Biology subjects

Shree, S.

Publications and source records attributed to Shree, S..

10 recordsLinked to original sources

bioRxiv2026

Neonatal T cell maternal microchimerism is common and associates with early pregnancy SARS2 vaccination

The fetus acquires a graft of maternal cells in utero, known as maternal microchimerism (MMc), which may be enriched for T cells. However, little is known regarding the determinants of T cell trafficking into the fetus or regarding the prevalence of T cell MMc in healthy neonates. Using a closely followed cohort, we determined the association between maternal vaccination and acute infection and the detection of T cell MMc in cord blood, as measured using polymorphism-specific qPCR. We identified an association between early pregnancy SARS2 mRNA vaccination, a potent T cell stimulus, and increased detection of T cell MMc.

immunology↗

bioRxiv2026

A sensitive and specific microchimerism detection method using custom designed genotyping and companion qPCR assays

Since first reports of the presence of a small number of cells from one individual persisting in another, researchers have developed a number of methods to detect microchimerism. Molecular based detection platforms range from gel electrophoresis to quantitative PCR to Next-Gen Sequencing. Central to all approaches is the need for informative polymorphic loci to effectively distinguish microchimeric cells from the host. However, we observed in our studies of maternal microchimerism that currently published approaches can limit the number of samples available for assessment due to the inability to identify an informative marker of the donor in the recipient. This was most striking in our studies of populations of African descent in which the use of HLA-based markers, a common approach, prevented the testing of 30-50% of participants. Other potential options were cost prohibitive or lacked sufficient sensitivity. To address this challenge, we developed a novel, simple, and low-cost approach that leverages global population frequency databases for polymorphism selection, reduces cost by amplifying multiple targets in one multiplex assay to complete genotyping, and has high sensitivity and specificity using rigorously designed qPCR assays to quantify microchimerism. Herein, we outline our design process, detailed assay protocol, and assessment of maternal microchimerism using this approach in two different populations of infants from the United States and South Africa.

molecular biology↗

bioRxiv2026

Direct engagement of mycobacterial RadA by c-di-AMP at a non-catalytic site suppresses its DNA-stimulated ATPase activity

Cyclic di-adenosine monophosphate couples DNA damage sensing to genome maintenance in mycobacteria, but how it regulates homologous recombination machinery beyond RecA remains unclear. Using Mycobacterium smegmatis RadA (MsRadA) as a model, we show by isothermal titration calorimetry that c-di-AMP binds to MsRadA, providing biophysical evidence for an interaction previously inferred only from genetic studies. C-di-AMP binding is independent of magnesium ions and is not affected by prior occupancy of the ATP-binding site by the ATP analogue {beta},{gamma}-methylene adenosine 5'-triphosphate, indicating that c-di-AMP binds at a site distinct from the canonical ATPase pocket. Functionally, c-di-AMP reduces MsRadA DNA-dependent ATPase activity. Structural modelling and molecular dynamics (MD) simulations show that c-di-AMP is stably bound in the central channel of the RadA hexamer, non-covalently engaged with the DNA-binding residues Arg197, Lys226, and Arg234, which are contributed by two adjacent subunit interfaces of the hexamer. In the modelled RadA-ATP-c-di-AMP-DNA quaternary complex, c-di-AMP was predicted at a peripheral site near, but distinct from, the ATP-binding pocket. Together, these findings identify RadA as a direct physical target of c-di-AMP within the homologous recombination pathway and provide a framework for testing how c-di-AMP regulates RadA during DNA damage.

biophysics↗

bioRxiv2026

Small-Molecule Inhibitors Allosterically Control Bruton's Tyrosine Kinase Assembly on the Membrane

Bruton's tyrosine kinase (Btk) is the target of small-molecule, active site inhibitors for treatment of lymphomas and leukemias. Beyond its catalytic activity, Btk participates in signaling assemblies through scaffolding functions that can contribute to drug resistance. Here, we show that Btk forms micron-scale assemblies on membranes and that this behavior is allosterically controlled by ligand binding at the active site. Using supported lipid bilayers and total internal reflection fluorescence microscopy, we find that nucleotides and inhibitors including ibrutinib, acalabrutinib, and dasatinib promote assembly, whereas fenebrutinib and pirtobrutinib prevent assembly. Membrane organization is independent of catalysis but correlates with ligand-dependent changes in Btk conformational flexibility measured by hydrogen-deuterium exchange. These results reveal a previously unrecognized connection between active-site ligand binding and mesoscale membrane organization, demonstrating that kinase inhibitors can control Btk behavior beyond suppression of catalytic activity.

