombination process resembling T-DNA integration. Plant J 20:295–304

    PubMed  Google Scholar 

  • Depicker A, Sanders M, Meyer P (2005) Transgene silencing. In: Meyer P (ed) Plant epigenetics. Blackwell, Oxford, pp 1–32

    Google Scholar 

  • Du Q, Thonberg H, Wang J, Wahlestedt C, Liang Z (2005) A systematic analysis of the silencing effects of an active siRNA at all single-nucleotide mismatched target sites. Nucleic Acids Res 33:1671–1677

    PubMed  Article  CAS  Google Scholar 

  • Elbashir SM, Martinez J, Patkaniowska A, Lendeckel W, Tuschl T (2001) Functional anatomy of siRNAs for mediating efficient RNAi in Drosophila melanogaster embryo lysate. EMBO J 20:6877–6888

    PubMed  Article  CAS  Google Scholar 

  • Hamilton AJ, Voinnet O, Chappell L, Baulcombe DC (2002) Two classes of short interfering RNA in RNA silencing. EMBO J 21:4671–4679

    PubMed  Article  CAS  Google Scholar 

  • Kasai A, Watarai M, Yumoto S, Akada S, Ishikawa R, Harada T, Niizeki M, Senda M (2004) Influence of PTGS on chalcone synthase gene family in yellow soybean seed coat. Breed Sci 54:355–360

    Article  CAS  Google Scholar 

  • Kusaba M, Miyahara K, Iida S, Fukuoka H, Takano T, Sassa H, Nishimura M, Nishio T (2003) Low glutelin content 1: a dominant mutation that suppresses the glutelin multigene family via RNA silencing in rice. Plant Cell 15:1455–1467

    PubMed  Article  CAS  Google Scholar 

  • Marchuck D, Drumm M, Saulino A, Collins FS (1991) Construction of T-vectors, a rapid and general system for direct cloning of unmodified PCR products. Nucleic Acids Res 19:1154

    Article  Google Scholar 

  • Matsumura H, Watanabe S, Harada K, Senda M, Akada S, Kawasaki S, Dubouzet EG, Minaka N, Takahashi R (2005) Molecular linkage mapping and phylogeny of the chalcone synthase multigene family in soybean. Theor Appl Genet 110:1203–1209

    PubMed  Article  CAS  Google Scholar 

  • Melquist S, Bender J (2003) Transcription from an upstream promoter controls methylation signaling from an inverted repeat of endogenous genes in Arabidopsis. Genes Dev 17:2036–2047

    PubMed  Article  CAS  Google Scholar 

  • Miernyk JA (2001) The J-domain proteins of Arabidopsis thaliana: an unexpectedly large and diverse family of chaperones. Cell Stress & Chaperones 6:209–218

    PubMed  Article  CAS  Google Scholar 

  • Miernyk JA, Coop NE (2000) AtJ20 (Accession No. AF214107), a plastid-localized type III J-domain protein from Arabidopsis thaliana. (PGR00–027). Plant Physiol 122(2):620

    Google Scholar 

  • Miki D, Shimamoto K (2004) Simple RNAi vectors for stable and transient suppression of gene function in rice. Plant Cell Physiol 45:445–450

    Article  Google Scholar 

  • Nagai I (1921) A genetico-physiological study on the formation of anthocyanin and brown pigments in plants. Tokyo Univ Coll of Agr J 8:1–92

    CAS  Google Scholar 

  • Nagai I, Saito S (1923) Linked factors in soybean. Japan J Bot 1:121–136

    Google Scholar 

  • Owen FV (1928) Inheritance studies in soybeans. III. Seed coat color and summary of all other mendelian characters thus far reported. Genetics 13:50–79

    PubMed  CAS  Google Scholar 

  • Richards E (1987) Preparation of genomic DNA from plant tissue. In: Ausubel FM, Brent R, Kingston RE, Moore DD, Seidman JG, Smith JA, Struhl K (eds) Current protocols in molecular biology. Wiley, New York, pp 2.3.1–2.3.3

    Google Scholar 

  • Senda M, Jumonji A, Yumoto S, Ishikawa R, Harada T, Niizeki M, Akada S (2002a) Analysis of the duplicated CHS1 gene related to the suppression of the seed coat pigmentation in yellow soybeans. Theor Appl Genet 104:1086–1091

    PubMed  Article  CAS  Google Scholar 

  • Senda M, Kasai A, Yumoto S, Akada S, Ishikawa R, Harada T, Niizeki M (2002b) Sequence divergence at chalcone synthase gene in pigmented seed coat soybean mutants of the Inhibitor locus. Genes Genet Syst 77:341–350

    PubMed  Article  CAS  Google Scholar 

  • Senda M, Masuta C, Ohnishi S, Goto K, Kasai A, Sano T, Hong JS, MacFarlane S (2004) Patterning of virus-infected Glycine max seed coat is associated with suppression of endogenous silencing of chalcone synthase genes. Plant Cell 16:807–818

    PubMed  Article  CAS  Google Scholar 

  • Shah DM, Hightower RC, Meagher RB (1982) Complete nucleotide sequence of a soybean actin gene. Proc Natl Acad Sci USA 79:1022–1026

