Abstract
Key message
Vegetative-to-reproductive phase transition in female cannabis seedlings occurs autonomously with the de novo development of single flowers.
Abstract
To ensure successful sexual reproduction, many plant species originating from seedlings undergo juvenile-to-adult transition. This phase transition precedes and enables the vegetative-to-reproductive shift in plants, upon perception of internal and/or external signals such as temperature, photoperiod, metabolite levels, and phytohormones. This study demonstrates that the juvenile seedlings of cannabis gradually shift to the adult vegetative stage, as confirmed by the formation of lobed leaves, and upregulation of the phase-transition genes. In the tested cultivar, the switch to the reproductive stage occurs with the development of a pair of single flowers in the 7th node. Histological analysis indicated that transition to the reproductive stage is accomplished by the de novo establishment of new flower meristems which are not present in a vegetative stage, or as dormant meristems at nodes 4 and 6. Moreover, there were dramatic changes in the transcriptomic profile of flowering-related genes among nodes 4, 6, and 7. Downregulation of flowering repressors and an intense increase in the transcription of phase transition-related genes occur in parallel with an increase in the transcription of flowering integrators and meristem identity genes. These results support and provide molecular evidence for previous findings that cannabis possesses an autonomous flowering mechanism and the transition to reproductive phase is controlled in this plant mainly by internal signals.
This is a preview of subscription content, log in via an institution to check access.
Similar content being viewed by others
References
Abe M, Kobayashi Y, Yamamoto S et al (2005) FD, a bZIP protein mediating signals from the floral pathway integrator FT at the shoot apex. Science 309:1052–1056. https://doi.org/10.1126/science.1115983
Abe M, Kosaka S, Shibuta M et al (2019) Transient activity of the florigen complex during the floral transition in Arabidopsis thaliana. Development 146(7):dev171504. https://doi.org/10.1242/dev.171504
Andre CM, Hausman JF, Guerriero G (2016) Cannabis sativa: the plant of the thousand and one molecules. Front Plant Sci. https://doi.org/10.3389/fpls.2016.00019
Anwar N, Ohta M, Yazawa T, Sato Y, Li C, Tagiri A (2018) miR172 downregulates the translation of cleistogamy 1 in barley. Ann Bot 122:251–265. https://doi.org/10.1093/aob/mcy1058
Aukerman MJ, Sakai H (2003) Regulation of flowering time and floral organ identity by a microRNA and its APETALA2-LIKE target genes. Plant Cell 15:2730–2741. https://doi.org/10.1105/tpc.016238
Bassiri A, Irish EE, Poethig RS (1992) Heterochronic effects of teopod 2 on the growth and photosensitivity of the maize shoot. Plant Cell 4:497–504. https://doi.org/10.1105/tpc.4.4.497
Bolger AM, Lohse M, Usadel B (2014) Trimmomatic: a flexible trimmer for Illumina sequence data. Bioinformatics 30:2114–2120. https://doi.org/10.1093/bioinformatics/btu170
Buchfink B, Xie C, Huson DH (2015) Fast and sensitive protein alignment using DIAMOND. Nat Methods 12:59–60. https://doi.org/10.1038/nmeth.3176
Castillejo C, Pelaz S (2008) The balance between CONSTANS and TEMPRANILLO activities determines FT expression to trigger flowering. Curr Biol 18:1338–1343. https://doi.org/10.1016/j.cub.2008.07.075
Chang W, Guo Y, Zhang H et al (2020) Same actor in different stages: genes in shoot apical meristem maintenance and floral meristem determinacy in Arabidopsis. Front Ecol Evol. https://doi.org/10.3389/fevo.2020.00089
Chen X (2004) A microRNA as a translational repressor of APETALA2 in Arabidopsis flower development. Science 303:2022–2025. https://doi.org/10.1126/science.1088060
