Biology subjects

Kelly, M.

Publications and source records attributed to Kelly, M..

At least 37 records · Page 2Linked to original sources

bioRxiv2025

Molecular mechanisms regulating PDE11A4 age-related liquid-liquid phase separation (LLPS) and its reversal by selective, potent and orally-available PDE11A4 small molecule inhibitors both in vitro and in vivo

PDE11A is a little-studied phosphodiesterase family that breaks down cAMP and cGMP, with the PDE11A4 isoform enriched in the memory-related hippocampus. Age-related increases in hippocampal PDE11A expression occur in human and rodents, causing age-related cognitive decline of social memories. Interestingly, this age-related increase triggers PDE11A4 liquid-liquid phase separation (LLPS), causing the enzyme to accumulate in the brain in filamentous structures termed "ghost axons". Here we sought to identify molecular mechanisms regulating PDE11A4 LLPS and therapeutic approaches capable of reversing it. PDE11A4 LLPS was reduced by phosphorylation of PDE11A4-S163 or-S239 and the D355A mutation that blocks the effect of cGMP binding the PDE11A4 GAF-A domain. PDE11A4 LLPS was increased by inhibiting kinases with staurosporine or stimulating packaging/repacking via the trans-Golgi network by overexpressing TGN38 or RhoB. 8 PDE11 inhibitors (MLG-122, MLG-185, MLG-199, SMQ-02-57, SMQ-03-30, SMQ-03-20, tadalafil, and BC11-38) across 3 scaffolds reverse overexpression-related PDE11A4 LLPS in HT22 mouse hippocampal neuronal cells. This effect of PDE11A4 inhibitors occurs within minutes, is reversed upon washout of lower but not higher concentrations, and occurs in part by reducing PDE11A4 homodimerization. PDE11A4 inhibitors also rescued exacerbated PDE11A4 LLPS triggered by aging-like S117D/S124D phosphomimic mutations, staurosporine, or TGN38/RhoB overexpression. In vivo, orally-administered 30mg/kg SMQ-03-20 reversed age-related increases in PDE11A4 ghost axons and neuroinflammation in old mice. Thus, PDE11A inhibitors that reverse age-related PDE11A4 LLPS in HT22 hippocampal cells also reduce PDE11A4 ghost axons and neuroinflammation in the aged mouse brain, indicating therapeutical potential.

neuroscience↗

bioRxiv2025

Diatom biodiversity response to shrinking glaciers in the Peruvian Andes

Alpine river biodiversity is under threat as climate change is driving extensive glacier retreat worldwide, altering aquatic habitats and restructuring biotic communities. However, with the exception of macroinvertebrates, the responses of aquatic groups to deglaciation have received little attention, particularly in tropical ecosystems where species may respond differently owing to additional pressures such as distinct wet and dry seasons, limited snowmelt and extreme high altitude and thus lower oxygen availability. Utilising a continuum of glacier cover across 23 rivers in the Peruvian Andes (Cordilleras Blanca and Vilcanota), we demonstrate that diatoms, abundant primary producers in glacier-fed rivers, show regionally consistent increased -diversity and density responses with deglaciation. However, {beta}-diversity was found to decline, potentially reflecting the reduction of habitat heterogeneity and niche space both within and between alpine rivers as glaciers are lost. High regional biodiversity, with just 23% of taxa found in both regions, was closely linked to the distinct underlying geologies of the Andes which influence water chemistry. Thirty-eight taxa were found exclusively at sites with glacier cover [≥] 25%, which is considerably higher than numbers recorded in European alpine rivers. However, 31 taxa were identified only at non-glacial sites suggesting deglaciation could open up more habitat for these species. Peruvian rivers overall showed similar responses to glacier retreat as temperate alpine systems, but the large number of taxa found only at mid-high glacial influence sites suggests tropical diatom assemblages may be especially vulnerable to climate change.

ecology↗

bioRxiv2025

Transient Acute Neuronal Activation Response Caused by High Concentrations of Oligonucleotides in the Cerebral Spinal Fluid