    PubMed  Article  CAS  Google Scholar 

  • Sijen T, Vijn I, Rebocho A, van Blokland R, Roelofs D, Mol JNM, Kooter JM (2001) Transcriptional and posttranscriptional gene silencing are mechanistically related. Curr Biol 11:436–440

    PubMed  Article  CAS  Google Scholar 

  • Smith NA, Singh SP, Wang M-B, Stoutjesdijk P, Green A, Waterhouse PM (2000) Total silencing by intron-spliced hairpin RNAs. Nature 407:319–320

    PubMed  Article  CAS  Google Scholar 

  • Todd JJ, Vodkin LO (1993) Pigmented soybean (Glycine max) seed coats accumulate proanthocyanidins during development. Plant Physiol 102:663–670

    PubMed  CAS  Google Scholar 

  • Todd JJ, Vodkin LO (1996) Duplications that suppress and deletions that restore expression from a chalcone synthase multigene family. Plant Cell 8:687–699

    PubMed  Article  CAS  Google Scholar 

  • Tuteja JH, Clough SJ, Chan W-C, Vodkin LO (2004) Tissue-specific gene silencing mediated by a naturally occurring chalcone synthase gene cluster in Glycine max. Plant Cell 16:819–835

    PubMed  Article  CAS  Google Scholar 

  • Vance V, Vaucheret H (2001) RNA silencing in plants- defense and counterdefense. Science 292:2277–2280

    PubMed  Article  CAS  Google Scholar 

  • Voinnet O (2001) RNA silencing as a plant immune system against viruses. Trends Genet 17:449–459

    PubMed  Article  CAS  Google Scholar 

  • Voinnet O (2005) Induction and suppression of RNA silencing: insights from viral infections. Nat Rev Genet 6:206–220

    PubMed  Article  CAS  Google Scholar 

  • Wang CS, Todd JJ, Vodkin LO (1994) Chalcone synthase mRNA and activity are reduced in yellow soybean seed coats with dominant I alleles. Plant Physiol 105:739–748

    PubMed  Article  CAS  Google Scholar 

  • Wang CS, Vodkin LO (1994) Extraction of RNA from tissues containing high levels of procyanidins that bind RNA. Plant Mol Biol Rep 12:132–145

    CAS  Google Scholar 

  • Waterhouse PM, Wang MB, Lough T (2001) Gene silencing as an adaptive defence against viruses. Nature 411:834–842

    PubMed  Article  CAS  Google Scholar 

  • Wesley SV, Helliwell CA, Smith NA, Wang MB, Rouse DT, Liu Q, Gooding PS, Singh SP, Abbott D, Stoutjesdijk PA, Robinson SP, Gleave AP, Green AG, Waterhouse PM (2001) Construct design for efficient, effective and high-throughput gene silencing in plants. Plant J 27:581–590

    PubMed  Article  CAS  Google Scholar 

  • Woodworth CM (1933) Genetics of the soybean. J Amer Soc Agron 25:36–51

    Google Scholar 

  • Zabala G, Vodkin L (2003) Cloning of the pleiotropic T locus in soybean and two recessive alleles that differentially affect structure and expression of the encoded flavonoid 3’ hydroxylase. Genetics 163:295–309

    PubMed  CAS  Google Scholar 

Download references

Acknowledgements

This work was supported in part by Grants-in-Aid for Scientific Research from the Ministry of Education, Culture, Sports, Science and Technology, Japan, and from Hirosaki University (Priority Area, 2003–2006), Japan, and a research grant from the Iijima Memorial Foundation for the Promotion of Food Science and Technology, Japan.

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Author notes

  1. Setsuzo Yumoto

    Present address: Soybean Breeding Research Subteam, National Agricultural Research Center for Tohoku Region, Kariwano, Daisen, Akita, 019-2112, Japan

Affiliations

  1. Faculty of Agriculture and Life Science, Hirosaki University, Bunkyo-cho 3, Hirosaki, Aomori, 036-8561, Japan

    Atsushi Kasai & Mineo Senda

  2. Department of Medical Technology, Hirosaki University School of Health Sciences, Hon-cho 66, Hirosaki, Aomori, 036-8564, Japan

    Kosuke Kasai

  3. Hokkaido Prefectural Tokachi Agricultural Experiment Station, S9-2, Shinsei, Memuro-cho, Kasai-gun, Hokkaido, 082-0071, Japan

    Setsuzo Yumoto

Authors
  1. Atsushi Kasai
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  2. Kosuke Kasai
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  4. Mineo Senda
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Correspondence to Mineo Senda.

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Kasai, A., Kasai, K., Yumoto, S. et al. Structural features of GmIRCHS, candidate of the I gene inhibiting seed coat pigmentation in soybean: implications for inducing endogenous RNA silencing of chalcone synthase genes. Plant Mol Biol 64, 467–479 (2007). https://doi.org/10.1007/s11103-007-9169-4

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Keywords

  • CHS genes
  • Endogenous RNA silencing
  • I gene
  • Inverted repeat
  • Seed coat
  • Soybean