Biology subjects

Awad, S.

Publications and source records attributed to Awad, S..

8 recordsLinked to original sources

bioRxiv2026

Serial vaccination expands and refines human CD4+ T cell memory

CD4+ T cells coordinate protective immunity against pathogens. However, a major unresolved question is how human CD4+ T cell memory is established and evolves following primary and repeated vaccination. Using COVID-19 mRNA vaccination as a model, we tracked 50 distinct antigen-specific populations directly ex vivo with peptide-MHC class II tetramers in eight SARS-CoV-2-naive individuals from pre-vaccine baseline through memory time points after three mRNA doses. Our findings identify the primary vaccine series as the main driver of memory pool size. It leverages pre-existing memory while preferentially recruiting high-avidity T cells, establishing an immunodominance hierarchy dominated by a small subset of precursors. Booster vaccination refines both the magnitude and quality of T cell memory. It increases select populations and enhances differentiation of subdominant CD4+ T cells. Populations that did not become more abundant after boosting retained their polyfunctional potential. Beyond establishing memory to the ancestral spike, vaccinations broadened responses by recruiting cross-reactive T cells recognizing viral variants. Collectively, these findings reveal how human CD4+ T cell memory evolves through sequential immunizations to generate a functionally diverse and broadly responsive memory repertoire against future viral challenges.

immunology↗

bioRxiv2026

RUNX1 aberrations in blast-phase CML induce the RBP SPATS2L which promotes growth, survival and stress granule assembly

The RUNX1 transcription factor is a critical regulator of hematopoiesis and frequently mutated in myeloid malignancies. In the myeloproliferative neoplasm, chronic myeloid leukemia (CML), secondary somatic RUNX1 mutations and RUNX1::MECOM/EVI1, are associated with tyrosine kinase inhibitor (TKI) resistance and progression to the blast-phase (BP-CML). Research has predominantly focussed on transcriptional dysregulation mediated by RUNX1 mutations in myeloid malignancies, whilst post-transcriptional dysregulation remains comparatively unexplored. To address this, we used orthogonal organic phase separation (OOPS), to characterise the RNA-binding proteome of RUNX1 deficient BP-CML cells. RUNX1 depleted BP-CML cells exhibited significant alterations to RBP abundance involved in stress response pathways and translation/ribosome-biogenesis (RiBi). Furthermore, RUNX1 depletion or expression of RUNX1::EVI1 in BP-CML cells induced expression and RNA binding activity of SPATS2L, a component of stress granules (SG); membraneless cytoplasmic condensates protecting mRNAs from degradation, promoting survival under stress. Whilst RUNX1 depletion increased SG-assembly, SPATS2L depletion reduced SG-assembly in BP-CML cells and inhibited the growth and survival of multiple BP-CML cell lines. The translation inhibitor homoharringtonine (HHT), used historically in TKI-resistant CML, ablated SG-assembly in BP-CML cells with RUNX1 depletion, and, primary BP-CML cells with LOF/hypomorphic RUNX1 mutations (characterised by defective DNA-binding/CBF{beta}-interaction) were preferentially sensitised to HHT. Finally, suppressing SPATS2L expression induced by RUNX1 depletion, increased the HHT-sensitivity of RUNX1 depleted BP-CML cells, suggesting SPATS2L contributes to therapeutic resistance in CML with RUNX1 mutations. This study suggests that SPATS2L and SG induction could be critical to RUNX1-mutant leukemias, and, provides preliminary evidence for a mutationally-targeted approach in CML with RUNX1 aberrations.

cancer biology↗

bioRxiv2026

Muller glia mediated regeneration restores neuronal diversity and retinal circuit organization in the adult zebrafish

The ability to regenerate neurons with appropriate identities and circuit connectivity is a fundamental challenge in regenerative biology. Unlike mammals, adult zebrafish robustly regenerate retinal neurons after injury through the reprogramming of Muller glia. However, the extent to which regenerated neurons faithfully reconstruct molecular identity, cellular diversity, and circuit organization remains unclear. Here, we combined inducible lineage tracing, single-cell RNA sequencing and high-resolution morphological analysis to define the molecular identities and structural organization of regenerated neurons following photoreceptor-selective light lesion or NMDA-induced inner retinal injury. Both injury paradigms regenerated all major retinal cell classes, although the relative abundance of regenerated cell types reflected the pattern of neuronal loss. Across major neuronal classes and subtypes, regenerated neurons largely reestablished endogenous molecular identities, with the residual transcriptional differences primarily reflecting ongoing maturation. Regenerated amacrine and bipolar neurons recovered subtype diversity, characteristic dendritic morphologies, and laminar organization. Regenerated retinal ganglion cells likewise restored broad molecular diversity and appropriate retinotectal projections, while a subset underwent microglia-mediated refinement. Together, these findings demonstrate that Muller glia-mediated regeneration largely reconstructs neuronal identity, cellular diversity and key features of retinal circuit organization, providing insights into understanding how complex neuronal tissues are rebuilt after injury.

neuroscience↗

bioRxiv2025

AAV-mediated regeneration of neurons in injury-induced and inherited retinal degeneration

