Biology subjects

Nawaz, S.

Publications and source records attributed to Nawaz, S..

7 recordsLinked to original sources

bioRxiv2026

PlantNetX: A web-based transcriptomic resource integrating bulk and single-cell RNA-seq for plant functional genomics.

Bulk RNA sequencing and single-cell RNA sequencing provide complementary information on tissue and cell-type-specific gene expression. Bulk RNA sequencing enables the construction of gene association networks that identify co-expressed genes involved in shared pathways, whereas single-cell RNA sequencing maps their expression to cell types. However, most platforms provide access to either bulk RNA sequencing or single-cell RNA sequencing analysis, making it difficult to connect tissue-level co-expression with cell-type-specific expression. PlantNetX was developed as a web-based platform that integrates both data types. Although PlantNetX currently focuses on rice (Oryza sativa) and includes 70 quality-controlled RNA sequencing datasets comprising 1,198 sequencing libraries, together with nine single-cell RNA sequencing datasets containing more than 580,000 cells, including recently released datasets not consistently represented in existing platforms, it was designed to incorporate additional plant species, datasets, and analytical tools. PlantNetX provides Mutual Rank-based co-expression analysis, global and tissue-specific gene association networks, interactive visualization, and cell-type-specific expression summaries. The platform was validated with published examples of plant cell-wall biosynthesis and root-hair growth and retrieved gene association and expression patterns. Under standardized testing conditions, PlantNetX had a shorter mean response time than the other databases assessed. PlantNetX will support research in plant cell-wall biosynthesis, pathway discovery, functional genomics, and crop improvement. HighlightsPlantNetX integrates gene co-expression networks with cell-type expression, enabling fast identification and biological interpretation of candidate genes across tissues and individual cells. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=120 SRC="FIGDIR/small/741827v1_ufig1.gif" ALT="Figure 1"> View larger version (60K): org.highwire.dtl.DTLVardef@1207e06org.highwire.dtl.DTLVardef@31c3acorg.highwire.dtl.DTLVardef@12560b0org.highwire.dtl.DTLVardef@eeee7c_HPS_FORMAT_FIGEXP M_FIG C_FIG

bioinformatics↗

bioRxiv2024

Reprogramming Human Inflammatory Macrophages in Symptomatic Carotid Stenosis: Potential Mechanisms for Stabilisation of Atherosclerotic Carotid Plaques

BackgroundInflammation is a precursor to atherosclerotic plaque destabilisation, leading to ischaemic events such as stroke. Macrophage phenotypes can be altered by the microenvironment, and certain anti-inflammatory agents may, therefore, stabilise plaques and reduce the risk of recurrent ischaemic events. MethodsThirteen carotid plaques were obtained from stroke/ Transient Ischaemic Attack (TIA) patients undergoing carotid endarterectomy. An immunofluorescence stain was used to identify common macrophage markers (pan macrophage: CD68, pro-inflammatory: CD86, anti-inflammatory: MRC1), and a novel analysis technique was used to measure the prevalence of macrophage phenotypes in carotid plaques in relation to other histological features of instability. An in vitro model of human blood-derived macrophages was also developed to evaluate the effect of statins and glucocorticoids on macrophage-specific markers using RT-qPCR, Western Blot and immunofluorescence stain. The physiological effect of dexamethasone was further evaluated on macrophages and human carotid plaques cultured ex vivo. ResultsThe macrophage population (CD68+) in the carotid plaques was dominated by "double-positive" (CD86+MRC1+) macrophages (67.8%), followed by "M1-like" (CD86+MRC1-) (16.5%), "M2-like" (CD86-MRC1+) (8.7%) and "double-negative" (CD86-MRC1-) (7.0%) macrophages. M1-like macrophages were more prevalent in unstable plaque sections than stable ones (p=0.0022). Exposure to dexamethasone increased macrophage MRC1 gene expression in vitro and ex vivo. Dexamethasone also reduced Oxidised Low-Density Lipoprotein Receptor 1 (OLR1) gene and protein expression, leading to a decreased ox-LDL uptake in foam cell assays. This was, in turn, associated with reduced lipid uptake in macrophages, as shown by Oil Red O staining. ConclusionsHuman macrophages may be "switched" to a less inflammatory phenotype by exposure to clinically relevant concentrations of glucocorticoid, potentially mediated by a reduction in Oxidised LDL uptake. This effect was not observed following macrophage exposure to statins. Glucocorticoids may have a future role in preventing ischaemic events in patients with advanced atherosclerosis. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=77 SRC="FIGDIR/small/598440v1_ufig1.gif" ALT="Figure 1"> View larger version (30K): org.highwire.dtl.DTLVardef@bb389borg.highwire.dtl.DTLVardef@18151b8org.highwire.dtl.DTLVardef@11c28a1org.highwire.dtl.DTLVardef@1af6acd_HPS_FORMAT_FIGEXP M_FIG C_FIG HighlightsO_LIA high prevalence (68% in this study) of carotid plaque macrophages express both pro-inflammatory (CD86) and anti-inflammatory (MRC1) markers. These may represent a novel macrophage population. C_LIO_LIHuman macrophages may be "reprogrammed" to a less inflammatory phenotype following exposure to glucocorticoids. C_LIO_LIDexamethasone increased MRC1 and decreased OLR1 expression in macrophages derived from human blood samples in vitro and in cells derived from cultured human carotid plaque tissue ex vivo. This was associated with reduced oxLDL uptake and reduced lipid accumulation in the macrophages. C_LIO_LIDexamethasone has the potential to stabilise carotid atherosclerotic plaques in humans. C_LI

