Biology subjects

Groth, G.

Publications and source records attributed to Groth, G..

6 recordsLinked to original sources

bioRxiv2026

Structural Mapping of the EIN2-EIN3 Interaction Core and Its Integration with ENAP1 in Ethylene Signaling

Ethylene regulates diverse developmental processes, yet the molecular function of its central regulator, ETHYLENE INSENSITIVE 2 (EIN2), has remained unclear. Although EIN2 nuclear import is mediated by the Importin-/{beta} pathway, the molecular events initiated by EIN2 after nuclear entry were unknown. Here we show that EIN2 directly engages the transcription factor EIN3, establishing a mechanistic link between EIN2 nuclear accumulation and transcriptional activity. Microscale thermophoresis, yeast two-hybrid analysis and in planta FLIM-FRET consistently support this interaction. Domain mapping identifies EIN3 residues 86-173 as the core EIN2-binding region, and structural modeling refines the interface to a conserved segment within residues 86-120 that contacts a conserved region near the N-terminus of the EIN2-CEND fragment. In planta, EIN2 residues 1042-1214 are sufficient for EIN3 binding, revealing multiple interaction-competent surfaces with distinct affinities. The chromatin-associated protein ENAP1 also binds EIN2 and competes with EIN3, indicating a dynamic, concentration-dependent regulatory mechanism rather than a stable ternary complex. These findings define the molecular basis of the EIN2-EIN3 interaction and provide a mechanistic framework for EIN2-dependent transcriptional control in ethylene signaling.

plant biology↗

bioRxiv2025

Molecular Mechanism and Structural Models ofProtein-Mediated Copper Transfer to the Arabidopsisthaliana Ethylene Receptor ETR1 at the ER Membrane

In plants, the gaseous plant hormone ethylene regulates a wide range of developmental processes and stress responses. The small unsaturated hydrocarbon is detected by a family of receptors (ETRs) located in the membrane of the endoplasmic reticulum, which rely on a monovalent copper cofactor to detect this hydrocarbon. The copper-transporting P-type ATPase RAN1 (HMA7), located in the same membrane, is known to be essential for the biogenesis of ETRs. Still, the precise molecular mechanism by which the receptors acquire their copper cofactor remains unclear. A recent study by our laboratory demonstrated a direct interaction between RAN1 and soluble copper chaperones of the ATX1 family with the model ethylene receptor ETR1, providing initial insights into the mechanism by which copper is transferred from the cytosol to the membrane-bound receptors. In this study, we further investigated these interactions with respect to the function of individual domains in complex formation. To this end, we combined biochemical experiments and computational predictions and unraveled the processes and mechanisms by which copper is transferred to ETR1 at the molecular level.

plant biology↗

bioRxiv2023

A peptide that regulates metalation of the Arabidopsis ethylene receptor.

Ethylene signalling represents one of the classic hormonal pathways in plants, with diverse roles in development and stress responses. The dimeric ethylene receptor localizes to the endoplasmic reticulum (ER) and contains Cu(I) ions essential for ethylene binding and signal transduction. We previously discovered that mutants in the Arabidopsis gene POLARIS (PLS), encoding a 36 amino acid peptide, exhibit enhanced ethylene signalling responses, suggestive of reduced receptor activity, but the role and activity of the peptide in this signalling cascade has not been defined. Here we report PLS binds copper as a 1:2 thiol-dependent Cu(I):PLS2 complex, with an affinity of 3.79 ({+/-}1.5) x1019 M-2, via two cysteine residues also found in the related species Camelina sativa. These residues are also essential for biological function. This affinity precludes PLS as a cytosolic Cu chaperone. We demonstrate that PLS localizes to endomembranes and interacts with the transmembrane domain of receptor protein ETR1. PLS-ETR1 binding is increased in the presence of copper, and this interaction provides a Cu-dependent mechanism for mediating a repression of ethylene responses. PLS transcription is up-regulated by auxin and down-regulated by ethylene, and so PLS-ETR1 interactions also provide a mechanism to modulate ethylene responses in high auxin tissues.

plant biology↗

bioRxiv2022

Spectroscopic and QM/MM studies of the Cu(I) binding site of the plant ethylene receptor ETR1