Oligonucleotide (ON) therapeutics are promising as a disease-modifying therapy for central nervous system disorders. Intrathecal ON administration into the cerebral spinal fluid is a safe and effective delivery mode to the CNS. However, preclinical studies have shown acute toxicities following high-dose central ON delivery. Here we characterize a transient neurobehavioral change peaking 15 minutes after ON dosing and resolving after 120 minutes. Symptoms include shaking, muscle twitching, cramping, hyperactivity, stereotypic movements, hyperreactivity, vocalizations, tremors, convulsions, and seizures. These are collectively referred here as the acute neuronal activation response. Acute neuronal activation is observed in rats, mice, and non-human primates and is quantifiable using a simple scoring system. It is distinct from acute sedation seen with some phosphorothioate-modified antisense oligonucleotides, characterized by loss of spinal reflexes, ataxia, and sedation. The acute neuronal activation response is largely sequence-independent and is driven by ON chelation of divalent cations, particularly influenced by the divalent cations-to-ON ratio in the dosing solution. Acute neuronal activation can be safely mitigated by adjusting this ratio through magnesium supplementation in the ON formulation. We provide a comprehensive framework for quantifying and mitigating the acute neuronal activation response caused by high concentrations of centrally delivered ON therapeutics in preclinical species. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=79 SRC="FIGDIR/small/638138v1_ufig1.gif" ALT="Figure 1"> View larger version (32K): org.highwire.dtl.DTLVardef@b18796org.highwire.dtl.DTLVardef@13c9898org.highwire.dtl.DTLVardef@1463019org.highwire.dtl.DTLVardef@ffe719_HPS_FORMAT_FIGEXP M_FIG C_FIG

neuroscience↗

bioRxiv2025

Permissive central tolerance plus defective peripheral checkpoints licence pathogenic memory B cells in CASPR2-antibody encephalitis

Autoimmunity affects 10% of the population. Within this umbrella, autoantibody-mediated diseases targeting one autoantigen provide a unique opportunity to comprehensively understand the developmental pathway of disease-causing B cells and autoantibodies. While such autoreactivities are believed to be generated during germinal centre reactions, the roles of earlier immune checkpoints in autoantigen-specific B cell tolerance are poorly understood. We address this concept in patients with CASPR2-autoantibody encephalitis and healthy controls. In both groups, comparable and high ([~]0.5%) frequencies of unmutated CASPR2-reactive naive B cells were identified. By contrast, CASPR2-reactive memory B cells were exclusive to patients, and their B cell receptors demonstrated affinity-enhancing somatic mutations with heterogenous binding kinetics. These effector molecules possessed epitope-dependent pathogenic effects in vitro neuronal cultures and in vivo. The unmutated common ancestors of these memory B cells showed a distinctive balance between strong CASPR2 reactivity and very limited binding across the remaining human proteome. Our results are the first to propose mechanisms underlying autoantigen-specific tolerance in humans. We identify permissive central tolerance, defective peripheral tolerance and heterogenous autoantibody binding properties as sequential pathogenic steps which licence CASPR2-directed pathology. By leveraging the basic immunobiology, we rationally direct tolerance-restoring approaches in CASPR2-antibody diseases. This paradigm is applicable across autoimmune conditions.

immunology↗

bioRxiv2024

Age-related increases in PDE11A4 protein expression trigger liquid:liquid phase separation (LLPS) of the enzyme that can be reversed by PDE11A4 small molecules inhibitors

