A Quartet of PIF bHLH Factors Provides a Transcriptionally Centered Signaling Hub That Regulates Seedling Morphogenesis through Differential Expression-Patterning of Shared Target Genes in Arabidopsis

摘要

Dark-grown seedlings exhibit skotomorphogenic development. Genetic and molecular evidence indicates that a quartet of Arabidopsis Phytochrome (phy)-Interacting bHLH Factors (PIF1, 3, 4, and 5) are critically necessary to maintaining this developmental state and that light activation of phy induces a switch to photomorphogenic development by inducing rapid degradation of the PIFs. Here, using integrated ChIP–seq and RNA–seq analyses, we have identified genes that are direct targets of PIF3 transcriptional regulation, exerted by sequence-specific binding to G-box (CACGTG) or PBE-box (CACATG) motifs in the target promoters genome-wide. In addition, expression analysis of selected genes in this set, in all triple pif -mutant combinations, provides evidence that the PIF quartet members collaborate to generate an expression pattern that is the product of a mosaic of differential transcriptional responsiveness of individual genes to the different PIFs and of differential regulatory activity of individual PIFs toward the different genes. Together with prior evidence that all four PIFs can bind to G-boxes, the data suggest that this collective activity may be exerted via shared occupancy of binding sites in target promoters. Author Summary An important issue in understanding mechanisms of eukaryotic transcriptional regulation is how members of large transcription-factor families, with conserved DNA–binding domains (such as the 162-member Arabidopsis bHLH family), discriminate between target genes. However, the specific question of whether, and to what extent, closely related sub-family members, with potential overlapping functional redundancy (like the quartet of Phytochrome (phy)-Interacting bHLH transcription Factors (PIF1, 3, 4, and 5) studied here), share regulation of target genes through shared binding to promoter-localized consensus motifs does not appear to have been widely investigated. Here, using ChIP–seq analysis, we have identified genes that bind PIF3 to conserved, sequence-specific sites in their promoters; and, using RNA–seq, we have identified those genes displaying altered expression in various pif mutants. Integration of these data identifies those genes that are likely direct targets of transcriptional regulation by PIF3. Our data suggest that the PIF quartet members share directly in transcriptional activation of numerous target genes, potentially via redundant promoter occupancy, in a manner that varies quantitatively from gene to gene. This finding suggests that these PIFs function collectively as a signaling hub, selectively partitioning common upstream signals from light-activated phys at the transcriptional-network interface.