Biology subjects

Tong, L.

Publications and source records attributed to Tong, L..

At least 19 recordsLinked to original sources

bioRxiv2026

Pseudomonas aeruginosa R-bodies deliver an ADP deaminase toxin

Diverse bacteria produce proteinaceous polymeric coils called R-bodies (refractile bodies), which extend into ~10 m-long needles in response to environmental triggers. It has been proposed that R-bodies deliver toxins intracellularly after such bacteria are ingested by eukaryotes, but intra-host extension of R-bodies has not been directly observed and these toxins remain unidentified. We use Pseudomonas aeruginosa PA14 and Dictyostelium discoideum to show that R-bodies extend within amoebae. We also identify a novel toxin, DeaR (deaminase associated with the R-body), which contributes to predation resistance and forms a near-megadalton dodecamer with ADP deaminase activity that alters purine metabolism. Our findings elucidate the molecular basis of R-body-mediated toxicity during PA14 infection and provide a paradigm for understanding the potential of cytotoxic effectors delivered by R-bodies (CEDRs) in interkingdom interactions.

microbiology↗

bioRxiv2026

A Computational Pipeline for Retinal Capillary Blood Flow Measurement using Adaptive Optics Line Confocal Ophthalmoscopy

Background and Objective: High-speed and high-resolution retinal imaging using adaptive optics line confocal ophthalmoscopy (AOLCO) resolves individual red blood cells (erythrocytes) flowing through the smallest retinal capillaries, but quantifying their velocity first requires removing linear and nonlinear distortions caused by eye motion that may result in a displacement of tens to hundreds of pixels between frames. We present a computational pipeline that stabilizes AOLCO capillary video and measures erythrocyte velocity in true physical units, delivered as an interactive 3D Slicer extension and a batch command-line tool sharing one analysis core. Methods: Our video analysis pipeline has four stages: preprocessing, registration, postprocessing, and velocity measurement. During preprocessing, illumination is corrected, and closed-eye frames are identified. During registration, each frame is aligned to a reference frame using affine and, when needed, B-Spline registration, initialized by our robust and fast (GPU-based) translation-registration algorithm to accommodate large eye movements (e.g., microsaccades). During postprocessing, adjacent registered frames are differenced to suppress stationary structures, and a 2D projection of the frame-difference video reveals the capillary network. During velocity measurement, an operator manually identifies capillary segments on the 2D projection image. Spatiotemporal images are then generated along these segments from the frame-difference video, and blood-flow velocity curves are obtained from these images using the Radon transform. Results: We validate the velocimetry of the pipeline against synthetic ground-truth flow data, and the experiments show that the velocimetry recovers the speed within an error of 4% The GPU-based translation registration algorithm aligns the large eye motions (~90 um per frame on average) and runs about 3.6 x (100 Hz) to 38 x (400 Hz) faster than the baseline method. Across the full cohort (2344 videos, 57 subjects, 30 Hz to 400 Hz), our registration approach improves structural similarity (SSIM) of the video frames on every acquisition from 0.73 to 0.86. The recovered blood flow velocities are physiologically plausible, of the same order as published AO measurements. Conclusions: Our computational pipeline enables AOLCO velocimetry on the capillary-level.

bioengineering↗

bioRxiv2026

Single-cell splice isoform usage reveals distinct axes of cellular identity and senescence

Alternative splicing greatly expands the diversity of gene products encoded by the human genome. Single-cell transcriptomic atlases have characterized human cell types through gene-level expression, but short-read sequencing has limited the ability to resolve full-length isoforms and their functional consequences. Here, we present a cross-tissue single-cell long-read isoform atlas spanning 26 human tissues. We identify hundreds of thousands of novel isoforms along with their cell-type-specific usage, and discover that over one-third of expressed isoforms are absent from existing reference databases. We further demonstrate that isoform usage is a structured, measurable axis of cellular identity that is distinct from gene expression. Applying this framework to cellular senescence, we resolve p16INK4a and p14ARF transcripts from the CDKN2A locus in individual cells and uncover cell-type-dependent isoform remodeling associated with the p16INK4a senescence program. This isoform-resolved single-cell atlas offers a versatile framework to dissect the cellular logic of isoform regulation in senescence and beyond.

