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Maher, S.

Publications and source records attributed to Maher, S..

8 recordsLinked to original sources
bioRxiv2025

Identification and characterization of host-modulating effectors encoded by the Cluster F1 mycobacteriophage NormanBulbieJr

NormanBulbieJr (NBJ) is a temperate siphovirus isolated on the host Mycobacterium smegmatis mc2155 that encodes 102 gene products, 60 of which have no known function (NKF). Based on gene content, NBJ is classified as a Cluster F, Subcluster F1 phage and shares 70% of its encoded gene phamilies with Girr, another F1 mycobacteriophage that was recently analyzed in a genome-wide overexpression screen and found to encode 29 diverse gene products capable of inhibiting growth of M. smegmatis. Similar functional screens in other mycobacteriophages have uncovered a growing repertoire of diverse, phage-encoded bacterial growth inhibitors, providing prime candidates for dissecting novel bacterial-phage interactions. An arrayed overexpression library encoding all 102 genes was constructed and systematically screened using a plate-based cytotoxicity assay, identifying 29 genes that inhibit mycobacterial growth. Because mycobacteriophage genomes are also known to encode systems involved in phage-phage competition, we conducted additional phage defense assays for a subset of NBJ genes in our library. This analysis confirmed homotypic immunity by the predicted immunity repressor and identified gp45 as a second defense factor that protects against Cluster F phages and promotes NBJ lysogen stability. Additional mechanistic analyses indicate that gp45 functions in a secretion-dependent manner, acting after phage adsorption to reduce productive infection and early phage transcript accumulation. Escape mutant and bacterial two-hybrid analyses implicate Cluster F tape measure proteins as the likely target of gp45, consistent with disruption of an early step in phage genome delivery. Finally, we extended our analysis to explore the essentiality of all identified host-modulating genes in the NBJ life cycle, using CRISPR-enhanced recombineering to generate phage deletion mutants, revealing two host modulators that are critical for lytic growth. Conducted as part of the SEA-GENES undergraduate research consortium, this study adds to a growing functional genomics resource and provides new insights into the complex interactions between phage gene products and the mycobacterial host cell.

microbiology↗
bioRxiv2025

Functional Diversity in GII.4 Norovirus Entry: HBGA Binding and Capsid Clustering Dynamics

Human noroviruses (HuNoVs), especially GII.4 strains, are the leading cause of acute viral gastroenteritis worldwide, yet no approved vaccines or antivirals exist. The pandemic GII.4 Sydney 2012 strain enters cells via membrane wounding and clathrin-independent carrier (CLIC)-mediated endocytosis, but it is unclear whether this entry mechanism is conserved across GII.4 variants. We compared early binding and entry of multiple GII.4 variants using wildtype and mutant GII.4 virus-like particles (VLPs) and modified human intestinal enteroid (HIE) cultures. Only a subset of GII.4 variants, including GII.4 Sydney, form distinct, HBGA-dependent capsid clusters on the cell surface. Clustering strains display significantly enhanced membrane wounding and endocytosis compared to non-clustering strains and outcompete non-clustering strains in replication assays as shown by complete inhibition of GII.4 Sydney replication. Using mutant VLPs and a HBGA non-binding mutant (R345A), we identified two residues, V333 and R339, in the VP1 protruding domain as critical mediators of clustering and entry. Mutations of these residues disrupt clustering and endocytosis without affecting HBGA binding, suggesting a role in post-attachment processes. While clustering and endocytosis are contingent upon VLP binding to HBGAs, inhibitor studies show they are independent of host protein glycosylation and are driven by lipid raft remodeling regulated by cholesterol and ceramides. Quantitative analyses across multiple GII.4 variants reveal an apparent dichotomy between clustering and non-clustering phenotypes, with clustering variants exhibiting higher entry competence. This distinction offers insight into strain-specific cell entry mechanisms and may aid in identifying the elusive proteinaceous HuNoV cellular receptor(s) supporting targeted therapeutic development.

microbiology↗
bioRxiv2024

Diphenyleneiodonium chloride inhibits MYCN-amplified neuroblastoma by targeting MYCN induced mitochondrial alterations

