Biology subjects

Wengier, D. L.

Publications and source records attributed to Wengier, D. L..

6 recordsLinked to original sources

bioRxiv2024

Zinc-starved Brassicaceae Plants Secrete Peptides thatInduce Root Expansion

Zinc (Zn) deficiency is recognized as a global crisis as it is observed in half of all agricultural soils. However, the molecular mechanisms that drive plant physiological responses to soil Zn deficiency are not well understood. We used an untargeted metabolomics approach to search for metabolites exuded from roots during Zn deficiency stress, which led to the discovery of a collection of secreted small defensin-like peptides in Arabidopsis thaliana (named Zinc-Deficiency Responsive Peptides (ZDRPs)). Phylogenetic analysis and untargeted metabolomics revealed ZDRPs in at least eleven accessions of A. thaliana and nine members of the Brassicaceae family. Analysis of Arabidopsis gene mutants and overexpressing lines, in combination with chemical complementation experiments, unveiled a critical role of these peptides in plant root growth. We hypothesize that Brassicaceae secreted peptides enable plants to expand their root mass to reach Zn-rich soil layers and optimize Zn uptake. These data reveal a critical relationship between plant survival, Zn status, root morphology and peptide production. Taken together, our results expand our knowledge regarding micronutrient deficiency responses in plants and could enable in engineering approaches to make plants more resilient to low Zn conditions. SignificanceZinc deficiency is the most abundant micronutrient deficiency affecting about 50% of arable lands thus presenting a high burden for plant health and agriculture globally. In this study, we reveal a metabolic strategy by Brassicaceae to deal with low Zn concentrations. We characterize the role of peptides expressed upon zinc deficiency in a variety of important crop plants. The discovery of a cryptic class of peptides that are made by plant roots specifically suffering from Zn deficiency provides critical insight into the molecular mechanisms by which plants dynamically acclimate to nutrient-limited soils. The identification of peptides actively secreted by zinc-deprived plants has translational value for sustainable agriculture, human health, and bioengineering approaches to enable tolerance to low zinc.

plant biology↗

bioRxiv2023

RADIP LIGHT-INDUCED PHOSPHORYLATION CHANGES IN MICROTUBULE RELATED PROTEINS IN ARABIDOPSIS

Rapid hypocotyl elongation allows buried seedlings to reach the surface, where light triggers de-etiolation and inhibits hypocotyl growth mainly by phytochromes A, B and cryptochromes 1, 2. Dynamic phosphorylation/dephosphorylation events provide a mechanism to rapidly transduce light signals. Only recently we have begun to uncover the earliest phospho-signaling responders to light. Here, we report a large-scale phosphoproteomic analysis and identify 20 proteins that change their phosphorylation pattern after 20 min of white light pulse compared to darkness. Microtubule-associated proteins (MAPs) were highly overrepresented in this group. Among them, we studied CIP7 (COP1-INTERACTING-PROTEIN-7), which presented microtubule (MT) localization, in contrast to what was previously described. Phosphorylated isoform in Serine 915 (Sp915) of CIP7 was detected in etiolated seedlings but undetectable after a light pulse in the presence of photoreceptors, while its expression decays with long light exposure. The short hypocotyl phenotype and rearrangement of MTs in etiolated cip7 mutants are complemented by CIP7-YFP and the phospho-mimetic CIP7S915D-YFP, but not the phospho-null CIP7S915A-YFP suggesting that Sp915CIP7 is the active isoform that promotes hypocotyl elongation thorough MT reorganisation in darkness. Our results reveal that the small repertory of proteins that changes the phosphorylation status after a rapid light signal is tightly focused on MAPs; suggesting that phospho-regulation of microtubule-base processes are early targets during de-etiolation. The evidence on Sp915CIP7 supports this idea.

plant biology↗

bioRxiv2022

Arabidopsis pollen Prolyl-hydroxylases P4H4/6 are required for correct hydroxylation and secretion of LRX11 in pollen tubes

