Journal of Plant Research

, Volume 132, Issue 5, pp 617–627 | Cite as

BLADE-ON-PETIOLE genes are not involved in the transition from protonema to gametophore in the moss Physcomitrella patens

  • Yuki Hata
  • Satoshi Naramoto
  • Junko KyozukaEmail author
Regular Paper


The timing of the transition between developmental phases is a critical determinant of plant form. In the moss Physcomitrella patens, the transition from protonema to gametophore is a particularly important step as it results in a change from two-dimensional to three-dimensional growth of the plant body. It is well known that this transition is promoted by cytokinin (CK), however, the underlying mechanisms are poorly understood. Previously, it was reported that P. patens orthologs of BLADE-ON-PETIOLE (BOP) genes (PpBOPs) work downstream of CK to promote the transition to gametophore. To further understand the role of PpBOPs in the control of this transition, we performed functional analyses of PpBOP genes. We simultaneously disrupted the function of all three PpBOP genes in P. patens using CRISPR technology, however, no abnormal phenotypes were observed in the triple mutant during either the gametophytic or the sporophytic growth stages. CK treatment did not alter the phase change in the triple mutant. We conclude that PpBOP genes are unnecessary in the control of P. patens development under normal conditions. We propose that BOP genes are not involved in the control of developmental processes in bryophytes and other basal land plants, but may function in physiological processes such as in the defense response.


BLADE-ON-PETIOLE (BOPBryophyte Cytokinin Physcomitrella patens Triple loss-of-function mutant 



We thank Dr. Yuji Hiwatashi (Miyagi University) for providing the P. patens Gransden strain and for teaching us how to culture and transform P. patens. We thank Dr. Emiko Yoro (Rikkyo University) for teaching us sporophyte induction. We also thank Dr. Fabien Nogue (INRA Centre de Versailles-Grignon) for providing the pBNRF, pAct-Cas9 and sgRNA plasmids. This work was supported by JSPS/MEXT Kakenhi grants to JK (22119008, 22247004, 16K14748, 17H06475, 18K19198) and SN (17K17595).

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflicts of interest.

Supplementary material

10265_2019_1132_MOESM1_ESM.pdf (95 mb)
Supplementary material 1 (PDF 97303 kb)


