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Biology subjects

Ahmad, M.

Publications and source records attributed to Ahmad, M..

At least 19 recordsLinked to original sources

Semidilute Polymer Scaling Controls PEG-induced Protein Phase Separation

When does polyethylene glycol (PEG) mass concentration provide a common measure of crowding strength across polymer chain lengths? Here we use bovine serum albumin (BSA) and PEG as a controlled depletion system and measure both coexisting phases to determine binodals, tie lines, and critical points across PEG molecular weights of 4--100 kDa. Phase diagrams for PEG 20, 35, and 100 kDa approximately collapse in mass-concentration coordinates, consistent with semidilute polymer scaling. Generalized free volume theory (GFVT) describes the BSA binodals and tie lines. It also approximately describes binodals for three additional globular proteins in PEG 35K, extending the comparison across proteins spanning 7.2--66.4 kDa. For the binary PEG mixtures tested, GFVT approximates the binodals using an effective polymer size corresponding to the weight-average molecular weight. These results identify a regime in which PEG mass concentration (g/L) provides a transferable measure of depletion-induced crowding.

biophysics↗

Mitochondrial dysfunction and impaired osteogenic capacity define stress-induced osteoblast senescence

Cellular senescence has emerged as a key contributor to age-related skeletal deterioration; however, the defining characteristics of senescent osteoblasts remain incompletely understood, hindering efforts to identify the cellular mechanisms that drive age-associated bone loss and potential therapeutic targets. Here, we compared doxorubicin- and hydrogen peroxide-induced senescence and established a robust in vitro osteoblast model that enables the stable maintenance of stress-induced premature senescence. Doxorubicin-treated MC3T3-E1 cells exhibited persistent growth arrest, a pronounced senescence-associated secretory phenotype (SASP), and impaired osteogenic function accompanied by marked mitochondrial dysfunction, including reduced respiratory capacity and altered mitochondrial morphology. Transcriptomic comparison with aged murine bone revealed a partial overlap with in vivo aging-associated gene expression signatures, particularly among genes involved in extracellular matrix organization and skeletal development, supporting the physiological relevance of the model. Importantly, key features of senescence, including the senescence-associated mitochondrial phenotype and impaired osteogenic capacity, were recapitulated in primary human osteoblasts. Collectively, these findings establish a robust model of stress-induced osteoblast senescence and demonstrate that senescent osteoblasts exhibit mitochondrial dysfunction, impaired osteogenic capacity, and molecular features that resemble aspects of skeletal aging.

cell biology↗

Spatial Patterning of Otic Vesicles in Human Inner Ear Organoids

During development axial patterning of the otic vesicle sets up the blueprint for the adult inner ear. While human inner ear organoids recapitulate the initial steps of ear formation, whether and how vesicles are properly regionalised remains unknown. Here, we combine re-analysis of published single-cell transcriptomic datasets with 2D and 3D imaging to assess axial patterning in human stem-cell-derived otic vesicles using a new PAX2-reporter line. We find that the transcriptional signatures of individual cells do not conform to clearly defined regional characteristics, with some cells co-expressing conflicting regional markers, while markers that should be co-expressed are not. Some vesicles establish spatially segregated molecular domains. However, patterning is incomplete, stochastic, and restricted to a subset of vesicles. Within the same aggregates, patterned axes lack consistent orientation. Together, our findings point to incomplete segregation of transcriptional programs that are mutually exclusive in vivo and to the absence of morphogen gradients that are crucial for otic vesicle patterning. >One sentence summaryIn human inner ear organoids, otic vesicles show incomplete and disorganized regionalisation pointing to fundamental limits of self-organization, while suggesting new routes to increase cell complexity in vitro.

developmental biology↗

Climate change drives contrasting redistribution patterns in endemic and endangered Himalayan Gentiana

