Transcriptional repressor PRR5 directly regulates clock-output pathways

Abstract

The circadian clock is an endogenous time-keeping mechanism that enables organisms to adapt to external daily cycles. The clock coordinates biological activities with these cycles, mainly through genome-wide gene expression. However, the exact mechanism underlying regulation of circadian gene expression is poorly understood. Here we demonstrated that an Arabidopsis PSEUDO-RE-SPONSE REGULATOR 5 (PRR5), which acts in the clock genetic circuit, directly regulates expression timing of key transcription factors involved in clock-output pathways. A transient expression assay and ChIP-quantitative PCR assay using mutated PRR5 indicated that PRR5 associates with target DNA through binding at the CCT motif in vivo. ChIP followed by deep sequencing coupled with genome-wide expression profiling revealed the direct-target genes of PRR5. PRR5 direct-targets include genes encoding transcription factors involved in flowering-time regulation, hypocotyl elongation, and cold-stress responses. PRR5-target gene expression followed a circadian rhythm pattern with low, basal expression from noon until midnight, when PRR9, PRR7, and PRR5 were expressed. ChIPquantitative PCR assays indicated that PRR7 and PRR9 bind to the direct-targets of PRR5. Genome-wide expression profiling using a prr9 prr7 prr5 triple mutant suggests that PRR5, PRR7, and PRR9 repress these targets. Taken together, our results illustrate a genetic network in which PRR5, PRR7, and PRR9 directly regulate expression timing of key transcription factors to coordinate physiological processes with daily cycles.ChIP-seq | plant T he circadian clock in plants regulates a broad range of biological processes. For example, hypocotyl elongation is observed before dawn and cold-stress responses reach maximal levels in the afternoon in Arabidopsis thaliana (1, 2), all largely because of circadian coordination of these biological processes (clock-output) with daily cycles. The circadian clock mechanism controls the temporal regulation of numerous genes involved in output processes (3-5).A number of recent studies have described the genetic components of the clock in Arabidopsis. CIRCADIAN CLOCK-ASSOCI-ATED 1 (CCA1) and LATE ELONGATED HYPOCOTYL (LHY) encode morning-expressed MYB transcription factors (TFs) that directly repress TIMING OF CAB EXPRESSION 1 [TOC1, also called PSEUDO-RESPONSE REGULATOR 1 (PRR1)], EARLY FLOWERING 3 (ELF3), ELF4, and LUXARRHYTHMO (LUX) (6-10). ELF3 and LUX associate with upstream region of PRR9, and repress PRR9 expression (11, 12). Expression of PRR9 and PRR7 are activated by CCA1 and LHY (13). CCA1 and LHY are in turn repressed by four PRR proteins, PRR9, PRR7, PRR5, and TOC1 from early daytime through to around midnight (14-16). These TFs form a negative feedback loop for clock function (12,17,18). Evidence is accumulating that these TFs directly regulate the expression of genes involved in clock-output pathways. LUX, ELF3, and ELF4 together form the "evening complex" that directly represses expression of PHYTOCHROME INTERACTING FACTOR 4 (PIF4) and P...

LNK genes integrate light and clock signaling networks at the core of the Arabidopsis oscillator

“…Many clock-regulated genes with peak expression in the morning are repressed throughout the day and during the early night by members of the TOC1/PRR1 family of clock proteins. To determine whether the LNKs were regulated by members of this protein family, we reexamined data describing TOC1/PRR1 and PRR5 binding sites in the Arabidopsis genome using ChIP followed by sequencing (ChIP-seq) (31,33). Indeed, we found that the regulatory region of LNK3 was directly bound by TOC1/PRR1 (31).…”

PIFs: Systems Integrators in Plant Development

“…Interestingly, the PIFs themselves are PIF bound, which may be linked to the lightinduced expression of some PIFs, such as PIF7 (Leivar et al, 2008a), PIF4, and PIF5 (Leivar et al, 2009), during deetiolation and suggests the existence of a complex autoregulatory feedback mechanism. PIF4/5 also emerge as direct targets of circadian clock proteins TOC1, PRR5, and PRR7 (Huang et al, 2012;Nakamichi et al, 2012;Liu et al, 2013a), which might contribute to the regulation of the oscillatory expression of PIF4/5 as a repression mechanism complementary to the EC. In addition to the described binding by BZR1 , BRs might also target PIF5 through direct binding by BES1 .…”

“…In the case of cold, the circadian clock regulates the expression and gates the cold induction of the C-repeat binding factors CBF1, 2, and 3, a pathway with a major role in plant cold acclimation and freezing tolerance. The mechanism involves direct CBF induction in the morning by CCA1 and LHY and evening repression by PRR5, 7, and 9 (Dong et al, 2011;Nakamichi et al, 2012). PIF7 and PIF4 also regulate the circadian and diurnal expression of CBFs at warm temperatures, thus potentially participating in the photoperiodic control of the CBF cold acclimation pathway (Kidokoro et al, 2009;Lee and Thomashow, 2012).…”

The Multiple Signals That Control Tuber Formation

“…Accumulation of CDF1 mRNA levels is regulated by the transcription-repressing PRR protein family (Nakamichi et al, 2007(Nakamichi et al, , 2012. Mutations of the PRRs resulted in elevated expression of CDF1 (Nakamichi et al, 2007;Ito et al, 2008).…”

“…Mutations of the PRRs resulted in elevated expression of CDF1 (Nakamichi et al, 2007;Ito et al, 2008). PRR proteins function in the clock circuit by directly regulating the timing of expression of key TFs involved in clock-output pathways (Nakamichi et al, 2012;Hayama et al, 2017). In Arabidopsis, the PRRs act to promote the floral transition by increasing the rate of FT transcription (Hayama et al, 2017).…”