Chitwood DH, Ranjan A, Kumar R et al (2014) Resolving distinct genetic regulators of tomato leaf shape within a heteroblastic and ontogenetic context. Plant Cell 26:3616–3629. https://doi.org/10.1105/tpc.114.130112
Chung KS, Yoo SY, Yoo SJ et al (2010) BROTHER OF FT AND TFL1 (BFT), a member of the FT/TFL1 family, shows distinct pattern of expression during the vegetative growth of Arabidopsis. Plant Signal Behav 5:1102–1104. https://doi.org/10.4161/psb.5.9.12415
Conesa A, Götz S, García-Gómez JM et al (2005) Blast2GO: a universal tool for annotation, visualization and analysis in functional genomics research. Bioinformatics 21:3674–3676. https://doi.org/10.1093/bioinformatics/bti610
Feng S, Xu Y, Guo C et al (2016) Modulation of miR156 to identify traits associated with vegetative phase change in tobacco (Nicotiana tabacum). J Exp Bot 67:1493–1504. https://doi.org/10.1093/jxb/erv551
Floyd SK, Bowman JL (2007) The ancestral developmental tool kit of land plants. Int J Plant Sci 168:1–35. https://doi.org/10.1086/509079
Freiman A, Shlizerman L, Golobovitch S et al (2012) Development of a transgenic early flowering pear (Pyrus communis L.) genotype by RNAi silencing of PcTFL1-1 and PcTFL1-2. Planta 235:1239–1251. https://doi.org/10.1007/s00425-011-1571-0
Gorelick DA, Heishman SJ (2006) Methods for clinical research involving cannabis administration. Methods Mol Med 123:235–253. https://doi.org/10.1385/1-59259-999-0:235
Grof CPL (2018) Cannabis, from plant to pill. Br J Clin Pharmacol 84:2463–2467. https://doi.org/10.1111/bcp.13618
Hall J, Bhattarai SP, Midmore DJ (2012) Review of flowering control in industrial hemp. J Nat Fibers 9:23–36. https://doi.org/10.1080/15440478.2012.651848
Hedman H, Källman T, Lagercrantz U (2009) Early evolution of the MFT-like gene family in plants. Plant Mol Biol 70:359–369. https://doi.org/10.1007/s11103-009-9478-x
Hepworth SR, Valverde F, Ravenscroft D et al (2002) Antagonistic regulation of flowering-time gene SOC1 by CONSTANS and FLC via separate promoter motifs. EMBO J 21:4327–4337. https://doi.org/10.1093/emboj/cdf432
Hyun Y, Richter R, Coupland G (2017) Competence to flower: age-controlled sensitivity to environmental cues. Plant Physiol 173:36–46. https://doi.org/10.1104/pp.16.01523
Immink RGH, Posé D, Ferrario S et al (2012) Characterization of SOC1’s central role in flowering by the identification of its upstream and downstream regulators. Plant Physiol 160:433–449. https://doi.org/10.1104/pp.112.202614
Jin S, Nasim Z, Susila H, Ahn JH (2021) Evolution and functional diversification of FLOWERING LOCUS T/TERMINAL FLOWER 1 family genes in plants. Semin Cell Dev Biol 109:20–30. https://doi.org/10.1016/j.semcdb.2020.05.007
Jung J-H, Seo Y-H, Seo PJ et al (2007) The GIGANTEA-regulated microRNA172 mediates photoperiodic flowering independent of CONSTANS in Arabidopsis. Plant Cell 19:2736–2748. https://doi.org/10.1105/tpc.107.054528
Jung J-H, Lee S, Yun J et al (2014) The miR172 target TOE3 represses AGAMOUS expression during Arabidopsis floral patterning. Plant Sci 215–216:29–38. https://doi.org/10.1016/j.plantsci.2013.10.010
Karlgren A, Gyllenstrand N, Källman T et al (2011) Evolution of the PEBP gene family in plants: functional diversification in seed plant evolution. Plant Physiol 156:1967–1977. https://doi.org/10.1104/pp.111.176206
Kaufmann K, Wellmer F, Muiño JM et al (2010) Orchestration of floral initiation by APETALA1. Science 328:85–89. https://doi.org/10.1126/science.1185244
Kerstetter RA, Poethig RS (1998) The specification of leaf identity during shoot development. Annu Rev Cell Dev Biol 14:373–398. https://doi.org/10.1146/annurev.cellbio.14.1.373
Khosa J, Bellinazzo F, Kamenetsky Goldstein R et al (2021) PHOSPHATIDYLETHANOLAMINE-BINDING PROTEINS: the conductors of dual reproduction in plants with vegetative storage organs. J Exp Bot 72:2845–2856. https://doi.org/10.1093/jxb/erab064