genomics↗

bioRxiv2026

Programming Brain Cell-Type-Selective Delivery In Vivo with Transporter-Guided Therapeutics

Many diseases arise from dysfunction of defined cell populations, yet most therapeutics distribute broadly, limiting efficacy and causing toxicity. We developed ExACT, a platform for cell-type-selective intracellular delivery that exploits membrane transporters. In vivo screening of combinatorial fluorescent small-molecule libraries in mouse brain identified chemistries whose uptake is dictated by endogenous transporter expression, yielding compounds with preferential entry into neurons, astrocytes, pericytes and endothelial cells. One series showed strong selectivity for brain and retinal endothelium, where Slco1a4 mediated uptake. This selectivity principle extended to the human orthologue SLCO1A2, highly expressed in brain endothelium and oligodendrocytes, where it mediated selective uptake in a humanized mouse model and human iPSC-derived oligodendrocytes. Ectopic expression of SLCO1A2 in neurons via gene therapy created a synthetic entry port, conferring ExACT conjugate uptake on otherwise inaccessible cells. Bifunctional compounds linking transporter-targeting motifs to antisense oligonucleotides or small-molecule drugs retained pharmacological activity while conferring transporter-dependent cell-type selectivity, illustrating how transporter diversity can be harnessed for precision pharmacotherapy.

bioengineering↗

bioRxiv2026

Magnetically activatable insect viral vectors promote anticancer immunity through spatially confined gene disruption

Precise control of genome editing activity in vivo remains a major barrier to the clinical translation of CRISPR-based therapeutics, as current approaches cannot reliably restrict editing activity to target tissues 1,2,3. Here we show that a magnetically gated baculoviral CRISPR platform enables spatial control of genome editing activity 4. By coupling magnetic activation of viral transduction at target sites with complement-mediated inactivation of circulating vectors, this system functionally confines genome editing activity to defined spatial contexts, effectively decoupling editing activity from biodistribution. We further show that spatially confined genome editing can be coupled with endogenous antiviral sensing to drive coordinated immune modulation within tumours. In a syngeneic murine tumour model, local administration combined with magnetic activation restricts genome editing to tumours without detectable editing in major organs, remodels dendritic cell and CD8{square} T cell states, suppresses tumour growth, and extends survival. Together, these findings establish a generalizable strategy for spatial control of genome editing and localized immunomodulation in cancer.

bioengineering↗

bioRxiv2025

Tissue-to-Bytes: A Catalytic Digital Twin Platform for Consortium-Scale Integration of Single-Cell Omics Data in BRAIN Initiative Cell Atlas Network

Monolithic Electronic Health Record (EHR) architectures remain fundamentally misaligned with high-resolution, specialty-specific clinical workflows, contributing to significant physician burnout and challangies in secondary use of data. This study introduces the Enterprise Health Twin (EHT) framework, a paradigm grounded in the Digital Twin concept, with the goal to transform static clinical ledgers into dynamic decision-support systems. The EHT establishes continuous bidirectional feedback (feed-forward ingestion and feed-backward decision support) between the physical entities of clinical workflows and the digital counterparts of interfaces reflecting the evolving states of the workflow, for each individual patient in the EHT. We conceptualized, prototyped, and operationalized the EHT framework for epilepsy care for seven years, in production, at the point of care. Deployed at the Epilepsy Center of the University of Texas Health Science Center at Houston, a [~]400% increase productivity has been achieved in specialized surgical evaluation throughput without additional staffing. The EHT achieved de novo capture of ontology-anchored structured data at the point of care, mitigating cognitive load while growing the volume of high-resolution, annotated longitudinal datasets for clinical research, and driving an increase of care velocity in a learning healthcare system.