PDE11A is a little-studied phosphodiesterase sub-family that breaks down cAMP/cGMP, with the PDE11A4 isoform being enriched in the memory-related brain region called the hippocampus. Age-related increases in PDE11A expression occur in human and rodent hippocampus and cause age-related cognitive decline of social memories. Interestingly, the age-related increase in PDE11A4 protein ectopically accumulates in spherical clusters that group together in the brain to form linear filamentous patterns termed "ghost axons." The biophysical/physiochemical mechanisms underlying this age-related clustering of PDE11A4 are not yet known. As such, we determine here if age-related clustering of PDE11A4 may reflect liquid:liquid phase separation (LLPS), and if PDE11A inhibitors being developed for age-related cognitive decline can reverse this biomolecular condensation. We found that human and mouse PDE11A4 exhibit several LLPS-promoting sequence features including intrinsically disordered regions, non-covalent pi-pi interactions, and prion-like domains, with multiple bioinformatic tools predicting PDE11A4 undergoes LLPS. Consistent with these predictions, age-related PDE11A4 clusters were non-membrane bound spherical droplets that progressively fuse over time in a concentration-dependent manner. 5 different PDE11 inhibitors (tadalafil, BC11-38, SMQ-02-57, SMQ-03-30 and SMQ-03-20) across 3 scaffolds reversed PDE11A4 LLPS (a.k.a. remixing) in hippocampal HT22 cells, with PDE11A4 droplets reforming (a.k.a. de-mixing) following a 5-hour washout of low but not high concentrations of these compounds. Strikingly, a single oral administration of 30 mg/kg SMQ-03-20 substantially reduced the presence of PDE11A4 ghost axon in the aged mouse brain. Thus, PDE11A4 exhibits 4 defining criteria of LLPS, and PDE11A small molecule inhibitors reverse this age-related phenotype both in vitro and in vivo.

neuroscience↗

bioRxiv2024

Limited O-specific polysaccharide (OSP)-specific functional antibody responses in young children with Shigella infection in Bangladesh

Shigellosis is the second leading cause of diarrheal death in children younger than five years of age globally. At present, there is no broadly licensed vaccine against shigella infection. Previous vaccine candidates have failed at providing protection for young children in endemic settings. Improved understanding of correlates of protection against Shigella infection and severe shigellosis in young children living in endemic settings is needed. Here, we applied a functional antibody profiling approach to define Shigella-specific antibody responses in young children versus older individuals with culture-confirmed shigellosis in Bangladesh, a Shigella endemic area. We analyzed Shigella-specific antibody isotypes, FcR binding and antibody-mediated innate immune cell activation in longitudinal serum samples collected at clinical presentation and up to 1 year later. We found that higher initial Shigella O-specific polysaccharide (OSP)-specific and protein-specific IgG and Fc{gamma}R binding levels correlated with less severe disease regardless of patient age, but that individuals under 5 years of age developed a less prominent class switched, FcR-binding, functional and durable antibody response against both OSP and protein Shigella antigens than older individuals. Focusing on the largest cohort, we found that functional S. flexneri 2a OSP-specific responses were significantly induced only in individuals over age 5 years, and that these responses promoted monocyte phagocytosis and activation. Our findings suggest that in a Shigella endemic region, young children with shigellosis harbor a functional antibody response that fails to maximally activate monocytes; such a response may be important in facilitating subsequent innate cell clearance of Shigella, especially via recruitment and activation of polymorphonuclear cells capable of directly killing Shigella.

immunology↗

bioRxiv2024

A Novel Whole Tissue Explant Model of Hidradenitis Suppurativa

Hidradenitis Suppurativa (HS) is a relatively common and highly morbid inflammatory skin disease. Due to our relatively limited understanding of HSs pathogenesis, there are currently insufficient treatment options available, and many patients medical needs are not being met. This is partly due to a scarcity of ex vivo human assays and animal models that accurately recapitulate the disease. To address this deficit, we have developed a whole-tissue explant model of HS to examine its pathogenic mechanisms and the efficacy of potential treatments within intact human tissue. We measured cytokine protein and RNA within whole tissue maintained in an agar-media solution, finding that IL-6 and IL-8 concentrations trended upwards in both HS explants and healthy controls, while IL-17A, IL-1{beta}, and TNF- exhibited increases in HS tissue alone. We also show that the explants were responsive to treatment with both dexamethasone and IL-2. Not only do our results show that this model effectively delivers treatments throughout the explants, but they also elucidate which cytokines are related to the explant process regardless of tissue state and which are related to HS tissue specifically, laying the groundwork for future implementations of this model.