Khoury M, Cohen I, Bar-Sela G (2022) The two sides of the same coin”—medical cannabis, cannabinoids and immunity: pros and cons explained. Pharmaceutics 14:389. https://doi.org/10.3390/pharmaceutics14020389
Kim S, Soltis PS, Wall K, Soltis DE (2006) Phylogeny and domain evolution in the APETALA2-LIKE gene family. Mol Biol Evol 23:107–120. https://doi.org/10.1093/molbev/msj014
Kim D, Pertea G, Trapnell C et al (2013) TopHat2: accurate alignment of transcriptomes in the presence of insertions, deletions and gene fusions. Genome Biol 14:36. https://doi.org/10.1186/gb-2013-14-4-r36
Koltai H, Namdar D (2020) Cannabis phytomolecule “Entourage”: from domestication to medical use. Trends Plant Sci 25:976–984. https://doi.org/10.1016/j.tplants.2020.04.007
Kotoda N, Iwanami H, Takahashi S, Abe K (2006) Antisense expression of MdTFL1, a TFL1-like Gene, reduces the juvenile phase in apple. J Am Soc Hortic Sci 131:74–81. https://doi.org/10.21273/JASHS.131.1.74
Kunitake T, Tanigawa T, et al. (2009) Exposure of rooted cuttings to long days, or mother plants to high temperature, can avoid delayed flowering of Chrysanthemum morifolium Ramat. ‘Jinba’ for Winter and Spring Cut-flower Production. J Japan Soc Hort Sci 78:363–368. https://doi.org/10.2503/jjshs1.78.363
Lian H, Wang L, Ma N et al (2021) Redundant and specific roles of individual MIR172 genes in plant development. PLOS Biol 19:e3001044. https://doi.org/10.1371/journal.pbio.3001044
Lin W, Gupta SK, Arazi T, Spitzer-Rimon B (2021) MIR172d is required for floral organ identity and number in tomato. Int J Mol Sci 22:4659. https://doi.org/10.3390/ijms22094659
Lippman ZB, Cohen O, Alvarez JP et al (2008) The making of a compound inflorescence in tomato and related nightshades. PLoS Biol 6:e288. https://doi.org/10.1371/journal.pbio.0060288
Love MI, Huber W, Anders, S (2014) Moderated estimation of fold change and dispersion for RNA-seq data with DESeq2. Genome Biol 15:550. https://doi.org/10.1186/s13059-014-0550-8
Mathieu J, Yant LJ, Mürdter F et al (2009) Repression of flowering by the miR172 target SMZ. PLoS Biol 7:e1000148. https://doi.org/10.1371/journal.pbio.1000148
Matsoukas I (2014) Attainment of reproductive competence, phase transition, and quantification of juvenility in mutant genetic screens. Front Plant Sci. https://doi.org/10.3389/fpls.2014.00032
Mohamed R, Wang C-T, Ma C et al (2010) Populus CEN/TFL1 regulates first onset of flowering, axillary meristem identity and dormancy release in Populus. Plant J 62:674–688. https://doi.org/10.1111/j.1365-313X.2010.04185.x
Moon J, Suh S-S, Lee H et al (2003) The SOC1 MADS-box gene integrates vernalization and gibberellin signals for flowering in Arabidopsis. Plant J 35:613–623. https://doi.org/10.1046/j.1365-313X.2003.01833.x
Morea EGO, da Silva EM, de Silva GFF et al (2016) Functional and evolutionary analyses of the miR156 and miR529 families in land plants. BMC Plant Biol 16:40. https://doi.org/10.1186/s12870-016-0716-5
Nakamura S, Abe F, Kawahigashi H et al (2011) A wheat homolog of MOTHER OF FT AND TFL1 acts in the regulation of germination. Plant Cell 23:3215–3229. https://doi.org/10.1105/tpc.111.088492
Ó’Maoiléidigh DS, van Driel AD, Singh A et al (2021) Systematic analyses of the MIR172 family members of Arabidopsis define their distinct roles in regulation of APETALA2 during floral transition. PLoS Biol 19:e3001043. https://doi.org/10.1371/journal.pbio.3001043
Périlleux C, Bouché F, Randoux M, Orman-Ligeza B (2019) Turning meristems into fortresses. Trends Plant Sci 24:431–442. https://doi.org/10.1016/j.tplants.2019.02.004
Petit J, Salentijn EMJ, Paulo M-J et al (2020) Genetic architecture of flowering time and sex determination in hemp (Cannabis sativa L.): a genome-wide association study. Front Plant Sci. https://doi.org/10.3389/fpls.2020.569958
Poethig RS (2013) Vegetative phase change and shoot maturation in plants. Curr Top Dev Biol 105:125–152. https://doi.org/10.1016/B978-0-12-396968-2.00005-1