  1. Aoyama T, Hiwatashi Y, Shigyo M et al (2012) AP2-type transcription factors determine stem cell identity in the moss Physcomitrella patens. Development 139:3120–3129. CrossRefPubMedGoogle Scholar
  2. Ashton NW, Cove DJ (1977) The isolation and preliminary characterisation of auxotrophic and analogue resistant mutants of the moss, Physcomitrella patens. MGG Mol Gen Genet 154:87–95. CrossRefGoogle Scholar
  3. Boyle P, Le SuE, Rochon A et al (2009) The BTB/POZ domain of the arabidopsis disease resistance protein NPR1 interacts with the repression domain of TGA2 to negate its function. Plant Cell 21:3700–3713. CrossRefPubMedPubMedCentralGoogle Scholar
  4. Canet JV, Dobón A, Fajmonová J, Tornero P (2012) The BLADE-ON-PETIOLE genes of Arabidopsis are essential for resistance induced by methyl jasmonate. BMC Plant Biol 12:1–13. CrossRefGoogle Scholar
  5. Cho SH, Coruh C, Axtell MJ (2012) miR156 and miR390 regulate tasiRNA accumulation and developmental timing in Physcomitrella patens. Plant Cell 24:4837–4849. CrossRefPubMedPubMedCentralGoogle Scholar
  6. Couzigou J-M, Zhukov V, Mondy S et al (2012) NODULE ROOT and COCHLEATA maintain nodule development and are legume orthologs of arabidopsis BLADE-ON-PETIOLE genes. Plant Cell 24:4498–4510. CrossRefPubMedPubMedCentralGoogle Scholar
  7. Couzigou JM, Magne K, Mondy S et al (2016) The legume NOOT-BOP-COCH-LIKE genes are conserved regulators of abscission, a major agronomical trait in cultivated crops. New Phytol 209:228–240. CrossRefPubMedGoogle Scholar
  8. Cove DJ, Knight CD (1993) The moss Physcomitrella patens, a model system with potential for the study of plant reproduction. Plant Cell 5:1483–1488CrossRefPubMedPubMedCentralGoogle Scholar
  9. Fu ZQ, Yan S, Saleh A et al (2012) NPR3 and NPR4 are receptors for the immune signal salicylic acid in plants. Nature 486:228–232. CrossRefPubMedPubMedCentralGoogle Scholar
  10. Ha CM, Kim G-T, Kim BC et al (2003) The BLADE-ON-PETIOLE 1 gene controls leaf pattern formation through the modulation of meristematic activity in Arabidopsis. Development 130:161–172. CrossRefPubMedGoogle Scholar
  11. Ha CM, Jun JH, Nam HG, Fletcher JC (2004) BLADE-ON-PETIOLE1 encodes a BTB/POZ domain protein required for leaf morphogenesis in Arabidopsis thaliana. Plant Cell Physiol 45:1361–1370. CrossRefPubMedGoogle Scholar
  12. Ichihashi Y, Kawade K, Usami T et al (2011) Key proliferative activity in the junction between the leaf blade and leaf petiole of Arabidopsis. Plant Physiol 157:1151–1162. CrossRefPubMedPubMedCentralGoogle Scholar
  13. Ichihashi Y, Aguilar-Martíneza JA, Farhia M et al (2014) Evolutionary developmental transcriptomics reveals a gene network module regulating interspecific diversity in plant leaf shape. Proc Natl Acad Sci 111:E2616–E2621. CrossRefPubMedGoogle Scholar
  14. Izhaki A, Alvarez JP, Cinnamon Y et al (2018) The tomato BLADE ON PETIOLE and TERMINATING FLOWER regulate leaf axil patterning along the proximal-distal axes. Front Plant Sci 9:1–10. CrossRefGoogle Scholar
  15. Jun JH, Ha CM, Fletcher JC (2010) BLADE-ON-PETIOLE1 coordinates organ determinacy and axial polarity in Arabidopsis by directly activating ASYMMETRIC LEAVES2. Plant Cell 22:62–76. CrossRefPubMedPubMedCentralGoogle Scholar
  16. Khan M, Xu M, Murmu J et al (2012) Antagonistic interaction of BLADE-ON-PETIOLE1 and 2 with BREVIPEDICELLUS and PENNYWISE regulates Arabidopsis inflorescence architecture. Plant Physiol 158:946–960. CrossRefPubMedGoogle Scholar
  17. Khan M, Xu H, Hepworth SR (2014) BLADE-ON-PETIOLE genes: setting boundaries in development and defense. Plant Sci 215–216:157–171. CrossRefPubMedGoogle Scholar
  18. Kofuji R, Hasebe M (2014) Eight types of stem cells in the life cycle of the moss Physcomitrella patens. Curr Opin Plant Biol 17:13–21. CrossRefPubMedGoogle Scholar
  19. Le Bail A, Scholz S, Kost B (2013) Evaluation of reference genes for RT qPCR Analyses of structure-specific and hormone regulated gene expression in Physcomitrella patens gametophytes. PLoS One 8:1–10. CrossRefGoogle Scholar
  20. Lopez-Obando M, Hoffmann B, Géry C et al (2016) Simple and efficient targeting of multiple genes through CRISPR-Cas9 in Physcomitrella patens. G3 6:3647–3653. CrossRefPubMedGoogle Scholar
  21. Magne K, George J, Berbel Tornero A et al (2018) Lotus japonicus NOOT-BOP-COCH-LIKE1 is essential for nodule, nectary, leaf and flower development. Plant J 94:880–894. CrossRefPubMedGoogle Scholar