neuroscience↗

bioRxiv2025

Phase separation drives SNARE complexes formation

BackgroundNeuronal exocytosis is mainly driven by the assembly of the soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) complexes. However, little is known about the organization principle of the SNARE complex. MethodsProtein condensates formed by SNARE complex were imaged by confocal microscope. Fluorescence recovery after photobleaching (FRAP) assay together with fusion and division assays at the cellular level and in vitro studies with purified proteins were performed to characterize the dynamic properties of protein condensates. The effect of SNARE complex phase separation on the recruitment of synaptic vesicles was detected by immunofluorescence. ResultsWe discover that phase separation drives SNARE complex formation and aggregation; in addition, nonintrinsically disordered regions (non-IDRs) of the syntaxin1 protein is necessary for the formation of these biological condensates. Functionally, phase separation of the SNARE complex can be regulated by the major cofactors of the fusion machinery and has the ability to recruit synaptic vesicles in neurons. ConclusionsOur study here establishes that phase separation is a promising way to mediate the formation and aggregation of the SNARE complex, and further identified that the non-IDRs of syntaxin1 is necessary for the phase separation of the SNARE complex. Our work answers an vital scientific question: does the SNARE complex function as multiple copies that are dispersed or clustered together to ensure sustained neurotransmitter release. In sum, phase separation provides an ideal working model for SNARE complex-mediated membrane fusion and neurotransmitter release.

neuroscience↗

bioRxiv2025

SUMO2 Promotes Histone Pre-mRNA Processing by Stabilizing Histone Locus Body Interactions and Facilitating U7 snRNP Assembly

Histone mRNAs are the only non-polyadenylated mRNAs in eukaryotic cells and require specialized processing in the histone locus body (HLB), a nuclear body where essential processing factors, including the U7 snRNP, are concentrated. Recent studies have revealed that misregulation of histone pre-mRNA processing can lead to polyadenylation of histone mRNAs and disruption of histone protein homeostasis. Despite links to human disease, the factors contributing to polyadenylation of histone mRNAs and the mechanisms underlying HLB assembly and U7 snRNP biogenesis remain unclear. Here, we report novel functions of the small ubiquitin-related modifier 2 (SUMO2) in promoting histone pre-mRNA processing. Using a SUMO2 knockout osteosarcoma cell line, we identified a defect in 3 end cleavage and a global increase in histone mRNA polyadenylation. Subsequent analysis of HLBs revealed increased dynamics and reduced levels of the U7 snRNP complex. By over-expressing U7 snRNP-specific components, Lsm11 and U7 snRNA, we rescued U7 snRNP levels and processing defects in SUMO2 knockout cells. Through analysis of Lsm11, we identified a SUMO-interacting motif in its N-terminus required for efficient formation of U7 snRNP. Collectively, we demonstrate that SUMO2 promotes histone pre-mRNA 3 end processing by stabilizing HLB interactions and facilitating U7 snRNP assembly.

molecular biology↗

bioRxiv2024

Epigenetic and 3D genome reprogramming during the aging of human hippocampus

Age-related cognitive decline is associated with altered physiology of the hippocampus. While changes in gene expression have been observed in aging brain, the regulatory mechanisms underlying these changes remain underexplored. We generated single-nucleus gene expression, chromatin accessibility, DNA methylation, and 3D genome data from 40 human hippocampal tissues spanning adult lifespan. We observed a striking loss of astrocytes, OPC, and endothelial cells during aging, including astrocytes that play a role in regulating synapses. Microglia undergo a dramatic switch from a homeostatic state to a primed inflammatory state through DNA methylome and 3D genome reprogramming. Aged cells experience erosion of their 3D genome architecture. Our study identifies age-associated changes in cell types/states and gene regulatory features that provide insight into cognitive decline during human aging.

genomics↗

bioRxiv2024

mTORC2-mediated cell-cell interaction promote BMP4-induced WNT activation and mesoderm differentiation

