Biology subjects

Li, M.-L.

Publications and source records attributed to Li, M.-L..

6 recordsLinked to original sources

bioRxiv2025

Human-specific features of the cerebellum and ZP2-regulated synapse development

Understanding the unique features of the human brain compared to non-human primates has long intrigued humankind. The cerebellum refines motor coordination and cognitive functions, contributing to the evolutionary development of human adaptability and dexterity. To identify shared and divergent features across primates, we conducted single-nucleus transcriptomic and chromatin accessibility profiling of the adult cerebellar cortex in humans, chimpanzees, macaques, and marmosets. We revealed human-specific transcriptomic and regulatory features, particularly those involved in synaptogenesis. Notably, we identified an enrichment of the sperm receptor zona pellucida glycoprotein 2 (ZP2) and its potential interactors, known for their roles in gamete interaction, in human granule cells. Experimental data show that ZP2 expression in human granule cells is induced by pontine mossy fibers, reducing synaptic proteins at pontocerebellar glomerular synapses, and decreasing cerebellar neuron electrophysiological activity. This unexpected co-option of ZP2 in human-specific synapse regulation provides insights into the evolutionary specialization of the human cerebellum.

neuroscience↗

bioRxiv2025

Fibroblast contractility drives network reorganization and epithelial proliferation in intestinal polyposis

Fibroblasts are critical regulators of epithelial homeostasis through mechanics and signaling. However, the regulatory principles governing fibroblast behaviors are largely unknown. Fibroblast dysregulation has emerged as a pathological contributor in epithelial diseases such as intestinal polyposis. Here, using an inducible Bmp-loss-of-function polyposis model, we define stepwise mechanisms unraveling how dysregulated signaling perturbs fibroblast behaviors, which in turn, disrupt their function to regulate epithelial homeostasis. Intriguingly, the first initiating event leading to epithelial polyps was architectural and impacted the fibroblasts, not the epithelium. Bmp signaling inhibition caused fibroblasts to become hypercontractile, leading to their reorganization from an elaborate network into collapsed clusters beneath crypts. Disrupted fibroblast mechanics and compartmentalization preceded epithelial hyperproliferation in polyps. Using in vitro models, we show that fibroblast hypercontractility not only disrupted fibroblast organization, but also enhanced the ability of fibroblasts to support epithelial growth, leading to epithelial dysregulation. Overall, our studies reveal stepwise regulatory mechanisms underlying fibroblast signaling, mechanics and organization critical for their function to regulate epithelial homeostasis.

cell biology↗

bioRxiv2023

Enterovirus Evolution Reveals the Mechanism of an RNA-Targeted Antiviral and Determinants of Viral Replication

Selective pressures on positive-strand RNA viruses provide opportunities to establish target site specificity and mechanisms of action of antivirals. Here, Enterovirus-A71 revertant viruses with resistant mutations in the SLII IRES domain (SLIIresist) were selected at low doses of the antiviral DMA-135. The EV-A71 revertant viruses were resistant to DMA-135 at concentrations that robustly inhibit replication of wild-type virus. EV-A71 IRES structures harboring the suppressor mutations induced efficient expression of reporter Luciferase mRNA in the presence of non-cytotoxic doses of DMA-135 whereas DMA-135 dose-dependently inhibited Luciferase expression from the wild-type IRES element. NMR studies indicate that the resistant mutations change the structure of SLII at the bulge loop binding site of DMA-135 and at part of an extended surface recognized by host RNA-binding protein AUF1. Comparisons of biophysical analysis of complexes formed between AUF1, DMA-135, or either SLII or SLIIresist show that DMA-135 stabilizes a ternary complex with AUF1-SLII but not AUF1-SLIIresist. Further studies demonstrate that the hnRNP A1 protein retains binding affinity for SLIIresist, illustrating that DMA-135 inhibition and viral resistance do not perturb the SLII-hnRNP A1 arm of the regulatory axis. Taken together, this work demonstrates how viral evolution under selective pressures of small molecules can elucidate RNA binding site specificity, mechanisms of action, and provide additional insights into the viral pathways inhibited by the antiviral DMA-135.