O_LIRapid climate warming threatens mountain biodiversity, particularly species with narrow climatic niches and limited dispersal capacity. Mountain ecosystems are especially vulnerable because steep environmental gradients restrict opportunities for species redistribution. The Kashmir Himalaya, a globally important biodiversity hotspot experiencing accelerated warming, has already undergone an increase of approximately 0.8 {degrees}C during the 20th century and is projected to warm by 2.5-2.8 {degrees}C by the 2050s. Despite climatic changes, future persistence of many threatened plants remains poorly understood. C_LIO_LIHere, we evaluate present and future habitat suitability for two Himalayan taxa, Gentiana cachemirica, an endemic species, and Gentiana kurroo, a critically endangered species. Specifically, we quantify how climate change and topographic variability influence species distribution and persistence under multiple emission scenarios. C_LIO_LIWe applied species distribution models (SDMs) to presence data of both species and forecasted habitat suitability under four Shared Socioeconomic Pathways (SSP126, SSP245, SSP370, and SSP585). We hypothesized that G. cachemirica, a narrow-niche, low-dispersal species, is expected to lose habitat due to thermal sensitivity, while G. kurroo may persist or expand under favorable scenarios because of broader tolerance and higher dispersal. We also expected microclimatic refuges to buffer populations, whereas extreme warming would accelerate habitat decline. C_LIO_LIThe predictions of our SDMs under current conditions indicate a highly restricted and fragmented habitat for G. cachemirica, covering 651 km{superscript 2}, but a broader suitable area (2,452 km{superscript 2}) for G. kurroo. In agreement with our hypotheses, our forecasts indicate severe habitat contraction (55-70%) for G. cachemirica across all SSPs, but scenario-dependent responses for G. kurroo, including modest expansion under low-emission scenarios and severe declines under high-emission scenarios. Centroid analyses suggest pronounced climate-driven range shifts, with G. kurroo projected to migrate up to 33 km toward the east-southeast by 2100, while G. cachemirica is projected to display limited dispersal capacity. C_LIO_LISynthesis. Our findings suggest that climatic niche breadth, dispersal limitation, and topographic buffering strongly mediate species responses to warming in mountain ecosystems. Endemic specialists are projected to experience disproportionate habitat fragmentation and range restriction, highlighting the importance of conserving climatic refugia and elevational connectivity under rapid environmental change. C_LI

ecology↗

Splice-Aware Optimization Prevents Pervasive Missplicing of Natural and Synthetic cDNAs

Heterologous gene expression is widely used across biology and medicine, and often relies on codon optimization to increase protein yields. Here we uncover missplicing as a common and largely unrecognized failure mode of heterologous expression. Using systematically designed libraries comprising over 5,000 synthetic reporter genes and natural human cDNAs, we find that the majority of gene variants expressed in a human cell line are at least partially spliced, and in many variants the spliced isoform dominates, reducing protein output or ablating expression entirely. By analysing sequence determinants of expression across multiple human cell lines, we uncover a hierarchical architecture of regulatory control, where GC content establishes baseline mRNA levels, local sequence features influence splicing, and tissue-specific codon adaptation to tRNA pools fine-tunes translation efficiency. These findings enable us to develop predictive models of expression and splicing, benchmark current optimization strategies, and design a splice-aware optimization algorithm that substantially improves transgene performance.

synthetic biology↗

Bark beetle protein elicitors trigger biphasic immune responses in Norway spruce seedlings

Norway spruce (Picea abies) responds to attacks by the spruce bark beetle (Ips typographus) through the rapid activation of local defense mechanisms, but field studies can be difficult to standardize due to variable attack pressure and environmental heterogeneity. Here, we developed a phytotron-based assay that mimics early beetle-associated stress using insect-derived protein extracts, enabling reproducible molecular analyses under controlled conditions. Ten-week-old spruce seedlings were stem-treated with mock buffer or beetle protein extracts, followed by transcriptomic analyses of stem tissues and targeted metabolomic profiling of needles at 2 and 48 h post-inoculation. RT-qPCR analysis revealed rapid transcriptional activation of signaling and defense genes in Norway spruce, with NP-40-based protein extracts producing the most consistent early response. RNA-seq analysis revealed transcriptional dynamics, with 488 differentially expressed genes detected at 2 h and 84 at 48 h post-inoculation relative to mock-treated controls. Early responses at 2 h were characterized by activation of genes associated with immune perception and signal transduction. By 48 h, the response shifted toward accumulation of transcripts encoding defense proteins such as chitinases, defensins, proteinase inhibitors, and pathogenesis-related (PR) proteins. Importantly, a substantial proportion of differentially expressed genes overlapped with those previously identified in mature Norway spruce trees during pioneer bark beetle attack under field conditions, supporting the biological relevance of the assay. In contrast, targeted analyses of secondary metabolites performed in needle tissue revealed limited systemic changes across time points, suggesting that early induced defenses may remain largely localized to the stem. Together, these results demonstrate that beetle-derived proteins trigger a rapid and temporally structured defense response in Norway spruce seedlings and establish a reproducible elicitor-based platform that may provide a broadly applicable framework for investigating bark beetle-induced defenses across conifers. HighlightBark beetle protein elicitors trigger temporally structured immune responses in Norway spruce seedlings that overlap with responses observed in mature trees, with rapid immune signaling at 2 h followed by defense protein accumulation at 48 h.

