PIFs: systems integrators in plant development

Abstract

Phytochrome-interacting factors (PIFs) are members of the Arabidopsis thaliana basic helix-loop-helix family of transcriptional regulators that interact specifically with the active Pfr conformer of phytochrome (phy) photoreceptors. PIFs are central regulators of photomorphogenic development that act to promote stem growth, and this activity is reversed upon interaction with phy in response to light. Recently, significant progress has been made in defining the transcriptional networks directly regulated by PIFs, as well as the convergence of other signaling pathways on the PIFs to modulate growth. Here, we summarize and highlight these findings in the context of PIFs acting as integrators of light and other signals. We discuss progress in our understanding of the transcriptional and posttranslational regulation of PIFs that illustrates the integration of light with hormonal pathways and the circadian clock, and we review seedling hypocotyl growth as a paradigm of PIFs acting at the interface of these signals. Based on these advances, PIFs are emerging as required factors for growth, acting as central components of a regulatory node that integrates multiple internal and external signals to optimize plant development.

Figure 1.

Systems Integration of Environmental and Internal Signals through the PIF Transcription Factors. (A) Summary of the biological functions of the PIFs as integrators of environmental and internal signals to implement multiple facets of downstream morphogenesis throughout the plant life cycle, acting as positive (→) or negative (−|) regulators. (B) Simplified schematic illustration of PIF function in transducing light signals downstream of phy photoreceptors and in integrating information from other internal and environmental pathways to provide a coordinated transcriptional response that implements morphogenesis. (C) Structural and functional domains of the PIF subfamily of bHLH transcription factors. (D) PIFs are required to promote growth in etiolated seedlings, under diurnal SD conditions, and in response to shade. Visible phenotypes of wild-type and pifq mutant seedlings grown in darkness or in SDs for 3 d (left panel) or for 7 d in light (high R/FR) or simulated shade (low R/FR, 2 d in light + 5 d in shade) (right panel). (Right panel modified from Leivar et al. [2012b], Figure 1B.)

Figure 2.

Integration of Phytochrome-Regulated Transcriptional Networks through the PIF Transcription Factors. (A) PIL1 expression is reciprocally regulated by light and PIFs during deetiolation, shade, and diurnal conditions. The wild type and pif mutants were grown in the dark for 2 d and then transferred to Rc (left panel), in white light (WL; high R/FR) for 2 d and then transferred to simulated shade (low R/FR) (middle panel), or in SD for 2 d and then kept in SD for an additional day (right panel). (B) and (C) Comparative transcriptomic analysis of PIF-regulated genes during deetiolation, shade, and diurnal conditions. The percentage of genes that are bound by PIFq members is shown in parentheses. PIFq-bound genes were defined by combining the list of published PIF1-, PIF3-, PIF4-, and PIF5-bound genes (Oh et al., 2009, 2012; Hornitschek et al., 2012; Zhang et al., 2013), which results in a combined list of 5073 PIFq-bound genes that are targeted by one or more PIF, representing ∼15% of the Arabidopsis genome. A list of these genes is presented in Supplemental Data Set 1. (B) Comparison of the following sets of light/growth-responsive PIF-dependent genes: (1) During deetiolation, the 839 genes that respond rapidly to R light (1 h, R1) or in a sustained manner (2d, Rc) in wild-type seedlings compared with darkness and that show PIFq regulation in the dark (Leivar et al., 2009). (2) In response to shade, the 265 genes that respond rapidly (1 h, FR1) or slowly (3 h, FR3) to low R/FR in wild-type seedlings that show moderately to robustly (>1.5-fold) PIFq dependency (Leivar et al., 2012b). (3) In diurnal conditions, the 118 genes whose expression correlates with the stationary (S) or growth (G) phases that show moderate to robust (>1.5-fold) PIF4/5 dependency (Nozue et al., 2011). (C) The PIF-regulated genes in (B) were divided in PIF-induced and PIF-repressed genes. Most of the PIF-induced genes are light (R1, Rc) repressed, shade induced, and/or growth induced, whereas the PIF-repressed set shows the opposite light-responsive pattern. Genes showing complex patterns were removed from this analysis. (D) Mean fold change in expression relative to wild type–dark (left graph) or wild type–G (right graph) of the 22 PIF-induced central class genes defined in (C), showing reciprocal regulation by light and PIFs during deetiolation, shade, and diurnal conditions. ([A] is modified from Leivar et al. [2009], Figure 6; Leivar et al. [2012b], Figure 4; and Soy et al. [2012], Figure 3A.]