cell biology↗

bioRxiv2024

Absence seizures and sleep abnormalities in a rat model of GRIN2B neurodevelopmental disorder

Pathogenic mutations in GRIN2B are an important cause of severe neurodevelopmental disorders resulting in epilepsy, autism and intellectual disability. GRIN2B encodes the GluN2B subunit of N-methyl-D-aspartate receptors (NMDARs), which are ionotropic glutamate receptors critical for normal development of the nervous system and synaptic plasticity. Here, we characterized a novel Grin2b heterozygous knockout rat model with 24-hour EEG recordings. We found rats heterozygous for the deletion (Grin2b+/-) had a higher incidence of spontaneous spike and wave discharges, the electrographic correlate of absence seizures, than wild-type rats (Grin2b+/+). Spike and wave discharges were longer in duration and displayed higher overall spectral power in Grin2b+/- when compared to those in Grin2b+/+ animals. Heterozygous mutant rats also had abnormal sleep-wake brain state dynamics over the circadian cycle. Specifically, we identified a reduction in total rapid eye movement sleep and, altered distributions of non-rapid eye movement sleep and wake epochs, when compared to controls. This was accompanied by an increase in overall spectral power during non-rapid eye movement sleep in Grin2b+/-. The sleep-wake phenotypes were largely uncorrelated to the incidence of spike and wave discharges. We then tested the antiseizure efficacy of ethosuximide, a T-type voltage-gated calcium channel blocker used in the treatment of absence seizures, and memantine, a noncompetitive NMDAR antagonist currently explored as a mono or adjunctive treatment option in NMDAR related neurodevelopmental disorders. Ethosuximide reduced the number and duration of spike and wave discharges, while memantine did not affect the number of spike and wave discharges but reduced their duration. These results highlight two potential therapeutic options for GRIN2B related epilepsy. Our data shows the new rat Grin2b haploinsufficiency model exhibits clinically relevant phenotypes. As such, it could prove crucial in deciphering underlying pathological mechanisms and developing new therapeutically translatable strategies for GRIN2B neurodevelopmental disorders.

neuroscience↗

bioRxiv2023

Electrophysiological characterisation of iPSC-derived human β-like cells and an SLC30A8 disease model.

iPSC-derived human {beta}-like cells (BLC) hold promise for both therapy and disease modelling, but their generation remains challenging and their functional analyses beyond transcriptomic and morphological assessments remain limited. Here, we validate an approach using multicellular and single cell electrophysiological tools to evaluate BLCs functions. The Multi-Electrode Arrays (MEAs) measuring the extracellular electrical activity revealed that BLCs are electrically coupled, produce slow potential (SP) signals like primary {beta}-cells that are closely linked to insulin secretion. We also used high-resolution single-cell patch-clamp measurements to capture the exocytotic properties, and characterize voltage-gated sodium and calcium currents. These were comparable to those in primary {beta} and EndoC-{beta}H1 cells. The KATP channel conductance is greater than in human primary {beta} cells which may account for the limited glucose responsiveness observed with MEA. We used MEAs to study the impact of the type 2 diabetes protective SLC30A8 allele (p.Lys34Serfs*50) and found that BLCs with this allele have stronger electrical coupling. Our data suggest that with an adapted approach BLCs from pioneer protocol can be used to evaluate the functional impact of genetic variants on {beta}-cell function and coupling. Article highlightsO_ST_ABSWhy did we undertake this study?C_ST_ABSiPSC-derived beta like cells (BLCs) from pioneering protocols are known for variable {beta}-cell functionality and mixed cell populations which greatly limits downstream functional assessment. To overcome this challenge, we used electrophysiological tools to provide a detailed functional assessment of BLCs. We then wanted to apply this approach to identify additional functional differences from BLCs carrying a protective Type 2 Diabetes SLC30A8 allele. What is the specific question(s) we wanted to answer?Can an electrophysiological approach provide detailed functional characterisation of iPSC-derived BLCs? Is this approach sensitive enough to capture functional differences resulting from SLC30A8 loss of function (lof)? What did we find?We found that BLCs generated from pioneer protocol shared electrophysiological features with human pancreatic {beta}-cells, and that a T2D-protective SLC30A8 lof allele improves the electrical coupling activity of human {beta}-cells. What are the implications of our findings?Our findings validate the use of intra- and extra-cellular electrophysiology to assess and monitor the functions of BLCs. Our approach opens the perspective of using MEAs to live-monitor the differentiation quality of iPSC-derived BLCs and to determine the functional consequences of diabetes-associated variants.