plant biology↗

HOMEOSTATIC COUPLING OF CORTICAL AND BRAINSTEM DELTA RHYTHMS IN SLEEPING INFANT RATS

The emergence of the cortical delta rhythm (1-4 Hz) during quiet sleep (QS) is a major milestone in brain development. In rats, this milestone is achieved between 8 and 12 days of postnatal (P) age. We previously reported an age-dependent increase in PZ delta-rhythmic activity that is synchronized with cortical delta and entrained by breathing. Here, we ask whether this long-distance synchrony persists in response to perturbations to sleep homeostasis or respiration. First, using male and female P12 rats, we investigated the coupling strength between frontal cortex and PZ in response to a short but intense period of sleep deprivation. During recovery sleep, we observed a rebound in delta power in both PZ and cortex, even in the absence of increased QS duration, indicating that PZ and cortical delta power are equivalent markers of homeostatic sleep regulation. Analyses of phase-locking and lagged cross-correlation revealed persistent temporal coupling between the two rhythms such that cortical delta reliably lagged PZ delta regardless of changes in sleep pressure. Curiously, we also observed an increase in breathing depth during recovery sleep, which we confirmed in a separate cohort of pups. Next, using mild hypercapnia (5% CO2) to alter breathing frequency and depth, we produced decreases in cortical and PZ delta power along with decreases in the depth of breathing. These findings provide additional support for the notion that PZ and cortical delta rhythms function as distantly interconnected components within a developmentally emerging sleep-homeostatic system that is also intimately tied with the brainstem respiratory network. SIGNIFICANCE STATEMENTWe reported previously that the delta rhythm that defines slow-wave sleep is not confined to the forebrain but also occurs synchronously in the medulla. This study in infant rats uses two perturbations to assess the coupling strength of cortical and brainstem delta. Using sleep deprivation and hypercapnia, we show that delta power increases or decreases in lockstep in the two regions, respectively. Our results reinforce the notion that delta across these two regions is strongly coupled and adds a new dimension to our understanding of the interconnectedness of the delta rhythm and respiration.

neuroscience↗

Gram-positive bacterial cell wall components inhibit herpes simplex virus infection.

The role of the mucosal microbiome in viral infections remains unclear. Genital herpes, caused by herpes simplex virus 1 and 2 (HSV-1 and HSV-2), is among the most prevalent sexually transmitted infections worldwide. Despite evidence linking vaginal Lactobacillus to protection against sexually transmitted viruses, the specific microbial components and mechanisms that mediate this defense are not well understood. Here, we show that multiple cell wall components from diverse gram-positive bacteria, including lactobacilli, inhibit HSV-1 and HSV-2 infection in cells and in a mouse model of genital herpes infection. Peptidoglycan (PG) and lipoteichoic acid (LTA), both major components of the gram-positive bacterial cell wall, significantly reduced HSV infectivity in vitro and improved survival and disease outcomes in mice. We further showed that Lactobacillus crispatus surface layer proteins SlpA and SlpB bind HSV-1 and inhibit infection. Antiviral effects of cell wall components were dose-dependent, relied on intact PG structure, and, in the case of PG and LTA, were independent of TLR2-mediated host signaling. Collectively, our findings identify a species-independent antiviral function for gram-positive bacterial cell wall components against HSV and suggest that the composition of the mucosal microbiome may play an underappreciated role in suppressing mucosal herpes infection in humans.

microbiology↗

Unravelling the intraspecific variation in drought responses in seedlings of European black pine (Pinus nigra J.F. Arnold)

