Molecular cloning and characterization of five SmGRAS genes associated with tanshinone biosynthesis in Salvia miltiorrhiza hairy roots

PLoS One. 2017 Sep 27;12(9):e0185322. doi: 10.1371/journal.pone.0185322. eCollection 2017.

Abstract

The gibberellin-responsive element binding factor (GRAS) family of proteins plays an important role in the transcriptional regulation of plant development and hormone signaling. To date, there are no reports on GRAS family proteins expressed in Salvia miltiorrhiza. In this study, 28 ESTs that contained the GRAS domain were identified from a S. miltiorrhiza cDNA library. Of these, full-length sequences of five genes were cloned and sequence analysis indicated that all five proteins contain only one GRAS domain and therefore, belong to the GRAS family. The five genes were designated S. miltiorrhiza GRAS1-5 (SmGRAS1-5), which belong to groups I (SmGRAS2 and SmGRAS4), II (SmGRAS3), III (SmGRAS1), and VIII (SmGRAS5) respectively. Additionally, SmGRAS1-5 have different expression patterns in the reed head, stems, leaves, flowers, and roots of S. miltiorrhiza. In this study, the expression of SmGRAS1-5 was sensitive to Gibberellin (GA) stress and that of SmGRAS1, SmGRAS4 and SmGRAS5 was sensitive to Ethephon (Eth) stress respectively. Moreover, S. miltiorrhiza copalyl diphosphate synthases 1 (SmCPS1) and S. miltiorrhiza kaurene synthase like 1 (SmKSL1), which are two key enzymes gene in the diterpenoid biosynthesis pathway, were also response to GA and Eth stress. In addition, Dihydrotanshinone (DT-I) and Tanshinone I (T-I) content were enhanced by GA and Eth stress, Tanshinone IIA (T-IIA) content was increased by GA stress, and the accumulation of Cryptotanshinone (CT) was insensitive to both GA and Eth stress. Together, these results provide insights into functional conservation and diversification of SmGRASs and are useful information for further elucidating SmGRAS functions.

MeSH terms

  • Abietanes / biosynthesis*
  • Amino Acid Motifs
  • Amino Acid Sequence
  • Base Sequence
  • Biosynthetic Pathways / drug effects
  • Biosynthetic Pathways / genetics*
  • Cloning, Molecular
  • Conserved Sequence / genetics
  • Gene Expression Profiling
  • Gene Expression Regulation, Plant / drug effects
  • Genes, Plant*
  • Gibberellins / pharmacology
  • Organ Specificity / drug effects
  • Organ Specificity / genetics
  • Organophosphorus Compounds / pharmacology
  • Phenanthrenes / metabolism
  • Phylogeny
  • Plant Proteins / chemistry
  • Plant Proteins / genetics*
  • Plant Proteins / metabolism
  • Plant Roots / drug effects
  • Plant Roots / genetics*
  • Salvia miltiorrhiza / genetics*

Substances

  • Abietanes
  • Gibberellins
  • Organophosphorus Compounds
  • Phenanthrenes
  • Plant Proteins
  • tanshinone
  • ethephon

Grants and funding

These researches were supported by National Natural Science Foundation of China (81373908) and National "Twelfth Five-Year" Plan for Science & Technology Support (2015BAC01B03).