High-risk neuroblastoma is one of the most lethal childhood cancers. Half of these tumors are driven by MYCN gene amplification (MNA). Despite intensive chemo- and radiotherapy, only 40% of patients survive, and they often suffer from severe long-term side effects of these genotoxic treatments. Thus, new therapies are needed that are less toxic and more efficacious. Here, we identified diphenyleneiodonium (DPI) as a tool compound that preferentially targets MNA neuroblastoma. Using proteomic assays we investigated the DPI mode of action, finding that DPI induces the proteasomal degradation of MYCN and could reverse some alterations induced by high levels of MYCN. These include profound changes in the expression of proteins participating in the mitochondrial electron transport chain. Metabolic and biological assays suggested that alterations in mitochondrial function and the associated production of reactive oxygen species (ROS) are critical DPI targets in the context of MNA. DPI reduced the survival, and malignant transformation of neuroblastoma across a panel of cell lines at clinically achievable concentrations. DPI also shrank tumors and prevented metastatic spread in zebrafish models of MYCN-driven neuroblastoma. These findings suggest that processes impacted by complex I inhibitors could be valuable new targets for the development of non-genotoxic drugs against high-risk MNA neuroblastoma. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=151 SRC="FIGDIR/small/619268v2_ufig1.gif" ALT="Figure 1"> View larger version (32K): org.highwire.dtl.DTLVardef@167e953org.highwire.dtl.DTLVardef@10749e4org.highwire.dtl.DTLVardef@1842598org.highwire.dtl.DTLVardef@c05d82_HPS_FORMAT_FIGEXP M_FIG C_FIG

cancer biology↗
bioRxiv2024

Phytochemical Analysis and Nematicidal Activity of Medicinal Plants Against Meloidogyne javanica

This study examined the nematicidal potential of ethanol extracts derived from four plants, namely Actinidia deliciosa (Chev.), Carica papaya, Citrus paradise, and Raphanus sativus, against the root-knot nematode Meloidogyne javanica. Experimental results indicated a significant reduction in the survival of second-stage juveniles of M. javanica when exposed to these plant extracts. Notably, concentrations of 2% and 1% were found to be more effective in comparison to 0.5% and 0.25%, yielding statistically significant outcomes. Furthermore, the mortality rate of the nematodes exhibited a direct correlation with the duration of exposure for most plant extracts. The fruit extracts obtained from A. deliciosa, C. papaya, C. paradise, and R. sativus demonstrated a substantial decrease in nematode infestation compared to the control group. The nematicidal activity of KI3a (BuOH) significant from 0.1356 to 1.9076 over 72 hours, while P-KI-2a (EtOAc) showed rose from 0.1206 to 0.6456. In compare, GF-3 (EtOAc) and Carbofuran to 0.0179 and 0.0098, respectively. PA-1 (MeOH) and RS-3 (EtOAc) showed modest nematicidal potential. This research emphasizes the efficacy of plant extract in mitigating root-knot nematode infestation, offering a promising alternative to chemical managements. Additionally, comprehensive phytochemical analysis should be achieved to recognize the active chemical constituents responsible for their effects. Exploring synergistic effects between these compounds and other natural or synthetic agents could further improve their potential applications in sustainable agriculture, improving crop protection and promoting eco-friendly practices.

plant biology↗
bioRxiv2024

MYCN inhibits TrkC-mediated differentiation in neuroblastoma cells via disruption of the PKA signalling pathway

Neuroblastoma is a complex paediatric cancer with a spectrum of clinical outcomes ranging from spontaneous regression to aggressive metastatic disease. Low-risk patients achieve over 90% survival with no or minimal treatment, while high-risk patients face less than 50% survival despite intensive multimodal therapy. Half of the high-risk cases harbour amplification of the MYCN oncogene. In addition to MYCN status, Trk receptors have also been linked to prognosis. TrkA expression is seen with low-risk cases while TrkB expression often occurs in high-risk MYCN-amplified NB. While TrkA and TrkB are well studied in NB, the role of TrkC in neuroblastoma genesis is not clear. Therefore, this study investigates the interplay between MYCN status and NT-3/TrkC signalling in neuroblastoma. Using a panel of neuroblastoma cell lines with varying MYCN levels, we found that TrkC activation leads to neuronal differentiation of MYCN non-amplified cells, whereas it promotes proliferation of MYCN-amplified cells. Temporal phosphoproteomics revealed differential activation of the PKA pathway, which was crucial for TrkC-mediated differentiation. Manipulating the PKA pathway altered cell fate outcomes, underscoring its role. In MYCN-amplified cells, MYCN knockdown increased PKA and CREB activity, shifting the phenotype towards differentiation. Analysis of neuroblastoma patient data showed lower expression of PKA pathway genes in MYCN-amplified tumours. Additionally, miR-221, upregulated by MYCN, was identified as a suppressor of the PKA/CREB pathway. These findings highlight the context-dependent nature of NT-3/TrkC signalling influenced by MYCN; and suggest therapeutic potential in targeting the PKA pathway to induce differentiation of high-risk MYCN-amplified neuroblastoma.