O_LIUnderstanding intraspecific variation in drought tolerance is essential for predicting the adaptive capacity of forest species under climate change. Yet, the molecular basis of this variation remains poorly understood in ecologically and economically important conifers. C_LIO_LIWe integrated high-throughput phenotyping with metabolomics and transcriptomics under standardized soil drying to investigate drought responses across nine climatically distinct provenances of the conifer Pinus nigra. We tested whether drought tolerance--measured as decline in maximum quantum yield of the photosystem II (Fv/Fm)--varies among provenances, follows a climatic cline, and involves trade-off with growth. To identify the underlying molecular basis, we performed metabolomics and transcriptomics in four provenances representing contrasting drought tolerance. C_LIO_LIDrought tolerance varied significantly among provenances and was decoupled from growth, yet showed no differentiation along the climatic cline. Drought tolerant provenances differed from sensitive ones in both constitutive and drought-induced levels of flavonoid and diterpene metabolites. Transcriptomic profiles further highlighted provenance-specific differences in gene expression related to flavonoids. C_LIO_LIOur results demonstrate the utility of integrating automated phenotyping with molecular profiling to uncover the metabolic basis of drought adaptation, laying the groundwork for targeted studies on metabolite function and tolerance strategies in non-model conifers. C_LI

plant biology↗

Pioneer bark beetle attacks induce multifaceted localized defense responses in Norway spruce

Conifer forests worldwide are increasingly threatened by environmental stressors, especially bark beetle infestations. In a novel experimental field setup with controlled attacks on clonal trees, we investigated the molecular defense responses of 35-year-old Norway spruce trees to colonization attempts by the spruce bark beetle, Ips typographus. A multi-omics approach was used to study tree responses to bark beetle pioneers under natural conditions. At the site of the attacks, increased levels in jasmonic acid, together with changes in other phytohormones, induced differential gene expression of up to 1,900 genes from multiple defense-related pathways. These transcriptomic alternations were corroborated by proteomic and metabolomic shifts, which ultimately showed increased levels of compounds involved in deterring bark beetle attacks, such as phenolic aglycones, diterpene resin acids, mono- and sesquiterpenes, and lignin. Our study highlights the intricate yet rapid and effective local defense response of Norway spruce against pioneer bark beetle attacks and lays a foundation for in-depth studies of this and other conifers resistance to phloem-feeding insects.

plant biology↗

A Genome-wide CRISPR Screen Reveals ZDHHC8-Dependent Gαq Palmitoylation as a Key Regulator of GPCR Signaling

G protein-coupled receptors (GPCRs) that couple to the Gq signaling pathway control diverse physiological processes, yet the full complement of cellular regulators for this pathway remains unknown. Here, we report the first genome-wide CRISPR knockout screen targeting a Gq-coupled GPCR signaling cascade. Using a Drosophila model of adipokinetic hormone receptor (AkhR) signaling, we identified CG34449 (Zdhhc8), encoding a palmitoyl acyltransferase and its adapter protein CG5447, as a top hit required for robust Gq-mediated GPCR signaling. We show that Zdhhc8 enhances GPCR signaling through palmitoylation of Gq, which promotes its membrane localization and function. Loss of Zdhhc8 markedly reduces palmitoylation of Gaq resulting in attenuation of AkhR/Gq signaling and a reduction in receptor stability. Mechanistically, Zdhhc8 is necessary for palmitoylation of Gq. These findings uncover Zdhhc8-dependent Gq palmitoylation as a pivotal regulatory mechanism in GPCR signal transduction and highlight palmitoyl transferase as potential modulators of GPCR pathways.

cell biology↗

Computational Modeling and Visualization of Ischemic Effects on an Advanced Purkinje Network

The cardiac Purkinje network plays a vital role in the hearts electrical conduction system, ensuring efficient and synchronized ventricular contraction. When impaired--particularly by ischemia--it can trigger life-threatening arrhythmias. In this study, we present an advanced computational model of the Purkinje network that integrates cell-level heterogeneity, spatial organization, and localized ischemic zones with customizable severity gradients. Developed in Python using open-source libraries (NumPy, matplotlib, pandas, seaborn), the model generates rich visualizations of network structure, conduction velocities, ischemia severity, and electrophysiological parameters. Simulations demonstrate how ischemia alters conduction and refractoriness in a spatially dependent manner, providing insights into arrhythmogenic risk. This modeling framework can advance understanding of cardiac conduction under pathological conditions and support therapeutic development.