Rubio-Somoza I, Zhou C-M, Confraria A et al (2014) Temporal control of leaf complexity by miRNA-regulated licensing of protein complexes. Curr Biol 24:2714–2719. https://doi.org/10.1016/j.cub.2014.09.058
Ruzin SE (1999) Plant microtechnique and microcopy. Oxford University Press, New York
Ryu JY, Park C-M, Seo PJ (2011) The floral repressor BROTHER OF FT AND TFL1 (BFT) modulates flowering initiation under high salinity in Arabidopsis. Mol Cells 32:295–303. https://doi.org/10.1007/s10059-011-0112-9
Ryu JY, Lee H-J, Seo PJ et al (2014) The Arabidopsis floral repressor BFT delays flowering by competing with FT for FD binding under high salinity. Mol Plant 7:377–387. https://doi.org/10.1093/mp/sst114
Sablowski R (2007) Flowering and determinacy in Arabidopsis. J Exp Bot 58:899–907. https://doi.org/10.1093/jxb/erm002
Samach A, Onouchi H, Gold SE et al (2000) Distinct roles of CONSTANS target genes in reproductive development of Arabidopsis. Science 288:1613–1616. https://doi.org/10.1126/science.288.5471.1613
Song S, Wang G, Hu Y et al (2018) OsMFT1 increases spikelets per panicle and delays heading date in rice by suppressing EHD1, FZP and SEPALLATA-like genes. J Exp Bot 69:4283–4293. https://doi.org/10.1093/jxb/ery232
Spitzer-Rimon B, Duchin S, Bernstein N, Kamenetsky R (2019) Architecture and florogenesis in female Cannabis Sativa plants. Front Plant Sci 10:350. https://doi.org/10.3389/fpls.2019.00350
Trapnell C, Williams B, Pertea G et al. (2010) Transcript assembly and quantification by RNA-Seq reveals unannotated transcripts and isoform switching during cell differentiation. Nat Biotechnol 28:511–515. https://doi.org/10.1038/nbt.1621
Varkonyi-Gasic E, Wang T, Voogd C et al (2019) Mutagenesis of kiwifruit CENTRORADIALIS-LIKE genes transforms a climbing woody perennial with long juvenility and axillary flowering into a compact plant with rapid terminal flowering. Plant Biotechnol J 17:869–880. https://doi.org/10.1111/pbi.13021
Voogd C, Brian LA, Wang T et al (2017) Three FT and multiple CEN and BFT genes regulate maturity, flowering, and vegetative phenology in kiwifruit. J Exp Bot 68:1539–1553. https://doi.org/10.1093/jxb/erx044
Wahl V, Ponnu J, Schlereth A et al (2013) Regulation of flowering by Trehalose-6-Phosphate signaling in Arabidopsis thaliana. Science 339:704–707. https://doi.org/10.1126/science.1230406
Wang JW, Czech B, Weigel D (2009) miR156-regulated SPL transcription factors define an endogenous flowering pathway in Arabidopsis thaliana. Cell 138:738–749. https://doi.org/10.1016/j.cell.2009.06.014
Wang JW, Park MY, Wang LJ, Koo Y, Chen XY, Weigel D, Poethig RS (2011) miRNA control of vegetative phase change in trees. PLoS Genet 7:e1002012. https://doi.org/10.1371/journal.pgen.1002012
Wellmer F, Alves-Ferreira M, Dubois A et al (2006) Genome-wide analysis of gene expression during early Arabidopsis flower development. PLoS Genet 2:e117. https://doi.org/10.1371/journal.pgen.0020117
Wickland DP, Hanzawa Y (2015) The FLOWERING LOCUS T/TERMINAL FLOWER 1 gene family: functional evolution and molecular mechanisms. Mol Plant 8:983–997. https://doi.org/10.1016/j.molp.2015.01.007
Wigge PA, Kim MC, Jaeger KE et al (2005) Integration of spatial and temporal information during floral induction in Arabidopsis. Science 309:1056–1059. https://doi.org/10.1126/science.1114358
Wingler A (2018) Transitioning to the next phase: the role of sugar signaling throughout the plant life cycle. Plant Physiol 176:1075–1084. https://doi.org/10.1104/pp.17.01229
Wu G, Poethig RS (2006) Temporal regulation of shoot development in Arabidopsis thaliana by miR156 and its target SPL3. Development 133:3539–3547. https://doi.org/10.1242/dev.02521
Wu G, Park MY, Conway SR et al (2009) The sequential action of miR156 and miR172 regulates developmental timing in Arabidopsis. Cell 138:750–759. https://doi.org/10.1016/j.cell.2009.06.031
