Biology subjects

Evans, G.

Publications and source records attributed to Evans, G..

7 recordsLinked to original sources

bioRxiv2025

An ancient plant symbiotic fungus with distinct features identified through advanced fluorescence and Raman imaging

Mycorrhizal associations between fungi and plants are a fundamental aspect of terrestrial ecosystems. The occurrence of mycorrhizae is now well established in c. 85% of extant plants, yet the geological record of these associations is sparse. Fossil evidence from early Paleozoic terrestrial environments provides rare insights into these ancient symbioses, but imaging micro-scale fossil fungi in surrounding plant tissues is challenging. We employed brightfield microscopy, confocal laser scanning microscopy (CLSM), fluorescence lifetime imaging microscopy (FLIM), and Raman spectroscopy to investigate a newly identified endomycorrhizal fungus within a 407-million-year-old plant from the Windyfield Chert. An exquisite preservation and our advanced imaging approach enabled the description of the fungal structures to unprecedented detail. The fungus, Rugosomyces lavoisieriae gen. nov., & sp. nov., exhibits structural features resembling modern Glomeromycotina arbuscular mycorrhizal fungi. Combining CLSM with FLIM provides extra information by separating features based upon fluorescence lifetime. Raman spectroscopy indicates that fungal arbuscules and plant water-conducting cells have undergone geological alterations, resulting in a similar chemical composition. These findings enhance our understanding of ancient plant-fungal symbioses and demonstrate the potential of advanced imaging techniques in paleobotanical research. Significance statementMost living plants form mycorrhizal associations with fungi, but how this ancient evolutionary relationship developed is poorly understood. Exceptional fossil materials preserve direct evidence of endomycorrhizae, but imaging micro-scale fossil fungi inside plant cells is challenging. Here we use confocal microscopy combined with fluorescence lifetime imaging and Raman spectroscopy to study a new endomycorrhizal fungus inside a 400-million-year-old plant. We document structures resembling modern Glomeromycotina. We show that the residual organic matter of the fungus and the plant cell walls has a very short fluorescence lifetime, which provides a new means of imaging these fossils. Raman spectroscopy indicates that both arbuscules and water-conducting cells were geologically altered by their diagenetic history and that they now have a similar chemical composition.

plant biology↗

bioRxiv2024

How do gepotidacin and zoliflodacin stabilize DNA-cleavage complexes with bacterial type IIA topoisomerases? 1. Experimental definition of metal binding sites

One of the challenges for experimental structural biology in the 21st century is to see chemical reactions happen. Staphylococcus aureus (S. aureus) DNA gyrase is a type IIA topoisomerase that can create temporary double-stranded DNA breaks to regulate DNA topology. Drugs, such as gepotidacin, zoliflodacin and the quinolone moxifloxacin, can stabilize these normally transient DNA-strand breaks and kill bacteria. Crystal structures of uncleaved DNA with a gepotidacin precursor (2.1[A] GSK2999423) or with doubly cleaved DNA and zoliflodacin (or with its progenitor QPT-1) have been solved in the same P61 space-group (a=b{approx}93[A], c{approx}412[A]). This suggests it may be possible to observe the two DNA-cleavage steps (and two DNA-religation steps) in this P61 space-group. Here a 2.58[A] anomalous manganese dataset in this crystal form is solved, and four previous crystal structures (1.98[A], 2.1[A], 2.5[A] and 2.65[A]) in this crystal form are re-refined to clarify crystal contacts. The structures clearly suggest a single moving metal mechanism - presented in an accompanying (2nd) paper. A previously published 2.98[A] structure of a yeast topoisomerase II, which has static disorder around a crystallographic twofold axis, was published as containing two metals at one active site. Re-refined coordinates of this 2.98[A] yeast structure are consistent with other type IIA topoisomerase structures in only having one metal ion at each of the two different active sites.

biophysics↗

bioRxiv2023

A redox switch allows binding of ferrous and ferric ions in the cyanobacterial iron binding protein FutA from Prochlorococcus

The marine cyanobacterium Prochlorococcus is a main contributor to global photosynthesis, whilst being limited by iron availability. Cyanobacterial genomes typically encode two different types of FutA iron binding proteins: periplasmic FutA2 ABC transporter subunits bind Fe(III), while cytosolic FutA1 binds Fe(II). Owing to their small size and their economized genome Prochlorococcus ecotypes typically possess a single futA gene. How the encoded FutA protein might bind different Fe oxidation states was previously unknown. Here we use structural biology techniques at room temperature to probe the dynamic behavior of FutA. Neutron diffraction confirmed four negatively charged tyrosinates, that together with a neutral water molecule coordinate iron in trigonal bipyramidal geometry. Positioning of the positively charged Arg103 side chain in the second coordination shell yields an overall charge-neutral Fe(III) binding state in structures determined by neutron diffraction and serial femtosecond crystallography. Conventional rotation X-ray crystallography using a home source revealed X-ray induced photoreduction of the iron center with observation of the Fe(II) binding state; here, an additional positioning of the Arg203 side chain in the second coordination shell maintained an overall charge neutral Fe(II) binding site. Dose series using serial synchrotron crystallography and an XFEL X-ray pump-probe approach capture the transition between Fe(III) and Fe(II) states, revealing how Arg203 operates as a switch to accommodate the different iron oxidation states. This switching ability of the Prochlorococcus FutA protein may reflect ecological adaptation by genome streamlining and loss of specialized FutA proteins. Significance StatementOceanic primary production by marine cyanobacteria is a main contributor to carbon and nitrogen fixation. Prochlorococcus is the most abundant photosynthetic organism on Earth, with an annual carbon fixation comparable to the net global primary production from agriculture. Its remarkable ecological success is based on the ability to thrive in low nutrient waters. To manage iron limitation, Prochlorococcus possesses the FutA protein for iron uptake and homeostasis. We reveal a molecular switch in the FutA protein that allows it to accommodate binding of iron in either the Fe(III) or Fe(II) state using structural biology techniques at room temperature and provide a plausible mechanism for iron binding promiscuity.