The mechanistic target of rapamycin complex 2 (mTORC2) is essential for embryonic development but its underlying molecular mechanisms remain unclear. Here we show that disruption of mTORC2 in human embryonic stem cells (hESCs) considerably alters the Rho/Rac signaling dynamics and reduces cell adhesion. Despite this, mTORC2-deficient hESCs maintain self-renewal and expression of pluripotent markers when cultured in mouse-embryonic fibroblast conditioned medium supplemented with bFGF (MEF-CM). However, these hESCs exhibit significantly impaired mesoderm and endoderm differentiation in response to BMP4 and Activin, respectively, due to reduced WNT activation mediated by cell-cell interactions. Direct activation of the WNT pathway by a GSK3 inhibitor restores mesendoderm differentiation in mTORC2-deficient hESCs. Our study uncovers a novel mechanism by which mTORC2 regulates cell fate determination and highlights a critical link between the intercellular adhesion and the activation of canonical WNT genes.

cell biology↗

bioRxiv2024

Genome sequencing of 'Fuji' apple clonal varieties reveals evolutionary history and genetic mechanism of the spur-type morphology

Somatic variations arising during asexual reproduction can lead to the emergence of bud sports exhibiting advantageous traits, forming the basis for bud sport breeding in perennial plants. Here, we report a near-complete, fully phased genome assembly of Fuji apple, enabling comprehensive identification of somatic variants across 74 clonally propagated Fuji varieties. Phylogenetic analysis of Fuji sport varieties indicates that the emergence of spur-type and early maturation traits results from multiple independent formation events. A number of putative functional somatic variants are identified, including one spur-type specific deletion located in the promoter of the TCP transcription factor gene MdTCP11. DNA methylation level of the deletion-associated miniature inverted-repeat transposable element is lower in the spur-type varieties compared to the standard-type varieties, while the expression of MdTCP11 is significantly higher. Overexpression of MdTCP11 in apple decreases the plant height, highlighting its important role in the development of spur-type apple varieties. This study sheds light on the 80-year cloning history of Fuji and provides valuable candidate causative mutations for apple breeding.

plant biology↗

bioRxiv2024

Beyond enumeration: Phenotype independent "labeling-capture-release" process enabling precise detection of circulating tumour cells and downstream applications

Although strategies for circulating tumor cells (CTCs) enrichment have been proposed, the practical effects of clinical CTCs detection are far from satisfactory. Generally, the methodologies for CTCs detection aim at naturally occurring targets, but misdetection/interferences are prevalent due to the diverse phenotypes and subpopulations of CTCs with high heterogeneity. Herein, a CTCs isolation system based on the "labeling-capture-release" process is demonstrated for precise and high-efficient enrichment of CTCs from clinical blood samples. The mechanism which is based on abnormal glyco-metabolism of tumor cells including CTCs can be utilized for the surface decoration of CTCs with artificial azido groups. With the aid of bio-orthogonal plates designed with DBCO- and disulfide groups and exploiting the anti-fouling effects, the cells labeled with azido groups can be captured via a copper-free click reaction and released in a non-destructive manner during subsequent disulfide reduction. The technique is demonstrated to label multiple different types of tumor cells with the EpCAM+/- phenotypes and adherent/suspended status, and all the epithelial/interstitial/hybrid phenotypes of CTCs can be separated from clinical blood samples from 25 patients with 10 different cancer types. Moreover, our strategy is superior to the clinically approved CTCs detection system from the perspective of broad-spectrum and accurate recognition of heterogeneous CTCs. The capturing efficiency of this isolation system is over 80% and the release efficiency exceeds 90%. Most of the released CTCs survive with maintained glycolytic activity thus boding well for downstream applications such as drug susceptibility tests using viable CTCs.

bioengineering↗

bioRxiv2024

The L-lactate dehydrogenases of Pseudomonas aeruginosa are conditionally regulated but both contribute to survival during macrophage infection

