CRISPR/Cas9 mutants delineate roles of Populus FT and TFL1/CEN/BFT family members in growth, dormancy release and flowering

Abstract

Vegetative and reproductive phase change and phenology are economically and ecologically important traits. Trees typically require several years of growth before flowering and once mature, seasonal control of the transition to flowering and flower development is necessary to maintain vegetative meristems and for reproductive success. Members of two related gene subfamilies, FLOWERING LOCUST (FT) and TERMINAL FLOWER1 (TFL1)/CENTRORADIALIS (CEN)/BROTHER OF FT AND TFL1 (BFT), have antagonistic roles in flowering in diverse species and roles in vegetative phenology in trees, but many details of their functions in trees have yet to be resolved. Here, we used CRISPR/Cas9 to generate single and double mutants involving the five Populus FT and TFL1/CEN/BFT genes. ft1 mutants exhibited wild-type-like phenotypes in long days and short days, but after chilling to release dormancy showed delayed bud flush and GA3 could compensate for the ft1 mutation. After rooting and generating some phytomers in tissue culture, both cen1 and cen1ft1 mutants produced terminal as well as axillary flowers, indicating that the cen1 flowering phenotype is independent of FT1. CEN1 showed distinct circannual expression patterns in vegetative and reproductive tissues and comparison with the expression patterns of FT1 and FT2 suggest that the relative levels of CEN1 compared to FT1 and FT2 regulate multiple phases of vegetative and reproductive seasonal development.

Age-dependent seasonal growth cessation in Populus

“…Thus, we speculate that the age-dependent growth cessation is accompanied by VPC in Populus at an early tree age when the flowering induction has yet not been initiated. Populus FT and TFL1 family genes have evolved to coordinate the flowering regulation and the seasonal vegetative growth processes ( 25 ). This strategy largely depends on the spatiotemporal expressions of these genes ( 25 ).…”

“…Populus FT and TFL1 family genes have evolved to coordinate the flowering regulation and the seasonal vegetative growth processes ( 25 ). This strategy largely depends on the spatiotemporal expressions of these genes ( 25 ). In our study, the declined expression of FT2 is tightly related to VPC, while its expression is relatively stable in different ages of adult trees.…”

“…SVL regulates seasonal growth by antagonistically acting on the ABA and gibberellic acid (GA) pathways, which are the two crucial plant hormones involved in seasonal growth. In addition, FT1, another paralog of FT2, is strongly induced by cold in the winter ( 2 ) and has recently been shown to have a critical role in dormancy release and the regulation of bud break ( 24 , 25 ). Based on these studies, it is suggested that perennial trees have evolved the ability to integrate the regulation of flowering with the regulation of the seasonal cycling between active and dormant vegetative growth through common genes and signal cascades ( 4 ).…”

Advancing tree genomics to future proof next generation orchard production

“…The delayed maturity, long life span, and polycarpic growth of trees require a complex regulatory network to regulate the timing of developmental transitions and synchronise environmental signals with phenological events to ensure survival and successful reproduction (Brunner et al, 2017). While flowering gene homologs from woody perennials often show a conserved role in regulation of flowering in model annuals such as Arabidopsis and tobacco, functional studies in trees reveal shared and distinct functional aspects, highlighting the diverse roles of paralogs (Hsu et al, 2011;Sheng et al, 2022) and the pleiotropic effects of flowering genes on growth (Andréet al, 2022;Sheng et al, 2023), organ identity (Zhang et al, 2021), and phenology in trees (Ding and Nilsson, 2016). Therefore, targeting specific genes to reduce the years to reproduction in trees is a complex task that depends on the species in question.…”

A MADS-box gene-induced early flowering pear (Pyrus communis L.) for accelerated pear breeding

“…A similar effect was achieved using silencing of the TERMINAL FLOWER1 ( TFL1 ) genes in apple and pear ( Kotoda et al., 2006 ; Freiman et al., 2011 ; Flachowsky et al., 2012 ; Yamagishi et al., 2016 ). Recently, gene editing of CEN genes gave rise to early flowering kiwifruit species ( Varkonyi-Gasic et al., 2019 ; Herath et al., 2023 ), whilst mutagenesis of CEN1 and overexpression of FT1 or FT2 resulted in precocious flowering in poplar ( Sheng et al., 2023 ). Editing of TFL1 for early flowering was also performed in apple and pear ( Charrier et al., 2019 ); however, this approach has problems including too early ( in vitro ) flowering, complex gene editing profiles and mosaicism, low transformation efficiency for pear, and the recessive nature of the mutation.…”