Dissecting the response to photoperiod at the cell-type level

Photoperiod—the length of light and dark periods each day—together with ambient temperature, light, circadian clock, plant age, hormone levels, and vernalization, are critical factors fine tuning flowering time. The timing of flowering is an essential parameter in both agricultural and natural settings, as it influences seed number and therefore crop yield and plays a critical role in reproductive success in nature.

Flowering is initiated by the transition of the shoot apical meristem (SAM) from a vegetative into a reproductive state. In Arabidopsis thaliana, photoperiod was shown to be perceived in the leaf vasculature, especially in the phloem companion cells (PCCs) where light signaling and circadian networks converge to regulate the expression of key floral genes (reviewed in Giakountis and Coupland, 2008). A transition to long days results in increased expression of the florigen FLOWERING LOCUS T (FT) and its homolog TWIN SISTER OF FT (TSF), which act as long-distance signals between the PCCs of the leaf vasculature and the SAM (Turck et�al., 2008) to promote flowering.

Epigenetic mechanisms such as histone post-translational modifications and modulation of transcription factor binding have been shown to regulate gene expression in PCCs and therefore flowering time (Turck et�al., 2008; You et�al., 2019). However, the underlying cis-regulatory elements and chromatin architecture controlling transcriptional responses sensing photoperiod switches have remained poorly understood. To tackle these questions, Tian et�al. (2021) combined the isolation of cell-type-specific nuclei with the profiling of chromatin accessibility and gene expression.

Isolation of nuclei tagged in specific cell types (INTACTs; Deal and Henikoff, 2011) was used to obtain PCCS (vascular-) and epidermis-specific nuclei (see Figure�1). Genome-wide information on chromatin accessibility in these nuclei types was then achieved by an assay for transposase-accessible chromatin using sequencing (ATAC-seq; Buenrostro et�al., 2013). In ATAC-seq, a prokaryotic transposase is used to simultaneously fragment and tag accessible genomic regions with sequencing adaptors. As open chromatin is often correlated with active transcription and/or regulatory regions, ATAC-seq offers the opportunity to identify cis-regulatory elements and transcription factor-binding sites that control gene expression.

Figure 1.

Tissue-type profiling of chromatin accessibility and transcription in response to changes in photoperiod. Nuclei were isolated using the INTACT system (Deal and Henikoff, 2011) in transgenic plants expressing a vascular (GAS1) or an epidermis (FDH) specific “nuclear targeting fusion” (NTF) protein. After biotinylation of NTF by a constitutively expressed biotin-ligase (ACT2p:BirA), cell-type-specific nuclei were purified using magnetic streptavidin-coated beads. After non-labeled nuclei were discarded, chromatin accessibility and gene expression were assessed by ATAC-seq (Buenrostro et�al., 2013) and RNA-seq, respectively. To investigate the effect of changes in the photoperiod, 14 days old seedlings grown in short day (SD) were either transferred to long day (LD) for 3 days or remained in short day. (Adapted from Tian et�al. [2021, Figure�1]).

By combining INTACT with ATAC-seq, the authors identified around 20,000 accessible chromatin regions (ACRs), the majority which was localized in close proximity to protein-coding genes. Most ACRs were shared between vascular and epidermal nuclei, illustrating overall conserved mechanisms regulating chromatin accessibility across tissues. In contrast to epidermis-specific ACRs, vascular-specific ACRs showed a slight increase in distal ACRs and were enriched at genes involved in regulation of flowering time, consistently with the role of PCCs. To investigate the tissue-specific impact of day length changes on chromatin accessibility, seedlings grown in short days were subjected to 3 days of long day length (see Figure�1). INTACT-ATAC-seq revealed that a shift to long day increased chromatin accessibility, particularly in vascular nuclei. By contrast, open chromatin tended to decline in the epidermis in long days. RNA-seq further showed photoperiod-dependent open chromatin to be strongly correlated with elevated gene expression, consistent with the notion that transcription leads to increased chromatin accessibility and vice versa.

Tian et�al. (2021) used this large-scale data to identify putative new components of the genetic flowering time network. They found that, in both the vasculature and the epidermis, the chromatin proximal to the TREHALOSE-6-PHOSPHATE SYNTHASE9 (TPS9) gene was decompacted and TPS9 transcription was increased upon an inductive photoperiod. Consistently, plants lacking TPS9 showed a delayed flowering time compared with wild type. To validate that TPS9 promotes flowering in the vasculature, TPS9 was specifically downregulated in PCCs using amiRNAs, which lead to a late flowering phenotype. TPS9 is a catalytically inactive homolog of TPS1 and is expressed in the leaf vasculature where it activates FT transcription to promote flowering. Interestingly, trehalose-6-phosphate acts as a signaling metabolite coordinating sugar status and carbon assimilation. The identification of TPS9 as a flowering time regulator responding to changes in day length strengthens the important link between metabolism and the environment, and emphasizes the importance of PCCs as a signaling hub that integrates a wide range of signals to control systemic plant growth and development.

Conflict of interest statement. None declared.