biophysics↗

bioRxiv2022

Loss of NF1 causes tactile hypersensitivity and impaired synaptic transmission in a Drosophila model of autism spectrum disorder

Autism Spectrum Disorder (ASD) is a neurodevelopmental condition in which the mechanisms underlying its core symptomatology are largely unknown. Studying animal models of monogenic syndromes associated with ASD, such as neurofibromatosis type 1 (NF1), can offer insights into its aetiology. Here, we show that loss of function of the Drosophila NF1 ortholog results in larval tactile hypersensitivity, paralleling the sensory abnormalities observed in individuals with ASD. Mutant larvae also exhibit synaptic transmission deficits at the glutamatergic neuromuscular junction (NMJ), with increased spontaneous but reduced evoked release. Diminished expression of NF1 specifically within central cholinergic neurons induces both excessive neuronal firing and tactile hypersensitivity, suggesting the two may be linked. Furthermore, both impaired synaptic transmission and behavioural deficits are fully rescued via knockdown of Ras proteins. These findings validate NF1-/- Drosophila as a tractable model of ASD with the potential to elucidate important pathophysiological mechanisms.

neuroscience↗

bioRxiv2020

Formation of flavone-based wooly fibres by glandular trichomes of Dionysia tapetodes.

Dionysia tapetodes, a small cushion-forming mountainous evergreen in the Primulaceae, possesses a vast surface-covering of long silky fibres forming the characteristic "wooly" farina. This contrasts with some related Primula which instead possess a powdery farina. Using a combination of cell biology and analytical chemical techniques, we provide a detailed insight of wooly farina formation by glandular trichomes that produce a mixture of flavone and substituted flavone derivatives, including hydroxyflavones. Conversely, our analysis show that the powdery form consist almost entirely of flavone. The wooly farina in D. tapetodes is extruded through specific sites at the surface of the glandular head cell, characterised by a small complete gap in the plasma membrane, cell wall and cuticle. The data is consistent with formation and thread elongation occurring from within the cell. The putative mechanism of wool thread formation and its stability is discussed.

plant biology↗

bioRxiv2020

A Workflow for Protein Structure Determination from Thin Crystal Lamella by Micro-Electron Diffraction

Micro-Electron Diffraction (MicroED) has recently emerged as a powerful method for the analysis of biological structures at atomic resolution. This technique has been largely limited to protein nanocrystals which grow either as needles or plates measuring only a few hundred nanometres in thickness. Furthermore, traditional microED data processing uses established X-ray crystallography software that is not optimised for handling compound effects that are unique to electron diffraction data. Here, we present an integrated workflow for microED, from sample preparation by cryo-focused ion beam milling, through data collection with a standard Ceta-D detector, to data processing using the DIALS software suite, thus enabling routine atomic structure determination of protein crystals of any size and shape using microED. We demonstrate the effectiveness of the workflow by determining the structure of proteinase K to 2.0 [A] resolution and show the advantage of using protein crystal lamellae over nanocrystals.

biophysics↗

bioRxiv2020

Effects of breeding history and crop management on the root architecture of wheat

AimsSelection for optimal root system architecture (RSA) is important to ensure genetic gains in the sustainable production of wheat (Triticum aestivum L.). Here we examine the idea that past wheat breeding has led to changes in RSA and that future breeding efforts can focus directly on root traits to improve adaptation to a target environment. MethodsWe conducted three field trials using diverse wheat varieties, including modern and historic UK varieties and non-UK landraces, tested under contrasting tillage regimes (non-inversion tillage versus conventional ploughing) or different seeding rates (standard rate versus high rate). We used field excavation, washing and measurement of root crowns ( shovelomics) to characterise RSA traits, including: numbers of seminal, crown and nodal roots per plant, and crown root growth angle. ResultsWe found large differences among genotypes for all root traits. Modern varieties generally had fewer roots per plant than historic varieties. There were fewer crown roots and root angles were wider, on average, under shallow non-inversion tillage compared with conventional ploughing. Crown root numbers per plant also tended to be smaller at a high seeding rate compared with the standard rate. There were significant genotype-by-year, genotype-by-tillage and genotype-by-seeding-rate interactions for many root traits. ConclusionsSmaller root systems is likely to be a result of past selection and may have facilitated historical yield increases by reducing below-ground competition within the crop. The effects of crop management practices on RSA depend on genotype, suggesting that future breeding could select for improved RSA traits in resource-efficient farming systems.

plant biology↗