Pseudomonas aeruginosa is an opportunistic pathogen that thrives in environments associated with human activity, including soil and water altered by agriculture or pollution. Because L-lactate is a significant product of plant and animal metabolism, it is available to serve as a carbon source for P. aeruginosa in the diverse settings it inhabits. Here, we evaluate P. aeruginosas production and use of its redundant L-lactate dehydrogenases, termed LldD and LldA. We confirm that the protein LldR represses lldD and identify a new transcription factor, called LldS, that activates lldA; these distinct regulators and the genomic contexts of lldD and lldA contribute to their differential expression. We demonstrate that the lldD and lldA genes are conditionally controlled in response to lactate isomers as well as to glycolate and {square}-hydroxybutyrate, which, like lactate, are {square}-hydroxycarboxylates. We also show that lldA is induced when iron availability is low. Our examination of lldD and lldA expression across depth in biofilms indicates a complex pattern that is consistent with the effects of glycolate production, iron availability, and cross-regulation on enzyme preference. Finally, macrophage infection assays revealed that both lldD and lldA contribute to persistence within host cells, underscoring the potential role of L-lactate as a carbon source during P. aeruginosa-eukaryote interactions. Together, these findings help us understand the metabolism of a key resource that may promote P. aeruginosas success as a resident of contaminated environments and animal hosts. ImportancePseudomonas aeruginosa is a major cause of lung infections in people with cystic fibrosis, hospital-acquired infections, and wound infections. It consumes L-lactate, which is found at substantial levels in human blood and tissues. In this study, we investigated the spatial regulation of two redundant enzymes, called LldD and LldA, which enable L-lactate metabolism in P. aeruginosa biofilms. We uncovered mechanisms and identified compounds that control P. aeruginosas LldD/LldA preference. We also showed that both enzymes contribute to its ability to survive within macrophages, a behavior that is thought to augment the chronicity and recalcitrance of infections. Our findings shed light on a key metabolic strategy used by P. aeruginosa and have the potential to inform the development of therapies targeting bacterial metabolism during infection.

microbiology↗

bioRxiv2023

Mosquito virus diversity in Western and North-western Uganda

IntroductionSurveillance for mosquito borne arboviruses in Sub Saharan Africa has largely focussed on known viruses including Yellow fever virus (YFV), Rift Valley fever virus (RVFV), Chikungunya virus (CHIKV), West Nile virus (WNV) and Dengue viruses (DENV). Routine surveillance and outbreak investigations traditionally rely on serology, PCR and cell culture. Although such methods are useful, they are do not detect novel or unexpected viruses. MethodsThis study employed unbiased metagenomic next generation sequencing (MNGS) to characterise viruses circulating in mosquitoes of Arua and Kasese districts of Uganda collected systematically as part the ArboViral Infection (AVI) study. Adult mosquito sampling was carried out from multiple sites using light traps baited with solid carbon dioxide (indoors) and pyrethrum spray (outdoors). 10,026 mosquitoes were identified using appropriate morphological identification keys and separated into 96 pools by species and location of collection. Viral RNA extracted from homogenised mosquitoes was reverse transcribed to complimentary DNA and sequenced on the flow cell of an Illumina NextSeq platform. Bioinformatic analysis was performed using a customized in-house metagenomics pipeline. Downstream analyses were carried out in R version 4.2.1. Results97 viruses from 24 families and 31 genera were detected in 96 mosquito pools from Arua and Kasese districts. The abundance of viruses in different families was in the order Rhabdoviridae (33), Flaviviridae (28), Orthomyxoviridae (13), Mesoniviridae (9), Piconarviridae (9), Peribunyaviridae (9), Phasmaviridae (9), Iflaviridae (8), Phenuiviridae (7), Nodaviridae (5), Xinmoviridae (4), Reoviridae (4), Virusidae (2), Nairoviridae (2), Qinviridae (2), Togaviridae (2), Alphatetraviridae (2), Picornaviralesidae (2), Iridoviridae, (1), Nudiviridae (1), Parvoviridae (1), Permutetraviridae (1) and 22 viruses from different families remain unclassified. The Flaviviridae, Togaviridae, Rhabdoviridae, Peribunyaviridae, Phenuiviridae, Nairoviridae, Nodaviridae and Orthomyxoviridae harbor viruses that cause disease in humans and other mammals. Viruses from insect-specific virus families that were detected included the Chuviridae, Iflaviridae, Phasmaviridae and Alphatetraviridae. 92/173 (53%) of the viral genomes had full opening reading frames (ORFs) and diverged by >30% nucleotide pairwise distance from the nearest reference genome. ConclusionThe majority of viruses detected were novel species described for the first time with unknown potential to cause disease. The diversity of virus species in mosquitoes from Uganda, a hotspot for emerging arboviruses has been only partially characterized. This study, carried out in Western and North West Uganda illustrates the scale of richness and virus diversity in the region and the need to further characterise the virome in mosquitoes, especially those with a propensity to feed from human and animal hosts.