bioinformatics↗

Isolation of Cyclotide Gene from Solanaceae Family and Its Bioinformatics Analysis

Cyclotides are highly tolerant of sequence variability, aside from the conserved residues forming the cystine knot. Cyclotides have attractive natural movement capabilities including cytotoxicity, harmfulness to HIV, antimicrobials, insecticides, uterine contraction and inhibition of proteases, making them very important in pharmacy and horticulture. The medicinal cyclotides may be able to reimburse the safety of antimicrobials, which is a noticeable problem worldwide today. The present study was focused on the isolation of genes that encoding cyclotide, from plant species. DNA was isolated by CTAB method. The gene was amplified by PCR and after agarose gel electrophoresis confirmation that gene was sequenced followed by bioinformatics analysis. Petunia showed 90% to 97% identity with cds of reference sequence and 27 amino acids were identical in multiple alignment of protein sequence. In addition to providing information for the future investigation of Solanaceae species for cyclotides, this work will aid in understanding the plant families that produce cyclotides.

genetics↗

Sensing of dietary amino acids and regulation of calcium dynamics in adipose tissues through Adipokinetic hormone in Drosophila

Nutrient sensing and the subsequent metabolic responses are fundamental functions of animals, closely linked to diseases such as type 2 diabetes and various obesity-related diseases. Drosophila melanogaster has emerged as an excellent model for investigating metabolism and its associated disorders. In this study, we used live-cell imaging to demonstrate that the fly functional homolog of mammalian glucagon, Adipokinetic hormone (AKH), secreted from AKH hormone-producing cells (APCs) in the corpora cardiaca, stimulates intracellular Ca2+ waves in the larval fat body/adipose tissue to promote lipid metabolism. Further, we show that specific dietary amino acids activate the APCs, leading to increased intracellular Ca2+ and subsequent AKH secretion. Finally, a comparison of Ca2+ dynamics in larval and adult fat bodies revealed different mechanisms of regulation, highlighting the interplay of pulses of AKH secretion, extracellular diffusion of the hormone, and intercellular communication through gap junctions. Our study underscores the suitability of Drosophila as a powerful model for exploring real-time nutrient sensing and inter-organ communication dynamics.

genetics↗

Vaginal lactobacilli produce anti-inflammatory beta-carboline compounds.

The vaginal microbiome is a lactobacilli-dominant, human-adapted community consistently found in people around the world. Presence of lactobacilli-dominant vaginal microbial communities, apart from Lactobacillus iners, is associated with reduced vaginal inflammation and reduced levels of pro-inflammatory cytokines. Loss of lactobacilli-dominance is associated with inflammatory conditions such as bacterial vaginosis. It remains unclear if health-associated lactobacilli actively promote the anti-inflammatory immune homeostasis of the vaginal mucosa and if so, by which mechanisms. We have identified that Lactobacillus crispatus, a key vaginal bacterial species, secretes a family of {beta}-carboline compounds with anti-inflammatory activity. These compounds suppress NF{kappa}B and interferon signaling downstream of multiple pattern recognition receptors in human macrophages and significantly dampen type I interferon receptor activation. Topical vaginal application of an anti-inflammatory {beta}-carboline compound, perlolyrine, was sufficient to significantly reduce vaginal inflammation in a mouse model of genital herpes infection. Together, we identify a key family of compounds by which vaginal lactobacilli mediate host immune homeostasis and could highlight a new therapeutic avenue for vaginal inflammation.

microbiology↗

The origin, deployment, and evolution of a plant-parasitic nematode effectorome

Plant-parasitic nematodes constrain global food security. During parasitism, they secrete effectors into the host plant from two types of pharyngeal gland cells. These effectors elicit profound changes in host biology to suppress immunity and establish a unique feeding organ from which the nematode draws nutrition. Despite the importance of effectors in nematode parasitism, there has been no comprehensive identification and characterisation of the effector repertoire of any plant-parasitic nematode. To address this, we advance techniques for gland cell isolation and transcriptional analysis to define a stringent annotation of putative effectors for the cyst nematode Heterodera schachtii at three key life-stages. We define 659 effector gene loci: 293 "known" high-confidence homologs of plant-parasitic nematode effectors, and 366 "novel" effectors with high gland cell expression. In doing so we define a comprehensive "effectorome" of a plant-parasitic nematode. Using this effector definition, we provide the first systems-level understanding of the origin, deployment and evolution of a plant-parasitic nematode effectorome. The robust identification of the comprehensive effector repertoire of a plant-parasitic nematode will underpin our understanding of nematode pathology, and hence, inform strategies for crop protection.