immunology↗

bioRxiv2024

Deep learning guided design of dynamic proteins

Deep learning has greatly advanced design of highly stable static protein structures, but the controlled conformational dynamics that are hallmarks of natural switch-like signaling proteins have remained inaccessible to de novo design. Here, we describe a general deep-learning-guided approach for de novo design of dynamic changes between intra-domain geometries of proteins, similar to switch mechanisms prevalent in nature, with atom-level precision. We solve 4 structures validating the designed conformations, show microsecond transitions between them, and demonstrate that the conformational landscape can be modulated by orthosteric ligands and allosteric mutations. Physics-based simulations are in remarkable agreement with deep-learning predictions and experimental data, reveal distinct state-dependent residue interaction networks, and predict mutations that tune the designed conformational landscape. Our approach demonstrates that new modes of motion can now be realized through de novo design and provides a framework for constructing biology-inspired, tunable and controllable protein signaling behavior de novo.

bioengineering↗

bioRxiv2024

Ancient Borrelia genomes document the evolutionary history of louse-borne relapsing fever

Several disease-causing bacteria have transitioned from tick-borne to louse-borne transmission, a process associated with increased virulence and genome reduction. However, the historical time frame and speed of such evolutionary transitions have not been documented with ancient genomes. Here, we discover four ancient cases of Borrelia recurrentis, the causative agent of louse-borne relapsing fever, in Britain between [~]600 and 2,300 years ago, and sequence whole genomes up to 29-fold coverage. We estimate a recent divergence from the closest tick-borne ancestor, likely within the last [~]8,000 years. We reconstruct a chronology of gene losses and acquisitions using the pan-genome of related species, and show that almost all of the reductive evolution observed in B. recurrentis had occurred by [~]2,000 years ago, and was thus a rapid process after divergence. Our observations provide a new understanding of the origins of B. recurrentis and document complex reductive evolution in a specialist vector-borne pathogen.

evolutionary biology↗

bioRxiv2024

Dissolved organic matter fluxes and fatty acid composition from shallow-water and deep-water demo- and hexactinellid sponges from New Zealand

Sponges are an important component of shallow- and deep-water ecosystems enhancing eukaryotic biodiversity via diverse endo- and epibiota and by providing three dimensional habitats for benthic invertebrates and fishes. Sponge biodiversity is particularly high in the waters around New Zealand (Southwest Pacific), where we collected two shallow- and two deep-water sponge species (Tedania sp., Suberea meandrina, Farrea raoulensis, Artemisina sp.) for ex-situ incubation experiments to detect whether these sponges may process dissolved organic matter. We assessed the biochemical and phospholipid-derived fatty acids (PLFAs) and measured dissolved organic carbon (DOC) and total dissolved nitrogen (TDN) fluxes. Changes in fluorescent dissolved organic matter (FDOM) over time were analyzed and the results were linked to the bacterial communities of the sponge holobiont. Dried sponge tissue consisted of 17.5 {+/-} 3.75% organic (org.) C and of 4.34 {+/-} 1.02% total nitrogen (TN) with a natural stable isotope composition of -19.0 {+/-}0.25{per thousand} for {delta}13org. C and of 10.2 {+/-}2.43{per thousand} for {delta}15TN. None of the DOC fluxes was significant and only the release of TDN by Tedania sp. was significantly different from 0 mol TDN g org. C-1 d-1. We detected the presence of four fluorophores in the FDOM pool: 2 tryptophan- and protein-like fluorophores (C1, C2), 1 humic-like fluorophore (C3), and 1 tyrosine-like fluorophore (C4). The maximum fluorescence intensity Fmax of C1 decreased significantly in S. meandrina incubations, whereas Fmax of C2 grew in the same incubations. Fmax of C3 increased in Tedania sp. incubations, in which Fmax of C4 decreased. In comparison, Fmax of C4 in S. meandrina rose. The PLFA composition of sponge tissue was dominated by long-chain fatty acids, saturated fatty acids, and monosaturated fatty acids and most PLFAs were sponge- and bacteria-specific. The bacterial community of the demosponges Artemisina sp., S. meandrina and Tedania sp. consisted mostly of Proteobacteria and Chloroflexota, whereas the dominating bacteria phylum of the hexactinellid F. raoulensis was Proteobacteria. We proposed that the holobionts of S. meandrina and Tedania sp. contain bacteria that are involved in the transformation and degradation of DOM. In S. meandrina, Chloroflexota and Poribacteria may degrade tryptophan-like fluorophores to a chemically modified tryptophan-like and protein-like fluorophore, while producing a tyrosine-like fluorophore. In Tedania sp., Chloroflexota may contribute to the release of significant amounts of TDN by producing humic-like fluorophores, while degrading tyrosine-like fluorophores. Farrea raoulensis may not take up DOM due to a lack of Poribacteria and Chloroflexota or may use colloidal instead of truly dissolved DOC.

