Abstract
FLOWERING LOCUS M (FLM), a component of the thermosensory flowering time pathway in Arabidopsis thaliana, is regulated by temperature-dependent alternative splicing (AS). The main splicing variant, FLM-β, is a well-documented floral repressor that is down-regulated in response to increasing ambient growth temperature. Two hypotheses have been formulated to explain how flowering time is modulated by AS of FLM. In the first model a second splice variant, FLM-δ, acts as a dominant negative isoform that competes with FLM-β at elevated ambient temperatures, thereby indirectly promoting flowering. Alternatively, it has been suggested that the induction of flowering at elevated temperatures is caused only by reduced FLM-β expression. To better understand the role of the two FLM splice forms, we employed CRISPR/Cas9 technology to specifically delete the exons that characterize each splice variant. Lines that produced repressive FLM-β but were incapable of producing FLM-δ were late flowering. In contrast, FLM-β knockout lines that still produced FLM-δ flowered early, but not earlier than the flm-3 loss of function mutant, as would be expected if FLM-δ had a dominant-negative effect on flowering. Our data support the role of FLM-β as a flower repressor and provide evidence that a contribution of FLM-δ to the regulation of flowering time in wild-type A. thaliana seems unlikely.
Highlights
The correct timing of the transition from vegetative growth to flowering is critical to ensure reproductive success
Plants grown at 23°C were transformed by floral dipping using Agrobacterium tumefaciens-mediated gene transfer according to standard protocols
FLOWERING LOCUS M (FLM)-ΔE2 carries a deletion of 57 bp that covers most of exon 2, which is normally incorporated in the repressive FLM-β splice variant, apart from 2 bp at the 5’ end and the first 2 bp of intron 3 (Fig. 1)
Summary
The correct timing of the transition from vegetative growth to flowering is critical to ensure reproductive success. Due to its importance, flowering time is regulated by an intricate genetic network that integrates both endogenous and environmental signals such as photoperiod, namely day length, and temperature (Srikanth and Schmid, 2011). Two aspects regarding the regulation of flowering by temperature can be distinguished: the response to prolonged periods of cold, such as overwintering and vernalization, and the effects of ambient temperature. In Arabidopsis thaliana vernalization controls flowering through the MADS domain transcription factor FLOWERING LOCUS C (FLC), which is epigenetically silenced in response to non-freezing temperatures (Gendall et al, 2001; Bastow et al, 2004).
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