Loss of retinal neurons is a leading cause of irreversible vision impairment, yet adult mammalian retinas lack the ability to regenerate these cells. In this study, we developed an adeno-associated virus (AAV)-based strategy to reprogram endogenous Muller glia into retinal neurons. Using stringent genetic lineage tracing, immunohistochemistry, electrophysiology and single-cell multiomic profiling, we show that AAV delivery of a stabilized, phospho-insensitive Neurogenin2 variant (Neurog2-9SA) efficiently converts Muller glia into multiple types of retinal neurons, including bipolar, starburst amacrine, and a small population of photoreceptor-like cells, in both injury-induced and inherited retinal degeneration models. The generated neurons exhibit electrical properties of retinal neurons, light-evoked responses and integrate into existing retinal circuitry. Single-cell multiomics analysis reveal that Neurog2-9SA induces reprogramming by remodeling chromatin accessibility, activating neurogenic transcriptional networks, and represses glial identity programs. Inhibiting Notch signaling markedly enhances reprogramming efficiency. Together, these findings establish Neurog2-9SA as a potent and clinically relevant factor for AAV-mediated reprogramming and provide a foundation for approaches to regenerate neuronal cells and restore function in retinal diseases.

neuroscience↗

bioRxiv2024

Divergent effects of cytomegalovirus and rheumatoid arthritis on senescent CD4+ T cells

CD4+ T cell senescence has been linked to repeated antigen stimulation. However, how different types of chronic antigenic exposures impact T cell differentiation and function remains incompletely understood. Using rheumatoid arthritis (RA) and cytomegalovirus (CMV) as models for persistent stimulation in autoimmunity and chronic viral infection, we performed high-dimensional mass cytometry analyses to examine their effects on CD4+ T cell differentiation. We found that CMV seropositive adults have a significant population of highly differentiated CD27-CD28-CD4+ T cells that exhibited common features of senescence, including CD57 and cytotoxic granule expression. In contrast, CD27-CD28-CD4+ T cells were rare in RA patients and Epstein-Barr virus (EBV) or Herpes simplex virus (HSV) seropositive individuals. The few that were present exhibited a predominantly non-cytotoxic profile, with higher expression of TCF1, CD127, and Ki67. Among CMV seropositive individuals, RA was associated with reduced degranulation of cytotoxic granules and lower cytokine production by senescent CD4+ T cells. We did not find an association with age, sex, clinical characteristics, or medication usage by univariate linear regression analyses. However, in vitro tofacitinib treatment reduced T cell functional activity, suggesting contributions from both the disease and RA treatment. These data uncovered distinct influences from CMV and RA, and their combined impact on senescent CD4+ T cell differentiation and function.

immunology↗

bioRxiv2024

Viral-mediated Oct4 overexpression and inhibition of Notch signaling synergistically induce neurogenic competence in mammalian Muller glia.

Retinal Muller glia in cold-blooded vertebrates can reprogram into neurogenic progenitors to replace neurons lost to injury, but mammals lack this ability. While recent studies have shown that transgenic overexpression of neurogenic bHLH factors and glial-specific disruption of NFI family transcription factors and Notch signaling induce neurogenic competence in mammalian Muller glia, induction of neurogenesis in wild-type glia has thus far proven elusive. Here, we report that viral-mediated overexpression of the pluripotency factor Oct4 (Pou5f1) induces transdifferentiation of mouse Muller glia into bipolar neurons, and synergistically stimulates glial-derived neurogenesis in parallel with Notch loss of function. Single cell multiomic analysis shows that Oct4 overexpression leads to widespread changes in gene expression and chromatin accessibility, inducing activity of both the neurogenic transcription factor Rfx4 and the Yamanaka factors Sox2 and Klf4. This study demonstrates that viral-mediated overexpression of Oct4 induces neurogenic competence in retinal Muller glia, identifying mechanisms that could be used in cell-based therapies for treating retinal dystrophies.

neuroscience↗

bioRxiv2023

Genome-wide analysis of promoter contacts identifies novel regulators of late-stage adipogenesis

Adipogenesis is a multi-stage process essential for healthy fat storage and metabolic regulation. While early regulatory mechanisms are well characterized, the control of late-stage adipocyte differentiation remains poorly understood. Integrating CAGE-seq, promoter capture Hi-C, and a high-throughput siRNA screen of druggable genes, we report here that chromatin architecture rewiring promotes gene regulation changes essential for terminal adipogenesis. We identified nine clusters of dynamic promoter-anchored chromosomal interactions, many involving distal enhancers. Functional screening of genes engaged in these interactions revealed 19 novel regulators of late adipogenesis, including proteins with peptidase and ubiquitin ligase activity. Human genetic variant-to-gene mapping, coupled with cross-species chromatin interaction and synteny analyses, highlighted new gene-trait associations relevant to lipid traits (FXYD5, LAP3, SGPP1) and type 2 diabetes (FBXO17, FN3KRP, ZFAND6, TTC3). Our findings define the 3D gene regulatory landscape of late adipogenesis. The molecular links uncovered here provide mechanistic insight into metabolic disease risk and potential interventions.

genetics↗

bioRxiv2017

Evaluating Metagenome Assembly on a Simple Defined Community with Many Strain Variants

We evaluate the performance of three metagenome assemblers, IDBA, MetaSPAdes, and MEGAHIT, on short-read sequencing of a defined \"mock\" community containing 64 genomes (Shakya et al. (2013)). We update the reference metagenome for this mock community and detect several additional genomes in the read data set. We show that strain confusion results in significant loss in assembly of reference genomes that are otherwise completely present in the read data set. In agreement with previous studies, we find that MEGAHIT performs best computationally; we also show that MEGAHIT tends to recover larger portions of the strain variants than the other assemblers.

bioinformatics↗