Biology subjects

Heilmann, I.

Publications and source records attributed to Heilmann, I..

7 recordsLinked to original sources

bioRxiv2026

Dynamic Patterns of Nuclear Transcription Factor Abundance in Plant Basal Immunity Revealed by Spatial Proteomics of Arabidopsis Nuclei

O_LIThe control of amount of nuclear proteins is fundamental in regulating plant gene expression, but the mechanisms of quantitative dynamics of the nuclear proteome are largely unstudied during adaptive responses to pathogens. C_LIO_LIHighly specific labeling, enrichment and measurement of the nuclear proteome was performed using TurboID LC-MS of Arabidopsis thaliana leaves treated with the pathogen-associated molecular pattern (PAMP), flg22, and/or cycloheximide. The chosen experimental approach allowed discrimination of the effects of translation, nuclear protein import, trafficking of preexisting proteins, derepression, and nuclear protein turn-over upon elicitation of basal immunity. C_LIO_LIThe highly specific, deep coverage of proteins in the nucleus makes this study a resource for anyone interested in plant nuclear proteome dynamics and defense. C_LIO_LIAround 2,000 nuclear proteins were repeatedly quantified, including more than 300 transcription factors or other proteins related to transcription. Several proteins with documented activity in endosomes were newly synthesized and imported into nuclei upon PAMP challenge, suggesting alternative nuclear functions in PAMP-triggered immunity (PTI). Circadian clock components, including the transcription factor, CIRCADIAN CLOCK ASSOCIATED 1 (CCA1)-HIKING EXPEDITION (CHE), were depleted upon PAMP challenge, suggesting a safeguard against untimely induction of systemic acquired resistance (SAR). C_LIO_LIBased on proteomic patterns, proteins moonlighting in the nucleus as well as trafficking and turn-over regulation of the proteome are common elements during plant immunity. C_LI

plant biology↗

bioRxiv2024

Barley resistance and susceptibility to fungal cell entry involve the interplay of ROP signaling with phosphatidylinositol-monophosphates

Rho-of-plant small GTPases (ROPs) are regulators of plant polar growth and of plant-pathogen interactions. The barley ROP, RACB, is involved in susceptibility towards infection by the barley powdery mildew fungus Blumeria hordei (Bh) but little is known about the cellular pathways that connect RACB-signaling to disease susceptibility. Here we identify novel RACB interaction partners of plant or fungal origin by untargeted co-immunoprecipitation of constitutively active (CA) RACB tagged by green-fluorescent protein from Bh-infected barley epidermal layers and subsequent analysis by liquid chromatography-coupled mass spectrometry. Three of immunoprecipitated proteins, a plant phosphoinositide phosphatase, a plant phosphoinositide phospholipase and a putative Bh-effector protein are involved in the barley-Bh-pathosystem and support disease resistance or susceptibility, respectively. RACB and its plant interactors bind to overlapping anionic phospholipid species in vitro, and in case of RACB, this lipid-interaction is mediated by its carboxy-terminal polybasic region (PBR). Fluorescent markers for anionic phospholipids show altered subcellular distribution in barley cells during Bh-attack and under expression of a RACB-binding fungal effector. Phosphatidylinositol 4-phosphate, phosphatidylinositol 3,5-bisphosphate and phosphatidylserine show a distinct enrichment at the haustorial neck region, suggesting a connection to subcellular targeting of RACB at this site. The interplay of ROPs with anionic phospholipids and phospholipid-metabolizing enzymes may thus enable the subcellular enrichment of components pivotal for success or failure of fungal penetration.

plant biology↗

bioRxiv2023

Inter-Organellar Effects of Defective ER-localized Linolenic Acid Formation on Thylakoid Lipid Composition and Xanthophyll-Cycle Pigment De-epoxidation in the Arabidopsis fad3 mutant

Monogalactosyldiacylglycerol (MGDG) is the main lipid constituent of thylakoids and a structural component of photosystems and photosynthesis-related proteo-lipid complexes in green tissues. Previously reported changes in MGDG abundance upon stress-treatments are hypothesized to reflect mobilization of MGDG-based polyunsaturated lipid intermediates to maintain extraplastidial membrane integrity. While exchange of lipid intermediates between compartmental membranes is well documented, physiological consequences of mobilizing an essential thylakoid lipid, such as MGDG, for an alternative purpose are not well understood.Arabidopsis seedlings exposed to mild (50 mM) salt-treatment displayed significantly increased abundance of both MGDG and the extraplastidial lipid, phosphatidylcholine (PC). Interestingly, similar increases in MGDG and PC were observed in Arabidopsis fad3 mutant seedlings defective in ER-localized linolenic acid formation, in which compensatory plastid-to-ER-directed mobilization of linolenic acid-containing intermediates takes place. The postulated (salt) or evident (fad3) plastid-ER-exchange of intermediates concurred with altered thylakoid function according to parameters of photosynthetic performance. While salt-treatment of wild type seedlings inhibited photosynthetic parameters in a dose-dependent manner, interestingly the fad3 mutant did not show overall reduced photosynthetic quantum yield. By contrast, we observed a reduction specifically of non-photochemical quenching (NPQ) under high light, representing only part of observed salt effects. The decreased NPQ in the fad3 mutant was accompanied by reduced activity of the xanthophyll cycle, leading to a reduced concentration of the NPQ-effective pigment zeaxanthin. The findings suggest that altered ER-located fatty acid unsaturation and ensuing inter-organellar compensation impacts on aspects of thylakoids related to the function of specific enzymes, rather than globally affecting thylakoid function. Subject Areas(2) Environmental and stress responses (7) Membrane and transport

plant biology↗

bioRxiv2022

AI-guided cryo-EM probes a thermophilic cell-free system with succinyl-coA manufacturing capability

