Biology subjects

Liou, R. H.-C.

Publications and source records attributed to Liou, R. H.-C..

3 recordsLinked to original sources

bioRxiv2025

Annexin A2 mediates RIG-I-like receptor responses to viral infection

RIG-I and MDA5 are critical sensors of virus infection and detect many important human pathogens including SARS-CoV-2, influenza A virus (IAV), Zika virus and West Nile virus (WNV). MDA5 is activated by binding to double-stranded RNA generated during infection and induces strong pro-inflammatory and antiviral responses mediated largely via type I interferons. Activation of MDA5 is also implicated in monogenic (e.g. Aicardi-Goutieres syndrome) and complex polygenic (e.g. systemic lupus erythematosus) autoinflammatory diseases, demonstrating the importance of appropriate regulation of this pathway. Yet, how MDA5 is regulated is poorly defined. We employed SILAC-mass spectrometry to discover MDA5 binding partners in virus-infected cells. Our screen revealed Annexin A2 (ANXA2) as an interactor of MDA5 specifically during infection. Direct binding of ANXA2 and MDA5 was recapitulated in vitro and required Ca2+, and ANXA2 promoted MDA5 oligomerisation in cells. Loss of ANXA2 in human cells significantly blunted interferon responses to EMCV, WNV and IAV, demonstrating a role for ANXA2 in activation of both MDA5 and RIG-I. Using human induced pluripotent stem cells, we found that ANXA2 was required for potent neuronal cell responses to WNV infection and macrophage responses to multiple viral pathogens. Thus, we find that ANXA2 is a critical mediator of RIG-I-like receptor pathways.

immunology↗

bioRxiv2023

The efficient induction of human retinal ganglion-like cells provides a platform for studying optic neuropathies

Retinal ganglion cells (RGCs) are essential for vision perception. In glaucoma and other optic neuropathies, RGCs and their optic axons undergo degenerative change and cell death; this can result in irreversible vision loss. Here we developed a rapid protocol for directly inducing RGC differentiation from human induced pluripotent stem cells (iPSCs) by the overexpression of ATOH7, BRN3B and SOX4. The hiPSC-derived RGC-like cells (iRGCs) show robust expression of various RGC-specific markers by whole transcriptome profiling. A functional assessment was also carried out and this demonstrated that these iRGCs display stimulus-induced neuronal activity, as well as spontaneous neuronal activity. Ethambutol (EMB), an effective first-line anti-tuberculosis agent, is known to cause serious visual impairment and irreversible vision loss due to the RGC degeneration in a significant number of treated patients. Using our iRGCs, EMB was found to induce significant dose-dependent and time-dependent increases in cell death and neurite degeneration. Western blot analysis revealed that the expression levels of p62 and LC3-II were upregulated, and further investigations revealed that EMB caused a blockade of lysosome-autophagosome fusion; this indicates that impairment of autophagic flux is one of the adverse effects of that EMB has on iRGCs. In addition, EMB was found to elevate intracellular reactive oxygen species (ROS) levels increasing apoptotic cell death. This could be partially rescued by the co-treatment with the ROS scavenger NAC. Taken together, our findings suggest that this iRGC model, which achieves both high yield and high purity, is suitable for investigating optic neuropathies, as well as being useful when searching for potential drugs for therapeutic treatment and/or disease prevention.

cell biology↗

bioRxiv2020

Combinatorial approach of binary colloidal crystals (BCCs) and CRISPR activation to improve induced pluripotent stem cell differentiation into neurons

Conventional methods of neuronal differentiation for human induced pluripotent stem cells (iPSCs) are tedious and complicated, involving multi-stage protocols with complex cocktails of growth factors and small molecules. Artificial extracellular matrix with defined surface topography and chemistry represents a promising venue to improve the neuronal differentiation in vitro. In the present study, we test the impact of a type of colloidal self-assembled patterns called binary colloidal crystals (BCCs) in neuronal differentiation. We developed a CRISPR activation (CRISPRa) iPSC platform that constitutively expresses the dCas9-VPR system, which allows robust activation of the proneural transcription factor NEUROD1 to rapidly induce neuronal differentiation within seven days. We showed that the combinatorial use of BCCs can further improve this neuronal differentiation system. In particular, our results indicate that fine tuning of silica and polystyrene size is critical to generate specific topographies to improve neuronal differentiation and branching. BCCs with 5 m silica and 100 nm carboxylated polystyrene has the most prominent effect on increasing neurite outgrowth and more complex ramification, while BCCs with 2m silica and 65nm carboxylated polystyrene is better in promoting neuronal enrichment. These results indicate that biophysical cues can support rapid differentiation and improve neuronal maturation. In summary, our combinatorial approach of CRISPRa and BCCs provides a robust and rapid pipeline for in vitro production of human neurons. Specific BCCs can be adapted to late stages of neuronal differentiation protocols to improve neuronal maturation, which have important implications in tissue engineering, in vitro biological studies and disease modeling.

bioengineering↗