molecular biology↗

bioRxiv2020

Amilorides inhibit SARS-CoV-2 replication in vitro by targeting RNA structures

The SARS-CoV-2 pandemic, and the likelihood of future coronavirus pandemics, has rendered our understanding of coronavirus biology more essential than ever. Small molecule chemical probes offer to both reveal novel aspects of virus replication and to serve as leads for antiviral therapeutic development. The RNA-biased amiloride scaffold was recently tuned to target a viral RNA structure critical for translation in enterovirus 71, ultimately uncovering a novel mechanism to modulate positive-sense RNA viral translation and replication. Analysis of CoV RNA genomes reveal many conserved RNA structures in the 5-UTR and proximal region critical for viral translation and replication, including several containing bulge-like secondary structures suitable for small molecule targeting. Following phylogenetic conservation analysis of this region, we screened an amiloride-based small molecule library against a less virulent human coronavirus, OC43, to identify lead ligands. Amilorides inhibited OC43 replication as seen in viral plaque assays. Select amilorides also potently inhibited replication competent SARS-CoV-2 as evident in the decreased levels of cell free virions in cell culture supernatants of treated cells. Reporter screens confirmed the importance of RNA structures in the 5-end of the viral genome for small molecule activity. Finally, NMR chemical shift perturbation studies of the first six stem loops of the 5-end revealed specific amiloride interactions with stem loops 4, 5a, and 6, all of which contain bulge like structures and were predicted to be strongly bound by the lead amilorides in retrospective docking studies. Taken together, the use of multiple orthogonal approaches allowed us to identify the first small molecules aimed at targeting RNA structures within the 5-UTR and proximal region of the CoV genome. These molecules will serve as chemical probes to further understand CoV RNA biology and can pave the way for the development of specific CoV RNA-targeted antivirals.

microbiology↗

bioRxiv2020

Small Molecule Targeting IRES Domain Inhibits Enterovirus 71 Replication via an Allosteric Mechanism that Stabilizes a Ternary Complex

We herein report an RNA-targeting antiviral small molecule that reduces replication of the human enterovirus 71 (EV71) via stabilization of an inhibitory small molecule-RNA-protein ternary complex. The EV71 virus poses serious threats to human health, particularly in regions of Southeast Asia, and no FDA approved drugs or vaccines are available. We first screened an RNA-biased small molecule library using a peptide-displacement assay to identify ligands for the stem loop II structure of the EV71 internal ribosomal entry site, which was previously shown to impact viral translation and replication. One ligand, DMA-135, decreased viral translation and replication in cell-based studies in a dose-dependent manner with no significant toxicity. Structural, biophysical, and biochemical characterization support an allosteric mechanism in which DMA-135 induces a conformational change in the RNA structure that stabilizes a ternary complex with the AUF1 protein that then represses translation. This mechanism was further supported by pull-down experiments in cell culture. These detailed studies establish enterovirus RNA structures as promising drug targets while revealing an approach and mechanism of action that should be broadly applicable to functional RNA targeting.

biochemistry↗

bioRxiv2019

Real-time spectroscopic photoacoustic/ultrasound (PAUS) scanning with simultaneous fluence compensation and motion correction for quantitative molecular imaging

For over two decades photoacoustic (PA) imaging has been tested clinically, but successful human trials have been minimal. To enable quantitative clinical spectroscopy, the fundamental issues of wavelength-dependent fluence variations and inter-wavelength motion must be overcome. Here we propose a new real-time, spectroscopic photoacoustic/ultrasound (PAUS) imaging approach using a compact, 1-kHz rate wavelength-tunable laser. Instead of illuminating tissue over a large area, the fiber-optic delivery system surrounding an US array sequentially scans a narrow laser beam, with partial PA image reconstruction for each laser pulse. The final image is then formed by coherently summing partial images at a 50-Hz video rate. This scheme enables (i) automatic laser-fluence compensation in spectroscopic PA imaging and (ii) inter-wavelength motion correction using US speckle tracking, which have never been shown before in real-time systems. The 50-Hz video rate PAUS system is demonstrated in vivo using a murine model of drug delivery monitoring.

bioengineering↗