biophysics↗

bioRxiv2026

Human tissue-resident CD8 T cells contribute to trophoblast homeostasis in health and during acute inflammation

Previous studies have highlighted that some T cell subsets in tissues can provide signals to support tissue cell homeostasis and differentiation. If and how T cell-tissue cell signaling is altered in healthy compared to inflamed tissues is poorly understood. Here, we address if communication between human T cells and tissue cells changes from steady state to an acutely inflamed state in the human placenta. We used single cell analysis strategies to examine invasive cytotrophoblasts (iCTBs) and immune cells isolated from third trimester healthy and acutely inflamed human placentas. We performed cell communication analysis to predict cell-cell communication networks, and found evidence that iCTBs provided signals to support the recruitment of T cells, as well as the formation of tissue-resident memory CD8 T cells (Trm). In exchange, Trm provide signals to support iCTB homeostasis. During acute inflammation, iCTBs and macrophages underwent profound transcriptional changes, while most T cell subsets only underwent limited transcriptional changes. This was not due to T cell exhaustion or tolerance, as T cells were functionally intact. Cell communication analysis and validation at the protein level provide evidence that T cells can maintain their homeostatic support to iCTBs during acute inflammation.

immunology↗

bioRxiv2026

Hyperglycemia transcriptionally regulates the paranodal protein (Caspr1) in retinal neurons and modulates neurite extension

Hyperglycemia is a hallmark of diabetes, affecting neuronal structure and function by altering molecular signalling pathways. Here, we explore the role of hyperglycemia in regulating Caspr1 expression and its downstream effects on neurite outgrowth. Caspr1, a critical protein implicated in neurodegenerative diseases, was found to be significantly downregulated in N2a and 661W cell lines cultured under hyperglycemic conditions (25mM glucose) and, as a result, promoted neurite outgrowth. Knockout of Caspr1 using CRISPR-Cas9 further confirmed its inhibitory role on neurite outgrowth, as Caspr1-deficient cells exhibited enhanced neurite elongation. Caspr1 downregulation was mediated by decreased expression of C/EBP, a key transcription factor with a binding site on the Caspr1 promoter. Overexpression of C/EBP restored Caspr1 promoter activity and mRNA levels, establishing C/EBP as a critical regulator. Additionally, hyperglycemia was observed to inhibit Akt phosphorylation, which further contributed to Caspr1 downregulation. Adding insulin to the culture medium under hyperglycemic conditions shows inhibition of Akt phosphorylation and downregulation of Caspr1, resulting in a shorter length of neurites in retinal neurons. In vivo, studies in diabetic mouse models and diabetic patient samples demonstrated reduced expression of Caspr1 in retinal tissues. These results suggest that hyperglycemia regulates Caspr1 expression through Akt and C/EBP pathways, promoting neurite outgrowth in retinal neurons. In contrast, adding insulin to the medium under hyperglycemia downregulates the Caspr1 expression and reduces neurite length in retinal neurons. Targeting this pathway may offer new therapeutic approaches to mitigate neurodegeneration in diabetic retinopathy.

cell biology↗

bioRxiv2025

Neurons exploit stochastic growth to rapidly and economically build dense radially oriented dendritic arbors

Dendrites grow by stochastic branching, elongation, and retraction. A key question is whether such a mechanism is sufficient to form highly branched dendritic morphologies. Alternatively, are signals from other cells or is the topological hierarchy of the growing network necessary for dendrite geometry? To answer these questions, we developed a mean-field model in which branch dynamics is isotropic and homogenous (i.e., no extrinsic instruction) and depends only on the average lengths and densities of branches. Branching is modeled as density-dependent nucleation so there are no tree structures and no network topology. Despite its simplicity, the model predicted several key morphological properties of class IV Drosophila sensory dendrites, including the exponential distribution of branch lengths, the parabolic scaling between dendrite number and length densities, the tight spacing of the dendritic meshwork (which required minimal total branch length), and the radial orientation of branches. Stochastic growth also accelerated the overall expansion rate of the arbor. Therefore, stochastic dynamics is an economical and rapid space-filling mechanism for building dendritic arbors without external guidance or hierarchical branching mechanisms. Our model provides a general theoretical framework for understanding how macroscopic branching patterns emerge from microscopic dynamics.

