skip to main content


Title: Integrated omics networks reveal the temporal signaling events of brassinosteroid response in Arabidopsis
Abstract

Brassinosteroids (BRs) are plant steroid hormones that regulate cell division and stress response. Here we use a systems biology approach to integrate multi-omic datasets and unravel the molecular signaling events of BR response inArabidopsis. We profile the levels of 26,669 transcripts, 9,533 protein groups, and 26,617 phosphorylation sites fromArabidopsisseedlings treated with brassinolide (BL) for six different lengths of time. We then construct a network inference pipeline called Spatiotemporal Clustering and Inference of Omics Networks (SC-ION) to integrate these data. We use our network predictions to identify putative phosphorylation sites on BES1 and experimentally validate their importance. Additionally, we identify BRONTOSAURUS (BRON) as a transcription factor that regulates cell division, and we show thatBRONexpression is modulated by BR-responsive kinases and transcription factors. This work demonstrates the power of integrative network analysis applied to multi-omic data and provides fundamental insights into the molecular signaling events occurring during BR response.

 
more » « less
Award ID(s):
1818160
NSF-PAR ID:
10304750
Author(s) / Creator(s):
; ; ; ; ; ; ; ; ;
Publisher / Repository:
Nature Publishing Group
Date Published:
Journal Name:
Nature Communications
Volume:
12
Issue:
1
ISSN:
2041-1723
Format(s):
Medium: X
Sponsoring Org:
National Science Foundation
More Like this
  1. Brassinosteroids (BR) and Target of Rapamycin Complex (TORC) are two major actors coordinating plant growth and stress responses. BRs function through a signaling pathway to extensively regulate gene expression and TORC is known to regulate translation and autophagy. Recent studies have revealed connections between these two pathways, but a system-wide view of their interplay is still missing. • We quantified the level of 23,975 transcripts, 11,183 proteins, and 27,887phosphorylation sites in wild-type Arabidopsis thalianaand inmutants with altered levels of either BRASSINOSTEROID INSENSITIVE 2 (B IN2) or REGULATORY ASSOCIATED PROTEIN OF TOR 1B (RAPTOR1B), two key players in BR and TORC signaling, respectively.• We found that perturbation of BIN2 or RAPTOR1B levels affects a common set of gene-products involved in growth and stress responses. Furthermore, we used the multi-omic data to reconstruct an integrated signaling network. We screened 41candidate genes identified from the reconstructed network and found that loss of function mutants of many of these proteins led to an altered BR response and/or modulated autophagy activity.• Altogether, these results establish a predictive network that defines different layers of molecular interactions between BR- or TORC-regulated growth and autophagy. 
    more » « less
  2. Abstract

    The scarcity of accessible sites that are dynamic or cell type-specific in plants may be due in part to tissue heterogeneity in bulk studies. To assess the effects of tissue heterogeneity, we apply single-cell ATAC-seq toArabidopsis thalianaroots and identify thousands of differentially accessible sites, sufficient to resolve all major cell types of the root. We find that the entirety of a cell’s regulatory landscape and its transcriptome independently capture cell type identity. We leverage this shared information on cell identity to integrate accessibility and transcriptome data to characterize developmental progression, endoreduplication and cell division. We further use the combined data to characterize cell type-specific motif enrichments of transcription factor families and link the expression of family members to changing accessibility at specific loci, resolving direct and indirect effects that shape expression. Our approach provides an analytical framework to infer the gene regulatory networks that execute plant development.

     
    more » « less
  3. Abstract

    Plant steroid hormones brassinosteroids (BRs) regulate plant growth and development at many different levels. Recent research has revealed that stress‐responsive NAC (petunia NAM and Arabidopsis ATAF1, ATAF2, and CUC2) transcription factorRD26 is regulated byBRsignaling and antagonizesBES1 in the interaction between growth and drought stress signaling. However, the upstream signaling transduction components that activateRD26 during drought are still unknown. Here, we demonstrate that the function ofRD26 is modulated byGSK3‐like kinaseBIN2 and protein phosphatase 2CABI1. We show thatABI1, a negative regulator inabscisic acid (ABA)signaling, dephosphorylates and destabilizesBIN2 to inhibitBIN2 kinase activity.RD26 protein is stabilized byABAand dehydration in aBIN2‐dependent manner.BIN2 directly interacts and phosphorylatesRD26in vitroandin vivo.BIN2 phosphorylation ofRD26 is required forRD26 transcriptional activation on drought‐responsive genes.RD26 overexpression suppressed the brassinazole (BRZ)  insensitivity ofBIN2 triple mutantbin2 bil1 bil2, andBIN2 function is required for the drought tolerance ofRD26 overexpression plants. Taken together, our data suggest a drought signaling mechanism in which drought stress relievesABI1 inhibition ofBIN2, allowingBIN2 activation. Sequentially,BIN2 phosphorylates and stabilizesRD26 to promote drought stress response.

     
    more » « less
  4. Abstract

    Molecular biology aims to understand cellular responses and regulatory dynamics in complex biological systems. However, these studies remain challenging in non-model species due to poor functional annotation of regulatory proteins. To overcome this limitation, we develop a multi-layer neural network that determines protein functionality directly from the protein sequence. We annotate kinases and phosphatases inGlycine max. We use the functional annotations from our neural network, Bayesian inference principles, and high resolution phosphoproteomics to infer phosphorylation signaling cascades in soybean exposed to cold, and identify Glyma.10G173000 (TOI5) and Glyma.19G007300 (TOT3) as key temperature regulators. Importantly, the signaling cascade inference does not rely upon known kinase motifs or interaction data, enabling de novo identification of kinase-substrate interactions. Conclusively, our neural network shows generalization and scalability, as such we extend our predictions toOryza sativa,Zea mays,Sorghum bicolor, andTriticum aestivum. Taken together, we develop a signaling inference approach for non-model species leveraging our predicted kinases and phosphatases.

     
    more » « less
  5. null (Ed.)
    Diatoms are major contributors to global primary production and their populations in the modern oceans are affected by availability of iron, nitrogen, phosphate, silica, and other trace metals, vitamins, and infochemicals. However, little is known about the role of phosphorylation in diatoms and its role in regulation and signaling. We report a total of 2759 phosphorylation sites on 1502 proteins detected in Phaeodactylum tricornutum. Conditionally phosphorylated peptides were detected at low iron (n = 108), during the diel cycle (n = 149), and due to nitrogen availability (n = 137). Through a multi-omic comparison of transcript, protein, phosphorylation, and protein homology, we identify numerous proteins and key cellular processes that are likely under control of phospho-regulation. We show that phosphorylation regulates: (1) carbon retrenchment and reallocation during growth under low iron, (2) carbon flux towards lipid biosynthesis after the lights turn on, (3) coordination of transcription and translation over the diel cycle and (4) in response to nitrogen depletion. We also uncover phosphorylation sites for proteins that play major roles in diatom Fe sensing and utilization, including flavodoxin and phytotransferrin (ISIP2A), as well as identify phospho-regulated stress proteins and kinases. These findings provide much needed insight into the roles of protein phosphorylation in diel cycling and nutrient sensing in diatoms. 
    more » « less