ecology↗

bioRxiv2024

North Pontic crossroads: Mobility in Ukraine from the Bronze Age to the early modern period

The North Pontic region, which encompasses present-day Ukraine, was a crossroads of migration as it connected the vast Eurasian Steppe with Central Europe. We generated shotgun-sequenced genomic data for 91 individuals dating from around 7,000 BCE to 1,800 CE to study migration and mobility history in the region, with a particular focus on historically attested migrating groups during the Iron Age and the medieval period, such as Scythian, Chernyakhiv, Saltiv and Nogai associated peoples. We infer a high degree of temporal heterogeneity in ancestry, with fluctuating genetic affinities to present-day Western European, Eastern European, Western Steppe and East Asian groups. We also infer high heterogeneity in ancestry within geographically, culturally and socially defined groups. Despite this, we find that ancestry components which are widespread in Eastern and Central Europe have been present in the Ukraine region since the Bronze Age.

genomics↗

bioRxiv2024

Nutrient fluxes, oxygen consumption and fatty acid composition from deep-water demo- and hexactinellid sponges from New Zealand

Sponges are an important component of deep-water ecosystems enhancing eukaryotic biodiversity by hosting diverse endo- and epibiota and providing three dimensional habitats for benthic invertebrates and fishes. As holobionts they are important hosts of microorganisms which are involved in carbon and nitrogen cycling. While increasing exploration of deep-water habitats results in new sponge species being discovered, little is known about their physiology and role in nutrient fluxes. Around New Zealand (Southwest Pacific), the sponge biodiversity is particularly high, and we selected six deep-sea sponge clusters (Saccocalyx, Suberites, Tedania, Halichondria/ Dendoricella, Sceptrulophora, Lissodendoryx) for in-situ and ex-situ experiments. We investigated the biochemical composition of the sponges, measured oxygen consumption and inorganic nutrient fluxes, as well as bacterial and phospholipid-derived fatty acid (PLFA) compositions. Our aim was to assess differences in fluxes and fatty acid composition among sponge clusters and linking their bacterial communities to nitrogen cycling processes. All sponges excreted nitrite and ammonia. Nitrate and phosphate excretion were independent of phylum affiliation (Demospongiae, Hexactinellida). Nitrate was excreted by the Halichondria/ Dendoricella and Lissodendoryx clusters, whereas the Suberites, Tedania, and Sceptrulophora clusters consumed it. Phosphate was excreted by Sceptrulophora and Halichondria/ Dendoricella clusters and consumed by all other clusters. Silicon was consumed by all sponge clusters, except for Saccocalyx and Halichondria/ Dendoricella clusters. Oxygen consumption rates ranged from 0.17 to 3.56{+/-}0.60 mmol O2 g C d-1. The PLFA composition was very sponge-cluster dependent and consisted mostly of long-chain fatty acids. Most PLFAs were sponge-specific, followed by bacteria-specific PLFAs, and others. All sponge clusters, except for Suberites, were low-microbial abundance (LMA) sponges whose bacterial community composition was dominated by Proteobacteria, Bacteroidota, Planctomycetota, and Nitrospinota. The Suberites cluster consisted of high-microbial abundance (HMA) sponges with Proteobacteria, Chloroflexota, Acidobacteriota, and Actinobacteriota as dominant bacteria. Based on the inorganic nitrogen flux measurements, we identified three types of nitrogen cycling in the sponges: In type 1, sponges (Dendoricella spp. indet., Lissodendoryx cluster) respired aerobically and ammonificated organic matter (OM) to ammonium, fixed N2 to ammonium, and nitrified aerobically heterotrophically produced ammonium to nitrate and nitrite. In type 2, sponges (Halichondria sp., Sceptrulophora, Suberites, Tedania clusters) respired OM aerobically and ammonificated it to ammonium. They also reduced nitrate anaerobically to ammonium via dissimilatory nitrate reduction to ammonium. In type 3, ammonium was microbially nitrified to nitrite and afterwards to nitrate presumably by ammonium-oxidizing Bacteria and/ or Archaea.