developmental biology↗

bioRxiv2021

Dynamic Instability of Dendrite Tips Generates the Highly Branched Morphologies of Sensory Neurons

The highly ramified arbors of neuronal dendrites provide the substrate for the high connectivity and computational power of the brain. Altered dendritic morphology is associated with neuronal diseases. Many molecules have been shown to play crucial roles in shaping and maintaining dendrite morphology. Yet, the underlying principles by which molecular interactions generate branched morphologies are not understood. To elucidate these principles, we visualized the growth of dendrites throughout larval development of Drosophila sensory neurons and discovered that the tips of dendrites undergo dynamic instability, transitioning rapidly and stochastically between growing, shrinking, and paused states. By incorporating these measured dynamics into a novel, agent-based computational model, we showed that the complex and highly variable dendritic morphologies of these cells are a consequence of the stochastic dynamics of their dendrite tips. These principles may generalize to branching of other neuronal cell-types, as well as to branching at the subcellular and tissue levels.

biophysics↗

bioRxiv2019

Sensitive detection of highly fragmented cytomegalovirus nucleic acid in human cfDNA

Congenital human cytomegalovirus (CMV) infections are the leading cause of newborn hearing and central nervous system impairments worldwide. Currently, routine prenatal screening for congenital CMV is not performed in the United States and confirmation in suspected perinatal cases requires invasive sampling by amniocentesis. We hypothesized that detection of CMV from maternal cell-free DNA (cfDNA) plasma could provide a non-invasive indicator of congenital CMV infection. We analyzed sequence data from 2,208 individuals undergoing routine non-invasive prenatal aneuploidy screening at the University of Washington. CMV reads were identified in 117 (5.3%) samples. Positive samples were stratified based on CMV reads per million sample reads (RPM), resulting in ten samples being classified as strong positive (RPM > 0.3) and 107 as intermediate positive (0.01<RPM<0.3). Subsequent qPCR testing identified CMV in 9/10 strong positive samples and 2/32 intermediate positive samples. Median cfDNA insert size derived from CMV was significantly shorter than cfDNA derived from human chromosomes (103 vs 172 bp, p<0.0001), corresponding to the 3rd percentile of human cfDNA insert size. In addition, CMV cfDNA fragment lengths were distributed over a wider range than human cfDNA reads. These studies reveal the highly fragmented nature of CMV cfDNA and offer precise measurements of its length: these features likely explain discrepancies in serum CMV viral loads measurements determined by different qPCR assays, despite widespread efforts to standardize results. More work is required to determine how detection of CMV from maternal cfDNA can be best used as tool for congenital CMV screening or diagnosis.

microbiology↗

bioRxiv2019

Beyond the suprachiasmatic nucleus: circadian rhythmicity shapes astrocyte morphology and neuronal function in hippocampal area CA1

Most animal species operate according to a 24-hour period set by the suprachiasmatic nucleus (SCN) of the hypothalamus. The rhythmic activity of the SCN is known to modulate hippocampal-dependent memory processes, but the molecular and cellular mechanisms that account for this effect remain largely unknown. Here, we show that there are cell-type specific structural and functional changes that occur with circadian rhythmicity in neurons and astrocytes in hippocampal area CA1. Pyramidal neurons change the surface expression of NMDA receptors, whereas astrocytes change their proximity to synapses. Together, these phenomena alter glutamate clearance, receptor activation and integration of temporally clustered excitatory synaptic inputs, ultimately shaping hippocampal-dependent learning in vivo. We identify corticosterone as a key contributor to changes in synaptic strength. These findings identify important mechanisms through which neurons and astrocytes modify the molecular composition and structure of the synaptic environment, contribute to the local storage of information in the hippocampus and alter the temporal dynamics of cognitive processing.

neuroscience↗