Herein, we present the first spectroscopic characterization of the Cu(I) active site of the plant ethylene receptor ETR1. The X-ray absorption (XAS) and extended X-ray absorption fine structure (EXAFS) spectroscopy presented here establish that ETR1 has a low-coordinate Cu(I) site. The EXAFS resolves a mixed first coordination sphere of N/O and S scatterers at distances consistent with potential histidine and cysteine residues. This finding agrees with the coordination of residues C65 and H69 to the Cu(I) site, which are critical for ethylene activity and well-conserved. Further, the Cu K-edge XAS and EXAFS of ETR1 exhibit spectroscopic changes upon addition of ethylene that are attributed to modifications in the Cu(I) coordination environment, suggestive of ethylene binding. Results from umbrella sampling simulations of the proposed ethylene binding helix of ETR1 at a mixed quantum mechanics/molecular mechanics (QM/MM) level agree with the EXAFS fit distance changes upon ethylene binding, particularly in the increase of the distance between H69 and Cu(I), and yield binding energetics comparable to experimental dissociation constants. The observed changes in the copper coordination environment might be the triggering signal for the transmission of the ethylene response.

biophysics↗

bioRxiv2021

Biophysical and pharmacokinetic characterization of a small-molecule inhibitor of RUNX1/ETO tetramerization with anti-leukemic effects

Acute myeloid leukemia (AML) is a malignant disease of immature myeloid cells and the most prevalent acute leukemia among adults. The oncogenic homo-tetrameric fusion protein RUNX1/ETO results from the chromosomal translocation t(8;21) and is found in AML patients. The nervy homology region 2 (NHR2) domain of ETO mediates tetramerization; this oligomerization is essential for oncogenic activity. Previously, we identified the first-in-class small-molecule inhibitor of NHR2 tetramer formation, 7.44, which was shown to specifically interfere with NHR2, restore gene expression down-regulated by RUNX1/ETO, inhibit the proliferation of RUNX1/ETO-depending SKNO-1 cells, and reduce the RUNX1/ETO-related tumor growth in a mouse model. However, no biophysical and structural characterization of 7.44 binding to the NHR2 domain has been reported. Likewise, the compound has not been characterized as to physicochemical, pharmacokinetic, and toxicological properties. Here, we characterize the interaction between the NHR2 domain of RUNX1/ETO and 7.44 by biophysical assays and show that 7.44 interferes with NHR2 tetramer stability and leads to an increase in the dimer population of NHR2. The affinity of 7.44 with respect to binding to NHR2 is Klig = 3.95 {+/-} 1.28 M. By NMR spectroscopy combined with molecular dynamics simulations, we show that 7.44 binds with both heteroaromatic moieties to NHR2 and interacts with or leads to conformational changes in the N-termini of the NHR2 tetramer. Finally, we demonstrate that 7.44 has favorable physicochemical, pharmacokinetic, and toxicological properties. Together with biochemical, cellular, and in vivo assessments, the results reveal 7.44 as a lead for further optimization towards targeted therapy of t(8;21) AML.

biophysics↗

bioRxiv2019

A peptide pair coordinates regular ovule initiation patterns with seed number and fruit size

Ovule development in Arabidopsis thaliana involves pattern formation which ensures that ovules are regularly arranged in the pistils to reduce competition for nutrients and space. Mechanisms underlying pattern formation in plants, such as phyllotaxis, flower morphogenesis or lateral root initiation, have been extensively studied, and genes controlling the initiation of ovules have been identified. However, how a regular spacing of ovules is achieved is not known. Using natural variation analysis combined with quantitative trait locus analysis, we found that the spacing of ovules in the developing fruits is controlled by two secreted peptides, EPFL2 and EPFL9 (also known as Stomagen), and their receptors from the ERECTA (ER) family that act from the carpel wall and the placental tissue. We found that a signalling pathway controlled by EPFL9 acting from the carpel wall through the LRR-receptor kinases ER, ERL1 and ERL2 promotes fruit growth. Regular spacing of ovules depends on EPFL2 expression in the carpel wall and in the inter-ovule spaces, where it acts through ERL1 and ERL2. Loss of EPFL2 signalling results in shorter fruits and irregular spacing of ovules or even ovule twinning. The EPFL2 expression pattern between ovules is under negative-feedback regulation by auxin, which accumulates in the arising ovule primordia. We propose that the auxin-EPFL2 signalling module evolved to control the initiation and regular, equidistant spacing of ovule primordia, which serves to minimise competition between developing seeds. Together, EPFL2 and EPFL9 coordinate ovule patterning and thereby seed number with fruit growth through a set of shared receptors.

plant biology↗