Biology subjects

Ross, W.

Publications and source records attributed to Ross, W..

5 recordsLinked to original sources

bioRxiv2026

Preferential innervation of endometriosis by hyperexcitable Ret/GFRα1+ nociceptors associates with target GDNF and clinical pain

Endometriosis is a prevalent condition characterized by chronic pelvic pain that is frequently refractory to treatment. While the mechanisms underlying this pain remain poorly defined, clinical evidence often indicates that lesion innervation, but not disease stage (e.g. number and depth of lesions), correlate with pelvic pain severity. However, characterization of lesion-innervating neurons is incomplete, revealing an opportunity to identify novel, disease-modifying therapeutics. Here, we coupled functional analyses of lesion-innervating neurons in a mouse model with concurrent identification and characterization of lesion-innervating neurons from pain-phenotyped endometriosis patients. Following the confirmation of abdominal-directed pain-like behaviors in the mouse model, electrophysiological analysis revealed that lesion-innervating dorsal root ganglion (DRG) neurons are hyperexcitable compared to matched controls. These neurons are predominantly small-diameter and bind Isolectin B4, an established marker of the GDNF Family Ligand receptor, Ret. GDNF is concentrated within the stromal layer of both mouse and human lesions, adjacent to axons expressing the GDNF co-receptor, GFR1. Critically, clinical pain correlates with lesion GDNF level, axonal density, and neuronal GFR1 levels. These data provide evidence that endometrial lesions may recruit the Ret-positive subpopulation of nociceptors where they become sensitized and increase patient pain.

neuroscience↗

bioRxiv2023

Vocal complexity in the long calls of Bornean orangutans

Vocal complexity is central to many evolutionary hypotheses about animal communication. Yet, quantifying and comparing complexity remains a challenge, particularly when vocal types are highly graded. Male Bornean orangutans (Pongo pygmaeus wurmbii) produce complex and variable "long call" vocalizations comprising multiple sound types that vary within and among individuals. Previous studies described six distinct call (or pulse) types within these complex vocalizations, but none quantified their discreteness or the ability of human observers to reliably classify them. We studied the long calls of 13 individuals to: 1) evaluate and quantify the reliability of audio-visual classification by three well-trained observers, 2) distinguish among call types using supervised classification and unsupervised clustering, and 3) compare the performance of different feature sets. Using 46 acoustic features, we applied machine learning (i.e., support vector machines, affinity propagation, and fuzzy c-means) to identify call types and assess their discreteness. We additionally used Uniform Manifold Approximation and Projection (UMAP) to visualize the separation of pulses using both extracted features and spectrogram representations. Supervised approaches showed low inter-observer reliability and poor classification accuracy, indicating that pulse types were not discrete. We propose an updated pulse classification approach that is highly reproducible across observers and exhibits strong classification accuracy using support vector machines. Although the low number of call types suggests long calls are fairly simple, the continuous gradation of sounds seems to greatly boost the complexity of this system. This work responds to calls for more quantitative research to define call types and quantify gradedness in animal vocal systems and highlights the need for a more comprehensive framework for studying vocal complexity vis-a-vis graded repertoires.

animal behavior and cognition↗

bioRxiv2020

Proton wires mediate the optical signal for ArcLight-type Genetically Encoded Voltage Indicators

The genetically encoded voltage indicators, ArcLight and its derivatives, mediate voltage dependent optical signals by intermolecular, electrostatic interactions between neighboring fluorescent proteins (FPs) via proton wires. A random mutagenesis event placed a negative charge on the exterior of the FP resulting in a greater than 10-fold improvement of the voltage-dependent optical signal. Repositioning this negative charge on the exterior of the FP reversed the polarity of voltage-dependent optical signals suggesting the presence of hot spots capable of interacting with the negative charge on a neighboring FP thereby changing the fluorescent output. To explore the potential effect on the chromophore state, voltage-clamp fluorometry was performed with alternating excitation at 390 nm followed by excitation at 470 nm resulting in several mutants exhibiting voltage-dependent, ratiometric optical signals of opposing polarities. However, the kinetics, voltage ranges, and optimal FP fusion sites were different depending on the wavelength of excitation. These results suggest that the FP has external, electrostatic pathways capable of quenching fluorescence that are wavelength specific. ArcLight-derived GEVIs may therefore offer a novel way to map how conditions external to the {beta}-can structure can affect the fluorescence of the chromophore and transiently manipulate those pathways via conformational changes mediated by whole cell voltage clamp. Statement of SignificanceArcLight-type GEVIs utilize proton pathways that send charge information outside of the FP to the internal chromophore enabling voltage induced conformational changes to affect fluorescence. These pathways are excitation wavelength specific suggesting that different external positions affect the protonated and deprotonated states of the chromophore.