genomics↗

bioRxiv2023

Temperature elevation synergises with and enhances the type-I IFN-mediated restriction of MPXV

Fever is an evolutionary conserved host pro-inflammatory immune response that governs the regulation of multiple biological processes to control the outcome of infection. In January 2022, the World Health Organization (WHO) reported a global outbreak in mpox cases with a high incidence of human-to-human transmission. A frequent prodromal symptom of monkeypox virus (MPXV) infection is fever, with a febrile temperature range of 38.3 to 40.5 {degrees}C. However, the outcome of temperature elevation on MPXV infection remains poorly defined. Here, we isolated a circulating strain of MPXV from a patient who presented with fever (38.5 {degrees}C) and rash from the 2022 outbreak. Genomic sequencing identified this isolate to belong to the epidemic Clade IIb.B1. Transcriptomic analysis of infected cells demonstrated this virus to induce a strong IL6 pro-inflammatory immune response, consistent with a role for this pyrogen in the regulation of fever. We identify host-cell temperature at both physiological skin (33 {degrees}C) and clinical febrile temperatures (38.5 and 40 {degrees}C) to be a key determinant in the outcome of infection through the differential regulation of MPXV transcription and associated amplitude of host cytokine response to infection. Incubation of infected cells at 38.5 or 40 {degrees}C led to a restriction or ablation in MPXV replication, respectively. Importantly, this thermal inhibition was reversible upon temperature downshift to 37 {degrees}C without detriment to viral replication fitness. Co-stimulation of the type-I interferon (IFN) response led to a dose- and temperature-dependent inhibition in MPXV replication that restricted the re-establishment of infection upon temperature downshift and withdrawal of IFN as an immune stimulus. Our data identify febrile temperatures associated with mpox disease to be a critical component of the host pro-inflammatory immune response to infection which can synergise with the type-I IFN response to enhance the host-cell mediated restriction of MPXV.

microbiology↗

bioRxiv2023

Massively parallel in vivo Perturb-seq reveals cell type-specific transcriptional networks in cortical development

Systematic analysis of gene function across diverse cell types in vivo is hindered by two challenges: obtaining sufficient cells from live tissues and accurately identifying each cells perturbation in high-throughput single-cell assays. Leveraging AAVs versatile cell type tropism and high labeling capacity, we expanded the resolution and scale of in vivo CRISPR screens: allowing phenotypic analysis at single-cell resolution across a multitude of cell types in the embryonic brain, adult brain, and peripheral nervous system. We undertook extensive tests of 86 AAV serotypes, combined with a transposon system, to substantially amplify labeling and accelerate in vivo gene delivery from weeks to days. Using this platform, we performed an in utero genetic screen as proof-of-principle and identified pleiotropic regulatory networks of Foxg1 in cortical development, including Layer 6 corticothalamic neurons where it tightly controls distinct networks essential for cell fate specification. Notably, our platform can label >6% of cerebral cells, surpassing the current state-of-the-art efficacy at <0.1% (mediated by lentivirus), and achieve analysis of over 30,000 cells in one experiment, thus enabling massively parallel in vivo Perturb-seq. Compatible with various perturbation techniques (CRISPRa/i) and phenotypic measurements (single-cell or spatial multi-omics), our platform presents a flexible, modular approach to interrogate gene function across diverse cell types in vivo, connecting gene variants to their causal functions.