Xi W, Liu C, Hou X, Yu H (2010) MOTHER OF FT AND TFL1 regulates seed germination through a negative feedback loop modulating ABA signaling in Arabidopsis. Plant Cell 22:1733–1748. https://doi.org/10.1105/tpc.109.073072
Xu M, Hu T, Zhao J et al (2016) Developmental functions of miR156-regulated SQUAMOSA PROMOTER BINDING PROTEIN-LIKE (SPL) Genes in Arabidopsis thaliana. PLoS Genet 12:e1006263. https://doi.org/10.1371/journal.pgen.1006263
Yamaguchi N (2021) LEAFY, a pioneer transcription factor in plants: A mini-review. Front Plant Sci 12:701406. https://doi.org/10.3389/fpls.2021.701406
Yamaguchi A, Wu M-F, Yang L et al (2009) The microRNA-regulated SBP-Box transcription factor SPL3 is a direct upstream activator of LEAFY, FRUITFULL, and APETALA1. Dev Cell 17:268–278. https://doi.org/10.1016/j.devcel.2009.06.007
Yang L, Xu M, Koo Y et al (2013) Sugar promotes vegetative phase change in Arabidopsis thaliana by repressing the expression of miR156A and miR156C. eLife 2:e00260. https://doi.org/10.7554/eLife.00260
Yant L, Mathieu J, Dinh TT et al (2010) Orchestration of the floral transition and floral development in Arabidopsis by the bifunctional transcription factor APETALA2. Plant Cell 22:2156–2170. https://doi.org/10.1105/tpc.110.075606
Yoo SY, Kardailsky I, Lee JS et al (2004) Acceleration of flowering by overexpression of MFT (MOTHER OF FT AND TFL1). Mol Cells 17:95–101
Yu S, Cao L, Zhou C-M et al (2013) Sugar is an endogenous cue for juvenile-to-adult phase transition in plants. eLife 2:e00269. https://doi.org/10.7554/eLife.00269
Zhang B, Wang L, Zeng L et al (2015) Arabidopsis TOE proteins convey a photoperiodic signal to antagonize CONSTANS and regulate flowering time. Genes Dev 29:975–987. https://doi.org/10.1101/gad.251520.114
Zheng C, Ye M, Sang M, Wu R (2019) A Regulatory network for miR156-SPL module in Arabidopsis thaliana. Int J Mol Sci 20:E6166. https://doi.org/10.3390/ijms20246166
Zhu QH, Upadhyaya NM, Gubler F, Helliwell CA (2009) Over-expression of miR172 causes loss of spikelet determinacy and floral organ abnormalities in rice (Oryza sativa). BMC Plant Biol 9:149. https://doi.org/10.1186/1471-2229-9-149
Zhu Y, Klasfeld S, Jeong CW et al (2020) TERMINAL FLOWER 1-FD complex target genes and competition with FLOWERING LOCUS T. Nat Commun 11:5118. https://doi.org/10.1038/s41467-020-18782-1
Zhu Y, Klasfeld S, Wagner D (2021) Molecular regulation of plant developmental transitions and plant architecture via PEPB family proteins: an update on mechanism of action. J Exp Bot 72:2301–2311. https://doi.org/10.1093/jxb/eraa598
Acknowledgements
We thank Dr. Ross Peters (Agricultural Research Organization, the Volcani Institute) for his critical reading of the manuscript.
Funding
This work was partially supported by the Chief Scientist of the Israeli Ministry of Agriculture and Rural Development, grant no. 20-01-0177.
Author information
Authors and Affiliations
Corresponding author
Ethics declarations
Conflict of interest
The authors have no competing interests to declare that are relevant to the content of this article.
Additional information
Publisher's Note
Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.
Rights and permissions
Springer Nature or its licensor holds exclusive rights to this article under a publishing agreement with the author(s) or other rightsholder(s); author self-archiving of the accepted manuscript version of this article is solely governed by the terms of such publishing agreement and applicable law.
About this article
Cite this article
Spitzer-Rimon, B., Shafran-Tomer, H., Gottlieb, G.H. et al. Non-photoperiodic transition of female cannabis seedlings from juvenile to adult reproductive stage. Plant Reprod 35, 265–277 (2022). https://doi.org/10.1007/s00497-022-00449-0
Received:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s00497-022-00449-0