Major constituents of the plant cell walls are structural proteins that belong to the Hydroxyproline-rich glycoprotein family. Leucine-rich repeat extensis are contain a leucine-rich domain and a C-terminal domain with repetitive Ser-Pro(3-5) motifs plausible to be glycosylated. We have demonstrated that pollen-specific LRX8-11 from Arabidopsis thaliana are necessary to maintain the integrity of the pollen tube cell wall during polarized growth. In classical EXTs and likely in LRXs, proline residues are converted to hydroxyproline by Prolyl-4-hydroxylases, thus defining novel O-glycosylation sites. In this context, we aimed to determine whether hydroxylation and subsequent O-glycosylation of Arabidopsis pollen LRXs are necessary for their proper function and cell wall localization in pollen tubes. We hypothesized that pollen-expressed P4H4 and P4H6 catalyze the hydroxylation of the proline units present in Ser-Pro(3-5) motifs of LRX8-LRX11. Here, we show the p4h4-1 p4h6-1 double mutant exhibits a significant reduction in pollen germination rates and a slight reduction in pollen tube length. Pollen germination is also inhibited by specific P4Hs inhibitors, suggesting that prolyl hydroxylation is required for pollen tube development. Plants expressing pLRX11::LRX11-GFP in the p4h4-1 p4h6-1 background show partial relocalization of LRX11-GFP from the pollen tube tip apoplast to the cytoplasm. Finally, IP-MS- MS analysis revealed a decrease in oxidized prolines in LRX11-GFP in the p4h4-1 p4h6-1 background when compared to lrx11 plants expressing pLRX11::LRX11-GFP. Together, these results suggest that P4H4 and P4H6 are required for pollen germination and are also involved in LRX11 hydroxylation necessary for its localization at the cell wall of pollen tubes. One Sentence SummaryPollen-expressed P4H4 and P4H6 are required for pollen germination and for proper hydroxylation and secretion of LRX11 in pollen tubes.

plant biology↗

bioRxiv2022

Modular and programmable RNA sensing using ADAR editing in living cells

With the increasing availability of single-cell transcriptomes, RNA signatures offer a promising basis for targeting living cells. Molecular RNA sensors would enable the study of and therapeutic interventions for specific cell types/stats in diverse contexts, particularly in human patients and non-model organisms. Here we describe a modular and programmable design for live RNA sensing using adenosine deaminases acting on RNA (RADAR). We validated and then expanded our basic design, characterized its performance, and thoroughly analyzed its compatibility with the human/mouse transcriptomes. We also identified strategies to further boost output levels and improve the dynamic range. We show that RADAR is programmable and modular, and uniquely enables compact AND logic. In addition to being quantitative, RADAR can distinguish disease-relevant point mutations. Finally, we demonstrate that RADAR is a self-contained system with the potential to function in diverse organisms.

bioengineering↗

bioRxiv2020

Synaptic contributions to cochlear outer hair cell Ca2+ homeostasis

For normal cochlear function, outer hair cells (OHCs) require a precise regulation of intracellular Ca2+ levels. Influx of Ca2+ occurs both at the stereocillia tips and through the basolateral membrane. In this latter compartment, two different origins for Ca2+ influx have been poorly explored: voltage-gated Ca2+ channels (VGCC) at synapses with type II afferent neurons, and 910 cholinergic nicotinic receptors at synapses with medio-olivochlear complex (MOC) neurons. Using functional imaging in rodent OHCs, we report that these two Ca2+ entry sites are closely positioned, but present different regulation mechanisms. Ca2+ spread from MOC synapses is contained by cisternal Ca2+-ATPases. Considered a weak drive for transmitter release, we unexpectedly found that VGCC Ca2+ signals are comparable in size to those elicited by 910 and can be potentiated by ryanodine receptors. Finally, we showed that sorcin, a highly expressed gene product in OHCs with reported Ca2+ control function in cardiomy-ocytes, regulates basal Ca2+ levels and MOC synaptic activity in OHCs.

neuroscience↗

bioRxiv2017

Dissection of MAPK signaling specificity through protein engineering in a developmental context

Mitogen-activated protein kinases (MAPK) signaling affects many processes, some of which have different outcomes in the same cell. In Arabidopsis, activation of a MAPK cascade consisting of YODA, MKK4/5 and MPK3/6 inhibits early stages of stomatal developmental, but this ability is lost at the latest stage when guard mother cells (GMCs) transition to guard cells (GCs). Rather than downregulating cascade components, stomatal precursors must have a mechanism to prevent late stage inhibition because the same MKKs and MPKs mediate other physiological responses. Here, we artificially activated the MAPK cascade using MKK7, another MKK that can modulate stomatal development, and found that inhibition of stomatal development is still possible in GMCs. This suggests that MKK4/5, but not MKK7, are specifically prevented from inhibiting stomatal development. To identify regions of MKKs responsible for cell-type specific regulation, we used a domain swap approach with MKK7 and a battery of in vitro and in vivo kinase assays. We found that N-terminal regions of MKK5 and MKK7 establish specific signal-to-output connections like they do in other organisms, but they do so in combination with previously undescribed modules in the C-terminus. One of these modules encodes the GMC-specific regulation of MKK5, that when swapped with MKK7s, allows MKK5 to mediate robust inhibition of late stomatal development. Because MKK structure is conserved across species, the identification of new MKK specificity modules and signaling rules furthers our understanding of how eukaryotes create specificity in complex biological systems.

plant biology↗