Cell-free systems display tremendous potential for biotechnological applications, complementing in vitro reconstituted enzymatic processes and traditional expression systems. However, they often represent "black boxes" without much insight into their components. Here, we characterize a thermophilic cell-free system that produces succinyl-CoA and discern its intrinsic, non-stochastic organization. By employing biochemical, biophysical, and bioinformatic methods we resolve its molecular composition, 3D architecture and molecular function at atomic resolution. We further report the high-resolution cryo-EM structure of the reactions main component, the oxoglutarate dehydrogenase complex core (E2o), which displays various structural adaptations. These include hydrogen bonding patterns confining interactions of participating enzymes (E1o-E2o-E3), electrostatic tunneling that drives inter-communication between subunits, and the presence of a flexible subunit, the E3BPo connecting E2o and E3. This multi-scale analysis of a cell-free system provides a blueprint for structure-function studies of complex mixtures of biotechnological value.

biochemistry↗

bioRxiv2022

First direct observation of 'elongated' conformational states in α-synuclein upon liquid-liquid phase separation

-Synuclein (-syn) is an intrinsically disordered protein (IDP) that undergoes liquid-liquid phase separation (LLPS), fibrillation, and forms insoluble intracellular Lewys bodies in neurons, which are the hallmark of Parkinsons Disease (PD). Neurotoxicity precedes the formation of aggregates and is probably related to LLPS of -syn in the cell. The molecular mechanisms underlying the early stages of LLPS are still elusive. To obtain structural insights into -syn upon LLPS, we take advantage of cross-linking/mass spectrometry (XL-MS) and introduce an innovative approach, termed COMPASS (COMPetitive PAiring StatisticS). COMPASS unravels transient interactions between -syn molecules in liquid droplets. In this work, we show that the conformational ensemble of -syn shifts from a hairpin-like structure towards more elongated conformational states upon LLPS. We obtain insights into the critical initial stages of PD and establish a novel mass spectrometry-based approach that will aid to solve open questions in LLPS structural biology.

biochemistry↗

bioRxiv2020

Plasma membrane nano-organization specifies phosphoinositide effects on Rho-GTPases and actin dynamics in tobacco pollen tubes

Pollen tube growth requires coordination of cytoskeletal dynamics and apical secretion. The regulatory phospholipid, phosphatidylinositol 4,5-bisphosphate (PtdIns(4,5)P2) is enriched in the subapical plasma membrane of pollen tubes and can influence both actin dynamics and secretion. How alternative PtdIns(4,5)P2-effects are specified is unclear. Spinning disc microscopy (SD) reveals dual distribution of a fluorescent PtdIns(4,5)P2-reporter in dynamic plasma membrane nanodomains vs. apparent diffuse membrane labelling, consistent with spatially distinct coexisting pools of PtdIns(4,5)P2. Several PI4P 5-kinases (PIP5Ks) can generate PtdIns(4,5)P2 in pollen tubes. Despite localizing to one membrane region, AtPIP5K2 and NtPIP5K6 display distinctive overexpression effects on cell morphologies, respectively related to altered actin dynamics or membrane trafficking. When analyzed by SD, AtPIP5K2-EYFP associated with nanodomains, whereas NtPIP5K6-EYFP localized diffusely. Chimeric AtPIP5K2 and NtPIP5K6 variants with reciprocally swapped membrane-associating domains evoked reciprocally shifted effects on cell morphology upon overexpression. Overall, PI4P 5-kinase variants targeted to nanodomains stabilized actin, suggesting a specific function of PtdIns(4,5)P2-nanodomains. A distinct role of nanodomain-associated AtPIP5K2 in actin regulation is further supported by proximity to and interaction with the Rho-GTPase NtRac5, and by functional interplay with elements of ROP-signalling. Plasma membrane nano-organization may thus aid the specification of PtdIns(4,5)P2-functions to coordinate cytoskeletal dynamics and secretion in pollen tubes.

cell biology↗

bioRxiv2020

Optimal seasonal routines across animals and latitudes

Most animals face seasonal fluctuations in food availability and need to develop an annual routine that maximizes their lifetime reproductive success. Two particularly common strategies are reducing energy expenditure and building storage to sustain the animal in meager periods (winters). Here, we pose a simple and generic model for an animal that can decide, at each time during the season, on its level of foraging effort and on building energy stores. Using dynamic optimization, we identify the optimal annual routines that maximize the trade-off between energy and mortality over a life-long horizon. We investigate how the optimal strategies depend on the body size and longevity of the animal, and upon the seasonal variability in the environment. We find that with large fluctuations, the optimal annual routine for small animals is to develop a surviving egg/spore stage rather than to attempt to survive the winter. Medium sized animals invest heavily in reserves to allow long hibernation, while larger animals only need smaller reserves and a shorter hibernation period. In environments with smaller fluctuations, organisms do not need energy stores or hibernation but reduce foraging activities during spring and summer where their fitness is highest. Our optimization model can be used as a null hypothesis to explain the annual routines of animals of all body sizes across the globe.

ecology↗