neuroscience↗

bioRxiv2023

Inhibition of CPEB3 ribozyme elevates CPEB3 protein expression and polyadenylation of its target mRNAs, and enhances object location memory

A self-cleaving ribozyme that maps to an intron of the cytoplasmic polyadenylation element binding protein 3 (CPEB3) gene is thought to play a role in human episodic memory, but the underlying mechanisms mediating this effect are not known. We tested the activity of the murine sequence and found that the ribozymes self-scission half-life matches the time it takes an RNA polymerase to reach the immediate downstream exon, suggesting that the ribozyme-dependent intron cleavage is tuned to co-transcriptional splicing of the CPEB3 mRNA. Our studies also reveal that the murine ribozyme modulates maturation of its harboring mRNA in both cultured cortical neurons and the hippocampus: inhibition of the ribozyme using an antisense oligonucleotide leads to increased CPEB3 protein expression, which enhances polyadenylation and translation of localized plasticity-related target mRNAs, and subsequently strengthens hippocampal-dependent long-term memory. These findings reveal a previously unknown role for self-cleaving ribozyme activity in regulating experience-induced co-transcriptional and local translational processes required for learning and memory. Significance StatementCytoplasmic polyadenylation-induced translation is one of the key steps for regulating protein synthesis and neuroplasticity in the hippocampus. The CPEB3 ribozyme is a highly conserved mammalian self-cleaving catalytic RNA with unknown biological roles. In this study, we investigated how the intronic ribozyme affects the CPEB3 mRNA maturation and translation, and its subsequent effect on memory formation. Our findings show that the ribozyme activity is anti-correlated with CPEB3 mRNA splicing: inhibition of the ribozyme results in higher mRNA and protein levels, which contribute to long-term memory. Our studies offer new insights into the role of the CPEB3 ribozyme in neuronal translational control for the activity-dependent synaptic functions that underlie long-term memory and demonstrate a novel biological role for self-cleaving ribozymes.

neuroscience↗

bioRxiv2023

In vitro methylation of the U7 snRNP subunits Lsm11 and SmE by the PRMT5/MEP50/pICln methylosome

U7 snRNP is a multi-subunit endonuclease required for 3 end processing of metazoan replication-dependent histone pre-mRNAs. In contrast to the spliceosomal snRNPs, U7 snRNP lacks the Sm subunits D1 and D2 and instead contains two related proteins, Lsm10 and Lsm11. The remaining five subunits of the U7 heptameric Sm ring, SmE, F, G, B and D3, are shared with the spliceosomal snRNPs. The pathway that assembles the unique ring of U7 snRNP is unknown. Here, we show that a heterodimer of Lsm10 and Lsm11 tightly interacts with the methylosome, a complex of the arginine methyltransferase PRMT5, MEP50 and pICln known to methylate arginines in the C-terminal regions of the Sm proteins B, D1 and D3 during the spliceosomal Sm ring assembly. Both biochemical and Cryo-EM structural studies demonstrate that the interaction is mediated by PRMT5, which binds and methylates two arginine residues in the N-terminal region of Lsm11. Surprisingly, PRMT5 also methylates an N-terminal arginine in SmE, a subunit that does not undergo this type of modification during the biogenesis of the spliceosomal snRNPs. An intriguing possibility is that the unique methylation pattern of Lsm11 and SmE plays a vital role in the assembly of the U7 snRNP.

biochemistry↗