ecology↗

bioRxiv2024

Evaluating the epizootic and zoonotic threat of an H7N9 low pathogenicity avian influenza virus (LPAIV) variant associated with enhanced pathogenicity in turkeys

Between 2013-2017, A/Anhui/1/13-lineage (H7N9) low pathogenicity avian influenza virus (LPAIV) was epizootic in chickens in China causing mild disease, with 616 fatal human cases. Despite poultry vaccination, H7N9 has not been eradicated. Previously we demonstrated increased pathogenesis in turkeys infected with H7N9, correlating with the emergence of the L217Q (L226Q H3 numbering) polymorphism in the haemagglutinin (HA) protein. A Q217 containing virus also arose and is now dominant in China following vaccination. We compared infection and transmission of this Q217 containing turkey-adapted (ty-ad) isolate alongside the H7N9 (L217) wild-type (wt) virus in different poultry species, and investigated the zoonotic potential in the ferret model. Both wt and ty-ad viruses demonstrated similar shedding and transmission in turkeys and chickens. However, the ty-ad virus was significantly more pathogenic than the wt virus in turkeys but not in chickens, causing 100% and 33% mortality in turkeys respectively. Expanded tissue tropism was seen for the ty-ad virus in turkeys but not chickens, yet the viral cell receptor distribution was broadly similar in visceral organs of both species. The ty-ad virus required exogenous trypsin for in vitro replication yet had increased replication in primary avian cells. Replication was comparable in mammalian cells and the ty-ad virus replicated successfully in ferrets. The L217Q polymorphism also affected antigenicity. Therefore, H7N9 infection in turkeys can generate novel variants with increased risk through altered pathogenicity and potential HA antigenic escape. These findings emphasise the requirement for enhanced surveillance and understanding of A/Anhui/1/13-lineage viruses and their risk to different species.

microbiology↗

bioRxiv2024

Machine Learning Approaches in Label-Free Small Extracellular Vesicles Analysis with Surface-Enhanced Raman Scattering (SERS) for Cancer Diagnostics

