Location: Plant Gene Expression Center
2018 Annual Report
Objectives
The long-term goal of this program is to define the molecular mechanisms by which the phy family of sensory photoreceptors perceive and transduce informational light signals from the environment to photoresponsive genes in regulating plant adaptational growth and development. The central goal of this proposal is to define the mechanisms by which the phys induce and modulate the Shade-Avoidance Response (SAR) in the constantly fluctuating light environment encountered in dense crop vegetational canopies. Toward this goal, the specific objectives for this funding period are:
Objective 1: Define the biological functions of the multiple Phy-Interacting transcription Factor (PIF)-family members in controlling the shade-avoidance response, including dissection of the relative contributions of the individual PIFs to this process.
Objective 2: Define the direct gene-targets of shade-active-PIF transcriptional regulation, and determine whether, and to what extent, this regulation involves differential direct targeting of rapidly shade-responsive genes by the individual PIF-family members.
Objective 3: Define the mechanism by which genes that lack apparent PIF regulation during skotomorphogenesis, but acquire PIF-dependent shade responsiveness in green seedlings (‘shade-specific’ genes), acquire this capacity.
Approach
Objective 1: To define the biological functions of the multiple Phy-Interacting transcription Factor (PIF)-family members in controlling the Shade-Avoidance Response (SAR), including dissection of the relative contributions of the individual PIFs to this process, we will perform phenotypic analyses of our array of higher order, pentuple and sextuple combinations of the pif mutants. This will enable us to define the relative quantitative contributions of the individual PIFs to the various facets of the SAR.
Objective 2: To define the direct gene-targets of shade-active-PIF transcriptional regulation, and determine whether, and to what extent, this regulation involves differential direct targeting of rapidly shade-responsive genes by the individual PIF-family members, we will analyze the global expression profiles of this set of higher-order pif-mutant combinations in response to shade, using RNA-seq, and identify the PIF-bound subset of these genes by ChIP-seq analysis. Direct targets of transcriptional regulation by the individual PIFs will be identified by integrating the RNA-seq and ChIP-seq data for each PIF as we have done for the PIF quartet in dark-grown seedlings.
Objective 3: To define the mechanism by which genes that lack apparent PIF regulation during skotomorphogenesis, but acquire PIF-dependent shade responsiveness in green seedlings (‘shade-specific’ genes), acquire this capacity, we will use ChIP-seq analysis for altered accessibility of PIF-binding sites and altered histone marks, indicative of nucleosome modifications, in response to shade. This will enable us to interrogate the genomes of light- and dark-grown Arabidopsis seedlings for differential histone-mark signatures correlated with the acquisition of PIF-dependent shade-signal responsiveness.
Progress Report
This is the first report for project 2030-21000-051-00D, which started March 2018, and continues research from project 2030-21000-042-00D, “Molecular Mechanisms of Photoperception, Signaling and Gene Regulation by the Phytochrome Family.”
Under Objective 1, ARS and University of California, Berkeley scientists have performed phenotypic analyses of our array of higher order, pentuple and sextuple combinations of the Phy-Interacting transcription Factor (pif) mutants, to define the biological functions of these multiple PIF-family members in controlling the Shade-Avoidance Response (SAR). The data indicate that all five of the individual PIFs tested (PIF1, PIF3, PIF4, PIF5, and PIF7) contribute cooperatively, but differentially, to this process.
In support of Objective 2, ARS and UC Berkeley scientists have performed quantitative reverse transcription polymerase chain reaction (RT-qPCR) analysis of known PIF-quartet-dependent, rapidly shade-responsive genes in the selected pif-mutant combinations for assessment of potential contributions of individual PIFs to transcriptional activation of these genes. The data reveal a complex combinatorial pattern of transcriptional regulation, whereby the different PIFs can activate a given individual gene to different extents, and an individual PIF can activate different genes to different extents.
Related to Objective 3, ARS and UC Berkeley scientists are developing the assays and methodology necessary to permit interrogation of the genomes of light- and dark-grown Arabidopsis seedlings for differential histone-mark signatures correlated with the acquisition of PIF-dependent shade-signal responsiveness.
Accomplishments