Figure 3.

Modes of Transcriptional Regulation by the PIFs. (A) PIFs predominantly act as constitutive transcriptional activators of genes like PIL1 in the dark (etiolated seedlings), in response to shade ,or at night under diurnal conditions, and light reverses this activity through phy-induced removal or inactivation of the transcription factors. (B) PIFs also act as constitutive transcriptional repressors of a relatively smaller subset of light-induced genes, such as PSY, especially during deetiolation, suggesting that PIFs may have a dual activity depending on the promoter and developmental context. (C) PIFs act as constitutive coactivators of transiently light-induced genes like ELIP2 during deetiolation. This model implies the participation of an additional unknown transcriptional coactivator (represented as X) that is activated by light.

Figure 4.

Transcriptional and Posttranslational Regulation of the PIFs. (A) Posttranslational regulation of PIF transcriptional activity. PIF binding proteins exert regulation of PIF transcriptional activity by acting as coregulators (green circles), blocking their ability to bind to DNA (blue circles), or inhibiting their intrinsic capacity to activate transcription (orange circle). In addition, interaction with phy Pfr triggers PIF degradation or inhibits PIF activity. (B) Direct transcriptional regulation of the PIF genes. PIFs potentially are targeted by photomorphogenic-, clock-, hormone-, and development-related transcription factors (TF). Based on ChIP-seq and ChIP-chip data, the regulatory regions of PIF genes (rectangles) are bound by PIFs and other factors (circles). Lines connecting TF with the PIFs depict the binding that has been verified experimentally (see text for details).

Figure 5.

Interface of PIF and Hormonal Pathways to Regulate Seedling Photomorphogenesis. Simplified model depicting hormone regulation of PIF levels and/or activity (A) and direct PIF transcriptional regulation of hormonal pathway components upstream (A) or downstream (B) of the PIF proteins to regulate growth. Solid lines originating from PIFs represent direct transcriptional events that have been proposed elsewhere, whereas dotted lines represent new direct potential connections based on ChIP-seq and transcriptomic data (Supplemental Table 1). The question mark indicates a possible connection based on data for hook development (An et al., 2012). Arrows indicate induction, whereas T-lines indicate repressive action. Colors indicate genes that are directly induced (orange) or repressed (blue) by the PIFs. Integration with other signals is represented by the effect of temperature and sugars. tp, transport.

Figure 6.

Interface of PIF, Clock, and Hormones to Regulate Diurnal Growth. Simplified model depicting PIF-mediated hypocotyl growth under SD conditions integrating hormone and clock signals. PIF4 and PIF5 mRNA expression is repressed by the circadian clock during the beginning of the dark period and rises in the middle of the night to peak at dawn in SD-grown seedlings. By contrast, PIF3 and PIF1 mRNA remain constant throughout the day (top panel). phyA and phyB activities induce degradation of PIFs during the day, and phyB is active during the early night to keep PIF3 and probably PIF1 levels low. As the night proceeds, phyB activity decreases and PIF1 and PIF3 progressively accumulate and peak at the end of the night. For PIF4 and PIF5, coincidence of clock and light regulation ensures that protein accumulation peaks before dawn. DELLA proteins accumulate during the night-to-day transition and remain high during the day to block the DNA binding activity of PIFs during dawn and possibly throughout the day to inhibit residual PIF protein (second panel). The clock maintains light repression of hormonal pathway genes during early night and gates their expression at dawn. PIFs mediate the induction of GA-, auxin-, and BR-related genes at the end of the night and directly induce the expression of growth-related genes at the end of the night (exemplified by PIL1, HFR1, and XTR7) to induce hypocotyl growth before dawn (bottom panels). (Adapted from Soy et al. [2012], Figure 5.)