Early diagnosis remains of pivotal importance in reducing patient morbidity and mortality in cancer. To this end, liquid biopsy is emerging as a tool to perform broad cancer screenings. Small extracellular vesicles (sEVs), also called exosomes, found in bodily fluids can serve as important cancer biomarkers in these screenings. Our group has recently developed a label-free electrokinetic microchip to purify sEVs from blood. Herein, we demonstrate the feasibility to integrate this approach with surface-enhanced Raman scattering (SERS) analysis. SERS can be used to characterized extracted sEVs through their vibrational fingerprint that changes depending on the origin of sEVs. While these changes are not easily identified in spectra, they can be modeled with machine learning (ML) approaches. Common ML approaches in the field of spectral analysis use dimensionality reduction method that often function as a black box. To avoid this pitfall, we used Shapley additive explanations (SHAP) is a type of explainable AI (XAI) that bridges ML models and human comprehension by calculating the specific contribution of individual features to a models predictions, directly correlating model/decisions with the original data. Using these approaches we demonstrated a proof-of-concept model predictive of cancer from isolated sEVs, integrating the electrokinetic device and SERS. This work explores the use of explainable AI to perform diagnostic analysis on complex SERS data of clinical samples, while reporting interpretable biochemical information. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=74 SRC="FIGDIR/small/581099v2_ufig1.gif" ALT="Figure 1"> View larger version (18K): org.highwire.dtl.DTLVardef@1cbcb8aorg.highwire.dtl.DTLVardef@9ffc00org.highwire.dtl.DTLVardef@15999a9org.highwire.dtl.DTLVardef@1775726_HPS_FORMAT_FIGEXP M_FIG C_FIG

bioengineering↗

bioRxiv2023

The sleep quality- and myopia-linked PDE11A-Y727C variant impacts neural physiology by reducing catalytic activity and altering subcellular compartmentalization of the enzyme

Recently, a Y727C variant in the dual-specific 3,5-cyclic nucleotide phosphodiesterase 11A (PDE11A-Y727C) was linked to increased sleep quality and reduced myopia risk in humans. Given the well-established role that the PDE11 substrates cAMP and cGMP play in eye physiology and sleep, we determined if 1) PDE11A protein is expressed in the retina or other eye segments in mouse, 2) PDE11A-Y7272C affects catalytic activity and/or subcellular compartmentalization more so than the nearby suicide-associated PDE11A-M878V variant, and 3) Pde11a deletion alters eye growth or sleep quality in male and female mice. Western blots show distinct protein expression of PDE11A4, but not PDE11A1-3, in eyes of Pde11a WT--but not KO mice--that vary by eye segment and age. In HT22 and COS-1 cells, PDE11A4-Y727C reduces PDE11A4 catalytic activity far more than PDE11A4-M878V, with both variants reducing PDE11A4-cAMP more so than PDE11A4-cGMP activity. Despite this, Pde11a deletion does not alter age-related changes in retinal or lens thickness, axial length, nor vitreous or anterior chamber depth. Further, Pde11a deletion only minimally changes refractive error and sleep quality. That said, both variants also dramatically alter the subcellular compartmentalization of human and mouse PDE11A4, an effect occurring independently of dephosphorylating PDE11A4-S117/S124 or phosphorylating PDE11A4-S162. Rather, re-compartmentalization of PDE11A4-Y727C is due to the loss of the tyrosine changing how PDE11A4 is packaged/repackaged via the trans-Golgi network. Therefore, the protective impact of the Y727C variant may reflect a gain-of-function (e.g., PDE11A4 displacing another PDE) that warrants further investigation in the context of reversing/preventing sleep disturbances or myopia.

neuroscience↗

bioRxiv2023

Glioblastoma cells use an integrin- and CD44-mediated motor-clutch mode of migration in brain tissue

Glioblastoma (GBM) is an aggressive malignant brain tumor with 2-year survival rates of 6.7% [1], [2]. One key characteristic of the disease is the ability of glioblastoma cells to migrate rapidly and spread throughout healthy brain tissue[3], [4]. To develop treatments that effectively target cell migration, it is important to understand the fundamental mechanism driving cell migration in brain tissue. Here we utilized confocal imaging to measure traction dynamics and migration speeds of glioblastoma cells in mouse organotypic brain slices to identify the mode of cell migration. Through imaging cell-vasculature interactions and utilizing drugs, antibodies, and genetic modifications to target motors and clutches, we find that glioblastoma cell migration is most consistent with a motor-clutch mechanism to migrate through brain tissue ex vivo, and that both integrins and CD44, as well as myosin motors, play an important role in constituting the adhesive clutch.

cancer biology↗

bioRxiv2023

Dual targeting of hepatocyte DGAT2 and stellate cell FASN alleviates nonalcoholic steatohepatitis in mice.