  22. McKim SM, Stenvik G-E, Butenko MA et al (2008) The BLADE-ON-PETIOLE genes are essential for abscission zone formation in Arabidopsis. Development 135:1537–1546. CrossRefPubMedGoogle Scholar
  23. Moody LA (2019) The 2D to 3D growth transition in the moss Physcomitrella patens. Curr Opin Plant Biol 47:88–95. CrossRefPubMedGoogle Scholar
  24. Moody LA, Kelly S, Rabbinowitsch E, Langdale JA (2018) Genetic regulation of the 2D to 3D growth transition in the moss Physcomitrella patens. Curr Biol 28:473–478.e5. CrossRefPubMedPubMedCentralGoogle Scholar
  25. Nishiyama T, Hiwatashi Y, Sakakibara K et al (2000) Tagged mutagenesis and gene-trap in the moss, Physcomitrella patens by shuttle mutagenesis. DNA Res 7:9–17. CrossRefPubMedGoogle Scholar
  26. Norberg M (2005) The BLADE ON PETIOLE genes act redundantly to control the growth and development of lateral organs. Development 132:2203–2213. CrossRefPubMedGoogle Scholar
  27. Ortiz-Ramírez C, Hernandez-Coronado M, Thamm A et al (2016) A Transcriptome atlas of Physcomitrella patens provides insights into the evolution and development of land plants. Mol Plant 9:205–220. CrossRefPubMedGoogle Scholar
  28. Perroud P-F, Demko V, Johansen W et al (2014) Defective Kernel 1 (DEK1) is required for three-dimensional growth in Physcomitrella patens. New Phytol 203:794–804. CrossRefPubMedPubMedCentralGoogle Scholar
  29. Ponce de León I, Montesano M (2017) Adaptation mechanisms in the evolution of moss defenses to microbes. Front Plant Sci 8:1–14. CrossRefGoogle Scholar
  30. Puttick MN, Morris JL, Williams TA et al (2018) The interrelationships of land plants and the nature of the ancestral embryophyte. Curr Biol 28:733–745.e2. CrossRefPubMedGoogle Scholar
  31. Saleh O, Issman N, Seumel GI et al (2011) MicroRNA534a control of BLADE-ON-PETIOLE 1 and 2 mediates juvenile-to-adult gametophyte transition in Physcomitrella patens. Plant J 65:661–674. CrossRefPubMedGoogle Scholar
  32. Schumaker Karen S, Dietrich MA (1997) Programmed changes in form during moss development. Plant Cell Online 9:1099–1107. CrossRefGoogle Scholar
  33. Strotbek C, Krinninger S, Frank W (2013) The moss Physcomitrella patens: methods and tools from cultivation to targeted analysis of gene function. Int J Dev Biol 57:553–564. CrossRefPubMedGoogle Scholar
  34. Tavakol E, Okagaki R, Verderio G et al (2015) The barley Uniculme4 Gene encodes a BLADE-ON-PETIOLE-like protein that controls tillering and leaf patterning. Plant Physiol 168:164–174. CrossRefPubMedPubMedCentralGoogle Scholar
  35. Toriba T, Tokunaga H, Shiga T et al (2019) BLADE-ON-PETIOLE genes temporally and developmentally regulate the sheath to blade ratio of rice leaves. NAt Commun 10:619CrossRefPubMedPubMedCentralGoogle Scholar
  36. Whitewoods CD, Cammarata J, Venza ZN et al (2018) CLAVATA was a genetic novelty for the morphological innovation of 3D Growth in land plants. Curr Biol 28:2365–2376.e5. CrossRefPubMedPubMedCentralGoogle Scholar
  37. Wickett NJ, Mirarabc S, Nguyenc N et al (2014) Phylotranscriptomic analysis of the origin and early diversification of land plants. Proc Natl Acad Sci 111:E4859–E4868. CrossRefPubMedGoogle Scholar
  38. Winter D, Vinegar B, Nahal H et al (2007) An “electronic fluorescent pictograph” Browser for exploring and analyzing large-scale biological data sets. PLoS One 2:1–12. CrossRefGoogle Scholar
  39. Wu X-M, Yi YuL-BH, Li C-L et al (2012) The tobacco BLADE-ON-PETIOLE2 gene mediates differentiation of the corolla abscission zone by controlling longitudinal cell expansion. Plant Physiol 159:835–850. CrossRefPubMedPubMedCentralGoogle Scholar
  40. Xu M, Hu T, McKim SM et al (2010) Arabidopsis BLADE-ON-PETIOLE1 and 2 promote floral meristem fate and determinacy in a previously undefined pathway targeting APETALA1 and AGAMOUS-LIKE24. Plant J 63:974–989. CrossRefPubMedGoogle Scholar
  41. Xu C, Park SJ, Van Eck J, Lippman ZB (2016) Control of inflorescence architecture in tomato by BTB/POZ transcriptional regulators. Genes Dev 30:2048–2061. CrossRefPubMedPubMedCentralGoogle Scholar
  42. Zhang B, Holmlund M, Lorrain S et al (2017) BLADE-ON-PETIOLE proteins act in an E3 ubiquitin ligase complex to regulate PHYTOCHROME INTERACTING FACTOR 4 abundance. Elife 6:1–19. CrossRefGoogle Scholar

Copyright information

© The Botanical Society of Japan and Springer Japan KK, part of Springer Nature 2019

Authors and Affiliations

  1. 1.Tohoku University Graduate School of Life SciencesSendaiJapan

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