plant biology↗

"Cortical Delta" In The Medulla Of Sleeping Infant Rats

In early development, active sleep is the predominant sleep state before it is supplanted by quiet sleep. In rats, the developmental increase in quiet sleep is accompanied by the sudden emergence of the cortical delta rhythm (0.5-4 Hz) around postnatal day 12 (P12). We sought to explain the emergence of cortical delta by assessing developmental changes in the activity of the parafacial zone (PZ), a medullary structure thought to regulate quiet sleep in adults. We recorded from PZ in P10 and P12 rats and predicted an age-related increase in neural activity during increasing periods of delta-rich cortical activity. Instead, during quiet sleep we discovered sleep-dependent rhythmic spiking activity--with intervening periods of total silence--phase-locked to a local delta rhythm. Moreover, PZ and cortical delta were coherent at P12, but not at P10. PZ delta was also phase-locked to respiration, suggesting sleep-dependent modulation of PZ activity by respiratory pacemakers in the ventral medulla. Disconnecting the main olfactory bulbs from the cortex did not diminish cortical delta, indicating that the influence of respiration on delta at this age is not mediated indirectly through nasal breathing. Finally, we observed an increase in parvalbumin-expressing terminals in PZ across these ages, supporting a role for GABAergic inhibition in PZs rhythmicity. The discovery of delta-rhythmic neural activity in the medulla--when cortical delta is also emerging--opens a new path to understanding the brainstems role in regulating sleep and synchronizing rhythmic activity throughout the brain.

neuroscience↗

Variant annotation across homologous proteins (Paralogue Annotation) identifies disease-causing missense variants with high precision, and is widely applicable across protein families

BackgroundDistinguishing pathogenic variants from those that are rare but benign remains a key challenge in clinical genetics, especially for variants not previously observed and characterised in humans. In vitro and in vivo functional characterisation are typically resource intensive, and model systems may not accurately predict influence on human disease. Many in silico tools have been developed to predict which variants are disease-causing, but typically lack precision. Here we demonstrate the applicability of a framework, called Paralogue Annotation, that draws on information from previously-characterised variants in homologous proteins to predict whether variants in a gene of interest are likely disease causing. MethodsWe assessed the performance of Paralogue Annotation through three orthogonal approaches: (1) comparison to established in silico variant prediction tools using 47,016 missense variants from ClinVar across 3,524 genes representing a broad range of diverse protein classes, by calculating precision and sensitivity; (2) evaluation against large-scale functional assays of variant effect; and (3) comparing odd ratios calculated from case-control association tests for inherited cardiac arrhythmia syndromes, and neurodevelopmental disorders with epilepsy, stratifying variants by Paralogue Annotation. ResultsParalogue Annotation correctly annotates 4,328 ClinVar pathogenic variants, with 245 false positives, yielding a precision of 0.95. This increases to 0.99 with more stringent annotation parameters (requiring greater conservation of amino acids between annotated orthologues) at the expense of sensitivity. Compared to established tools, Paralogue Annotation has higher precision for the identification of pathogenic variants, albeit with lower sensitivity across diverse test sets. Extending the technique by transferring annotations between homologous protein domains, rather than full-length protein paralogues, increases sensitivity. Rare variants predicted pathogenic by Paralogue Annotation were more strongly disease-associated (increased odds ratio) than unstratified rare variants for six out of eight genes tested with case-control cohort approaches. ConclusionsWe demonstrate that Paralogue Annotation has high precision for predicting pathogenic missense variants, providing a useful line of evidence for clinical variant interpretation that is complementary to other approaches in use. As the number of characterised variants increases in reference datasets such as ClinVar, Paralogue Annotation will further increase in sensitivity and applicability.

bioinformatics↗