cancer biology↗
bioRxiv2024

Multimodal mechano-microscopy reveals mechanical phenotypes of breast cancer spheroids in three dimensions

Cancer cell invasion relies on an equilibrium between cell deformability and the biophysical constraints imposed by the extracellular matrix (ECM). However, there is little consensus on the nature of the local biomechanical alterations in cancer cell dissemination in the context of three-dimensional (3D) tumor microenvironments (TME). While the shortcomings of two-dimensional (2D) models in replicating in situ cell behavior are well known, 3D TME models remain underutilized because contemporary mechanical quantification tools are limited to surface measurements. Here, we overcome this major challenge by quantifying local mechanics of cancer cell spheroids in 3D TMEs. We achieve this using multimodal mechano-microscopy, integrating optical coherence microscopy-based elasticity imaging with confocal fluorescence microscopy. We observe that non-metastatic cancer spheroids show no invasion while showing increased peripheral cell elasticity in both stiff and soft environments. Metastatic cancer spheroids, however, show ECM-mediated softening in a stiff microenvironment and, in a soft environment, initiate cell invasion with peripheral softening associated with early metastatic dissemination. This exemplar of live-cell 3D mechanotyping supports that invasion increases cell deformability in a 3D context, illustrating the power of multimodal mechano-microscopy for quantitative mechanobiology in situ.

bioengineering↗
bioRxiv2022

Maternal and/or direct supplementation with a combination of a casein hydrolysate and yeast β-glucan on post-weaning performance and intestinal health in the pig

A 2 x 2 factorial experiment was conducted to investigate the effect of maternal supplementation from day 83 of gestation and/or direct supplementation from weaning of a bovine casein hydrolysate plus a yeast {beta}-glucan (CH-YBG) on pig performance and intestinal health on day ten post-weaning. Twenty cross bred gilts (Large White x Landrace) were randomly assigned to one of two dietary groups (n = 10 gilts/group): basal diet (basal sows) and basal diet supplemented with CH-YBG (supplemented sows) from day 83 of gestation until weaning (2g/sow/day). At weaning, 120 pigs (6 pigs/sow) were selected. The two dam groups were further divided, resulting in four experimental groups (10 replicates/group; 3 pigs/pen) as follows: 1) BB (basal sows + basal pigs); 2) BS (basal sows + supplemented pigs); 3) SB (supplemented sows + basal pigs); 4) SS (supplemented sows + supplemented pigs). Supplemented pigs were offered 0.5g CH-YBG/kg of feed for 10 days post-weaning. On day 10 post-weaning, 1 pig/pen was humanely sacrificed and samples were taken from the gastrointestinal tract for analysis. Pigs weaned from supplemented sows (SS, SB) had reduced faecal scores and incidence of diarrhoea (P<0.05) compared to pigs weaned from basal sows (BB, BS), with SS pigs not displaying the transient rise in faecal scores seen in the other three groups from day 3 to day 10 post-weaning (P<0.05). Pigs weaned from supplemented sows had reduced feed intake (P<0.05), improved feed efficiency (P<0.05), increased butyrate concentrations (P<0.05), increased abundance of Lactobacillus (P<0.05) and decreased abundance of Enterobacteriaceae and Campylobacteraceae (P<0.05) compared to pigs weaned from basal sows. In conclusion, a combination of both maternal and direct supplementation with CH-YBG is the desirable feeding strategy to ameliorate the post-weaning challenge in pig production systems.

developmental biology↗