Biology subjects

Hinckley, W. E.

Publications and source records attributed to Hinckley, W. E..

6 recordsLinked to original sources

bioRxiv2025

HHO5: A key orchestrator of dose-dependent nitrogen signaling pathways in Arabidopsis

A major goal in agriculture is to engineer crops that can maintain yield with less nitrogen (N) fertilizer input. Major orchestrators of plant responses to N include members of the HRS1 HOMOLOG (HHO) family of transcription factors (TFs). However, HHO TFs have been difficult targets for functional studies in planta due to their redundancy. Here, we highlight a unique role for a phylogenetically diverged HHO TF, HHO5, whose expression is regulated in an N-dose dependent fashion and is specifically expressed in phloem. We found that an HHO5 single mutant displays significant misregulation of N-dose dependent genes and plant growth rates. HHO5 is also unique as it displays a dual activator/repressor activity on N-dose dependent gene regulation. HHO5 specifically acts as a direct gene repressor when binding DNA targets. In contrast, genes activated by HHO5 include indirect targets regulated by TFs downstream of HHO5 (TF2s). To validate the influence of HHO5 via its direct TF2s, we used validated TF2 data to build a gene regulatory network that links HHO5-TF2 targets to [~]70% of the N-dose genes regulated by HHO5 in planta. By these means, we define HHO5 as a novel dual activator/repressor of plant N-dose signaling. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=110 SRC="FIGDIR/small/667803v1_ufig1.gif" ALT="Figure 1"> View larger version (46K): org.highwire.dtl.DTLVardef@11adaa9org.highwire.dtl.DTLVardef@9c221org.highwire.dtl.DTLVardef@a80eceorg.highwire.dtl.DTLVardef@163fcdf_HPS_FORMAT_FIGEXP M_FIG C_FIG

plant biology↗

bioRxiv2025

Transcription Factor (TF) validation using Dam-IT simultaneously captures genome-wide TF-DNA binding, direct gene regulation, and chromatin accessibility in plant cells

Transcription Factors (TFs) govern vast networks of gene regulation. However, TF-DNA binding and TF-gene regulation datasets are typically measured separately due to experimental constraints, making it challenging to disentangle true biological relationships from batch effects. To fill this gap, we developed DamID-seq Incorporating Transcriptomics (Dam-IT), which simultaneously captures TF-DNA binding, direct TF-gene regulation, and chromatin accessibility in the same batch of cells. Dam-IT uses a transient cell-based TF-target validation system that is scalable and flexible to many experimental designs. As proof of concept, we used Dam-IT to reveal that bZIP1 directly regulates genes by binding to DNA regions of relatively low chromatin accessibility, supporting a "Hit-and-Run" mechanism of transcription.

systems biology↗

bioRxiv2022

Uncovering the "ZIP code" for bZIP dimers reveals novel motifs, regulatory rules and one billion years of cis-element evolution

Many eukaryotic transcription factors (TF) form homodimer or heterodimer complexes to regulate gene expression. For example, dimerization properties of the basic leucine zipper (bZIP) family play a critical role in regulating the unique biological functions in all eukaryotes. However, the molecular mechanism underlying the binding sequence and functional specificity of homo- versus heterodimers remains elusive. To fill this gap, we developed a double DNA Affinity Purification sequencing (dDAP-seq) technique that maps heterodimer DNA binding sites in an endogenous genome context. Our genome-wide binding profiles of twenty pairs of C/S1 bZIP heterodimers and S1 homodimers in Arabidopsis revealed that heterodimerization significantly expands the DNA binding preferences of bZIP TFs. Analysis of the heterodimer target genes in stress response and development suggest heterodimerization gives rise to regulatory responses that are distinct from the homodimers. In addition to the classic ACGT elements recognized by plant bZIPs, we found that the C/S1 heterodimers bound to motifs that might share an origin with the GCN4 cis-elements in yeast that diverged from plants more than one billion years ago. Importantly, heterodimer binding specificities can be distinguished by their relative preference for ACGT motifs versus GCN4-related motifs. More broadly, our study demonstrates the potential of dDAP-seq in deciphering the DNA binding specificities of interacting TFs that are key for combinatorial gene regulation.

molecular biology↗

bioRxiv2021

Regulatory start-stop elements in 5' untranslated regions pervasively modulate translation

Sequence elements within the 5 untranslated region (UTR) of eukaryotic genes, e.g. upstream open reading frames (uORFs), control translation of eukaryotic genes. We describe an element consisting of a start codon immediately followed by a stop codon which is distinct from uORFs in the lack of an elongation step. Start-stops have been described for specific cases, but their widespread impact has been overlooked. Start-stop elements occur in the 5UTR of 1, 417 human genes and are more often occupied with a ribosome than canonical uORFs or control sequences. Start-stops efficiently halt ribosomes without evidence for accelerated RNA turnover, therefore acting as a barrier for the scanning of the small ribosomal subunit and repressing downstream translation. Our results suggest a model by which the ribosome undergoes repeated cycles of termination and partial ribosomal recycling, during which the large subunit detaches, but the 40S subunit with the Met-tRNAiMet remains associated with the mRNA to be rejoined by the 60S subunit. Start-stop elements occur in many transcription factors and signaling genes, and affect cellular fate via different routes. We investigate the start-stop element in several genes, i.e. MORF4L1, SLC39A1, and PSPC1, and in more detail in ATF4.

systems biology↗

bioRxiv2020

Gene Expression Changes Occurring at Bolting Time are Associated with Leaf Senescence in Arabidopsis

In plants, the vegetative to reproductive phase transition (termed bolting in Arabidopsis) generally precedes age-dependent leaf senescence (LS). Many studies describe a temporal link between bolting time and LS, as plants that bolt early, senesce early, and plants that bolt late, senesce late. However, the molecular mechanisms underlying this relationship are unknown and are potentially agriculturally important, as they may allow for the development of crops that can overcome early LS caused by stress-related early phase transition. We hypothesized that gene expression changes associated with bolting time were regulating LS. We used a mutant that displays both early bolting and early LS as a model to test this hypothesis. An RNA-seq time series experiment was completed to compare the early bolting mutant to vegetative WT plants of the same age. This allowed us to identify bolting time-associated genes (BAGs) expressed in an older rosette leaf at the time of inflorescence emergence. The BAG list contains many well characterized LS regulators (ORE1, WRKY45, NAP, WRKY28), and GO analysis revealed enrichment for LS and LS-related processes. These bolting associated LS regulators likely contribute to the temporal coupling of bolting time to LS.

plant biology↗

bioRxiv2019

The HAC1 Histone Acetyltransferase Promotes Leaf Senescence via Regulation of ERF022

Nutrient remobilization during leaf senescence nourishes the growing plant. Understanding the regulation of this process is essential for reducing our dependence on nitrogen fertilizers and increasing agricultural sustainability. Our lab is interested in chromatin changes that accompany the transition to leaf senescence. Previously, darker green leaves were reported for Arabidopsis thaliana hac1 mutants, defective in a gene encoding a histone acetyltransferase in the CREB-binding protein family. Here, we show that two Arabidopsis hac1 alleles display delayed age-related developmental senescence, but have normal dark-induced senescence. Using a combination of ChIP-seq for H3K9ac and RNA-seq for gene expression, we identified 44 potential HAC1 targets during age-related developmental senescence. Genetic analysis demonstrated that one of these potential targets, ERF022, is a positive regulator of leaf senescence. ERF022 is regulated additively by HAC1 and MED25, suggesting MED25 recruits HAC1 to the ERF022 promoter to increase its expression in older leaves.

plant biology↗