Crosstalk in the darkness: bulb vernalization activates meristem transition via circadian rhythm and photoperiodic pathway

Ben Michael et al. BMC Plant Biology (2020) 20:77 https://doi.org/10.1186/s12870-020-2269-x RESEARCH ARTICLE Open Access Crosstalk in the darkness: bulb vernalization activates meristem transition via circadian rhythm and photoperiodic pathway Tomer E. Ben Michael1,2, Adi Faigenboim1, Einat Shemesh-Mayer1, Itzhak Forer1, Chen Gershberg1, Hadass Shafran1, Haim D. Rabinowitch2 and Rina Kamenetsky-Goldstein1* Abstract Background: Geophytes possess specialized storage organs - bulbs, tubers, corms or rhizomes, which allow their survival during unfovarable periods and provide energy support for sprouting and sexual and vegetative reproduction. Bulbing and flowering of the geophyte depend on the combined effects of the internal and external factors, especially temperature and photoperiod. Many geophytes are extensively used in agriculture, but mechanisms of regulation of their flowering and bulbing are still unclear. Results: Comparative morpho-physiological and transcriptome analyses and quantitative validation of gene expression shed light on the molecular regulation of the responses to vernalization in garlic, a typical bulbous plant. Long dark cold exposure of bulbs is a major cue for flowering and bulbing, and its interactions with the genetic makeup of the individual plant dictate the phenotypic expression during growth stage. Photoperiod signal is not involved in the initial nuclear and metabolic processes, but might play role in the later stages of development, flower stem elongation and bulbing. Vernalization for 12 weeks at 4 °C and planting in November resulted in flower initiation under short photoperiod in December–January, and early blooming and bulbing. In contrast, non-vernalized plants did not undergo meristem transition. Comparisons between vernalized and non-vernalized bulbs revealed ~ 14,000 differentially expressed genes. Conclusions: Low temperatures stimulate a large cascades of molecular mechanisms in garlic, and a variety of flowering pathways operate together for the benefit of meristem transition, annual life cycle and viable reproduction results.The circadian clock appears to play a central role in the transition of the meristem from vegetative to reproductive stage in bulbous plant, serving as integrator of the low-temperature signals and the expression of the genes associated with vernalization, photoperiod and meristem transition. The reserved photoperiodic pathway is integrated at an upstream point, possibly by the same receptors. Therefore, in bulb, low temperatures stimulate cascades of developmental mechanisms, and several genetic flowering pathways intermix to achieve successful sexual and vegetative reproduction. Keywords: Allium sativum, Bulbing, Flowering, Reproductive meristem, Low temperature Background Perennial plant species carefully time their flowering with seasonal changes to ensure and maximize reproductive success. Flowering is induced, initiated and promoted both by internal and environmental cues, such as physiological age, gibberellin synthesis, photoperiod, and/or temperatures. The combined effect of two or more of these * Correspondence: 1 Institute of Plant Sciences, ARO, The Volcani Center, Rishon LeZion, Israel Full list of author information is available at the end of the article agents results in an integrated regulatory network that controls flowering time and quality [4, 89]. Despite the considerable genetic differences between and within species [9, 10, 18, 29], there is a great physiological similarity among plants from the temperate zone as many of them depend on vernalization for floral induction. Accumulation of chilling hours regulates essential changes in water status, in hormonal balance, in respiration and in carbohydrate mobilization [39], with a consequent increase in plant receptivity to changes in day length in the following spring [4]. © The Author(s). 2020 Open Access This article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated. Ben Michael et al. BMC Plant Biology (2020) 20:77 Dependence on low temperatures evolved several times in the history of plants, with the consequent involvement of a number of regulatory mechanisms [4, 10, 40, 74]. For instance, Arabidopsis remains vegetative when FRIGIDA (FRI) upregulates the floral repressor FLOWERING LOCUS C (FLC), which in turn downregulates the floral integrators FT, FD, and SOC1. Cold induction downregulates FLC, thus enabling the expression of floral integrators and consequently the meristem transition from the vegetative to the reproductive state [44, 60, 79, 80]. An alternative regulating mechanism is common in the monocot cereals, where FThomolog VERNALIZATION 3 (VRN3) is repressed by VRN2. Following cold induction, VRN1 represses VRN2 expression in the leaves, thus enabling the expression of VRN3 and the consequent transition of the meristem [74]. In bulb onion, FT-like genes control the initiation of both bulbing and flowering [45]. Under short photoperiod, high AcFT4 expression inhibits bulb formation by repressing AcFT1. This genotype-specific inhibitory effect gradually weakens when days elongate, and thereafter the inductive photoperiod downregulates AcFT4, with the consequent expression of bulb-promoting AcFT1. On the other hand, low temperatures in storage and/or in the field promote the upregulation of AcFT2, which encodes for flowering in the spring/summer. Vernalization effects on the upregulation of FT-like sequences (LiFTL) were also reported for Lilium longiflorum [47, 49]. In tulip, TgFT2 is considered to act as florigen, whereas TgFT1 and TgFT3 may have bulb-specific functions [47]. Unlike vernalization, the molecular mechanism of the photoperiodic pathway evolved in early times and is well conserved in plants [4, 91]. It involves stabilizing/destabilizing balance between photoreceptors, with the consequent timely induction and initiation of flowering by the expression of CONSTANS (CO), which is regulated by both, GIGANTEA (GI) and the circadian rhythm [84, 91]. In many plant species, flowering induction and initiation require sequential and combined effects of vernalization and photoperiod. Hence a ‘memory’ of the vernalization effect is maintained by epigenetic mechanisms [8, 86]. In Arabidopsis, these chromatin modifications suppress the floral repressor FLC [16, 86], while in cereals they upregulate the floral activator VRN1 [67, 90]. It is generally accepted that for both mono- and dicots, FTs homologs act as th (...truncated)