cell biology↗

bioRxiv2023

LRRC23 loss-of-function impairs radial spoke 3 head assembly and causes defective sperm motility underlying male infertility

Radial spokes (RS) are T-shaped multiprotein complexes on the axonemal microtubules. Repeated RS1, RS2, and RS3 couple the central pair to modulate ciliary and flagellar motility. Despite the cell type specificity of RS3 substructures, their molecular components remain largely unknown. Here, we report that a leucine-rich repeat-containing protein, LRRC23, is an RS3 head component essential for its head assembly and flagellar motility in mammalian spermatozoa. From infertile male patients with defective sperm motility, we identified a splice site variant of LRRC23. A mutant mouse model mimicking this variant produces a truncated LRRC23 at the C-terminus that fails to localize to the sperm tail, causing male infertility due to defective sperm motility. LRRC23 was previously proposed to be an ortholog of the RS stalk protein RSP15. However, we found that purified recombinant LRRC23 interacts with an RS head protein RSPH9, which is abolished by the C-terminal truncation. Evolutionary and structural comparison also shows that LRRC34, not LRRC23, is the RSP15 ortholog. Cryo-electron tomography clearly revealed that the absence of the RS3 head and the sperm-specific RS2-RS3 bridge structure in LRRC23 mutant spermatozoa. Our study provides new insights into the structure and function of RS3 in mammalian spermatozoa and the molecular pathogenicity of LRRC23 underlying reduced sperm motility in infertile human males.

cell biology↗

bioRxiv2022

Enhanced hippocampal LTP but typical NMDA receptor and AMPA receptor function in a novel rat model of CDKL5 deficiency disorder

Mutations in the X-linked gene cyclin-dependent kinase-like 5 (CDKL5) cause a severe neurological disorder characterised by early-onset epileptic seizures, autism and intellectual disability (ID). Impaired hippocampal function has been implicated in other models of monogenic forms of autism spectrum disorders and ID and is often linked to epilepsy and behavioural abnormalities. Many individuals with CDKL5 deficiency disorder (CDD) have null mutations and complete loss of CDKL5 protein, therefore in the current study we used a novel Cdkl5 KO rat model to elucidate the impact of CDKL5 loss on cellular excitability and synaptic function of CA1 pyramidal cells (PCs). We hypothesised abnormal pre and/or post synaptic function underlie the enhanced LTP we observe in the hippocampus of Cdkl5 KO rats. We tested this hypothesis using a combination of extracellular and whole-cell electrophysiological recordings, biochemistry, and histology. We show that NMDA receptor function and subunit expression are unaltered throughout development, and Ca2+ permeable AMPA receptor mediated currents are unchanged in Cdkl5 KO rats. We observe reduced mEPSC frequency accompanied by increased spine density in basal dendrites of CA1 PCs, however we find no evidence supporting an increase in silent synapses when assessed using a minimal stimulation protocol in slices. Additionally, we found no change in paired-pulse ratio, consistent with normal release probability in Cdkl5 KO rats and supported by typical expression of pre-synaptic proteins in synaptosome preparations. Together these data indicate a role for CDKL5 in hippocampal synaptic function and raise the possibility that altered intracellular signalling rather than synaptic deficits might contribute to the altered plasticity.

neuroscience↗

bioRxiv2020

Chromatin 3D interaction analysis of the STARD10 locus unveils FCHSD2 as a new regulator of insulin secretion

Genome-wide association studies have identified thousands of genetic variants associated with type 2 diabetes (T2D) risk. Using chromatin conformation capture we show that an enhancer cluster in the STARD10 T2D locus forms a defined 3D chromatin domain. A 4.1 Kb region within this region, carrying five disease-associated variants, physically interacts with CTCF-binding regions and with an enhancer possessing strong transcriptional activity. Analysis of human islet 3D chromatin interaction maps identifies FCHSD2 gene as an additional target of the enhancer cluster. CRISPR-Cas9-mediated deletion of the variant region, or of an associated enhancer, in human pancreatic beta cells impaired glucose-stimulated insulin secretion. Expression of both STARD10 and FCHSD2, but not ARAP1, was reduced in cells harboring CRISPR deletions, and expression of STARD10 and FCHSD2 was associated with the possession of variant alleles in human islets. Finally, CRISPR-Cas9-mediated loss of STARD10 or FCHSD2 impaired regulated insulin secretion. Thus, multiple genes at the STARD10 locus influence {beta} cell function.

cell biology↗