Nonalcoholic steatohepatitis (NASH) is a malady of multiple cell types associated with hepatocyte triglyceride (TG) accumulation, macrophage inflammation, and stellate cell-induced fibrosis, with no approved therapeutics yet available. Here, we report that stellate cell fatty acid synthase (FASN) in de novo lipogenesis drives the autophagic flux that is required for stellate cell activation and fibrotic collagen production. Further, we employ a dual targeting approach to NASH that selectively depletes collagen through selective stellate cell knockout of FASN (using AAV9-LRAT Cre in FASNfl/fl mice), while lowering hepatocyte triglyceride by depleting DGAT2 with a GalNac-conjugated, fully chemically modified siRNA. DGAT2 silencing in hepatocytes alone or in combination with stellate cell FASNKO reduced liver TG accumulation in a choline-deficient NASH mouse model, while FASNKO in hepatocytes alone (using AAV8-TBG Cre in FASNfl/fl mice) did not. Neither hepatocyte DGAT2 silencing alone nor FASNKO in stellate cells alone decreased fibrosis (total collagen), while loss of both DGAT2 plus FASN caused a highly significant attenuation of NASH. These data establish proof of concept that dual targeting of DGAT2 plus FASN alleviates NASH progression in mice far greater than targeting either gene product alone.

molecular biology↗

bioRxiv2023

Metabolic capabilities are highly conserved among human nasal-associated Corynebacterium species in pangenomic analyses

Corynebacterium species are globally ubiquitous in human nasal microbiota across the lifespan. Moreover, nasal microbiota profiles typified by higher relative abundances of Corynebacterium are often positively associated with health. Among the most common human nasal Corynebacterium species are C. propinquum, C. pseudodiphtheriticum, C. accolens, and C. tuberculostearicum. To gain insight into the functions of these four species, we identified genomic, phylogenomic, and pangenomic properties and estimated the metabolic capabilities of 87 distinct human nasal Corynebacterium strain genomes: 31 from Botswana and 56 from the USA. C. pseudodiphtheriticum had geographically distinct clades consistent with localized strain circulation, whereas some strains from the other species had wide geographic distribution spanning Africa and North America. All species had similar genomic and pangenomic structures. Gene clusters assigned to all COG metabolic categories were overrepresented in the persistent versus accessory genome of each species indicating limited strain-level variability in metabolic capacity. Based on prevalence data, at least two Corynebacterium species likely coexist in the nasal microbiota of 82% of adults. So, it was surprising that core metabolic capabilities were highly conserved among the four species indicating limited species-level metabolic variation. Strikingly, strains in the USA clade of C. pseudodiphtheriticum lacked genes for assimilatory sulfate reduction present in most of the strains in the Botswana clade and in the other studied species, indicating a recent, geographically related loss of assimilatory sulfate reduction. Overall, the minimal species and strain variability in metabolic capacity implies coexisting strains might have limited ability to occupy distinct metabolic niches. IMPORTANCEPangenomic analysis with estimation of functional capabilities facilitates our understanding of the full biologic diversity of bacterial species. We performed systematic genomic, phylogenomic, and pangenomic analyses with qualitative estimation of the metabolic capabilities of four common human nasal Corynebacterium species, along with focused experimental validations, generating a foundational resource. The prevalence of each species in human nasal microbiota is consistent with the common coexistence of at least two species. We identified a notably high level of metabolic conservation within and among species indicating limited options for species to occupy distinct metabolic niches, highlighting the importance of investigating interactions among nasal Corynebacterium species. Comparing strains from two continents, C. pseudodiphtheriticum had restricted geographic strain distribution characterized by an evolutionarily recent loss of assimilatory sulfate reduction in USA strains. Our findings contribute to understanding the functions of Corynebacterium within human nasal microbiota and to evaluating their potential for future use as biotherapeutics.

microbiology↗