biophysics↗

bioRxiv2019

E. coli TraR allosterically regulates transcription initiation by altering RNA polymerase conformation and dynamics

TraR and its homolog DksA are bacterial proteins that regulate transcription initiation by binding directly to RNA polymerase (RNAP) rather than to promoter DNA. Effects of TraR mimic the combined effects of DksA and its cofactor ppGpp. How TraR and its homologs regulate transcription is unclear. Here, we use cryo-electron microscopy to determine structures of Escherichia coli RNAP, with or without TraR, and of an RNAP-promoter complex. TraR binding induced RNAP conformational changes not seen in previous crystallographic analyses, and a quantitative analysis of RNAP conformational heterogeneity revealed TraR-induced changes in RNAP dynamics. These changes involve mobile regions of RNAP affecting promoter DNA interactions, including the {beta}lobe, the clamp, the bridge helix, and several lineage-specific insertions. Using mutational approaches, we show that these structural changes, as well as effects on {sigma}70 region 1.1, are critical for transcription activation or inhibition, depending on the kinetic features of regulated promoters.

biophysics↗

bioRxiv2019

Structural basis for transcription activation by Crl through tethering of σS and RNA polymerase

In bacteria, a primary {sigma} factor associates with the core RNA polymerase (RNAP) to control most transcription initiation, while alternative {sigma} factors are used to coordinate expression of additional regulons in response to environmental conditions. Many alternative {sigma} factors are negatively regulated by anti-{sigma} factors. In Escherichia coli, Salmonella enterica, and many other{gamma} -proteobacteria, the transcription factor Crl positively regulates the alternative {sigma}S regulon by promoting the association of {sigma}S with RNAP without interacting with promoter DNA. The molecular mechanism for Crl activity is unknown. Here, we determined a single-particle cryo-electron microscopy structure of Crl-{sigma}S-RNAP in an open promoter complex with a {sigma}S regulon promoter. In addition to previously predicted interactions between Crl and domain 2 of {sigma}S ({sigma}S), the structure, along with p-benzoylphenylalanine crosslinking, reveals that Crl interacts with a structural element of the RNAP {beta} subunit we call the {beta}-clamp-toe ({beta}CT). Deletion of the {beta}CT decreases activation by Crl without affecting basal transcription, highlighting the functional importance of the Crl-{beta}CT interaction. We conclude that Crl activates {sigma}S-dependent transcription in part through stabilizing {sigma}S-RNAP by tethering {sigma}S and the {beta}CT. We propose that Crl, and other transcription activators that may use similar mechanisms, be designated {sigma}-activators.\n\nSignificance StatementIn bacteria, multiple {sigma} factors can bind to a common core RNA polymerase (RNAP) to alter global transcriptional programs in response to environmental stresses. Many {gamma}-proteobacteria, including the pathogens Yersinia pestis, Vibrio cholera, Escherichia coli, and Salmonella typhimurium, encode Crl, a transcription factor that activates {sigma}S-dependent genes. Many of these genes are involved in processes important for infection, such as biofilm formation. We determined a high-resolution cryo-electron microscopy structure of a Crl-{sigma}S-RNAP transcription initiation complex. The structure, combined with biochemical experiments, shows that Crl stabilizes {sigma}S-RNAP by tethering {sigma}S directly to the RNAP.

biophysics↗