Biology subjects

zhang, q.

Publications and source records attributed to zhang, q..

4 recordsLinked to original sources

bioRxiv2023

Force sensing GPR133 is essential for normal balance and modulates vestibular hair cell membrane excitability via Gi signaling and CNGA3 coupling

The maintenance of normal balance sensing is a fundamental prerequisite for virtually every activity of daily life. As one set most important balance information collectors, vestibular hair cells convert mechanical stimuli from head movement into electrical signals through a mechanoelectrical transduction (MET) process. The molecular mechanism underlying equilibrioception and MET in vestibular hair cells is not well understood but is generally believed to be mediated by ion channels. However, whether these procedures are also mediated by other receptors, such as GPCRs, which are known to govern light, odorant and taste sensing, is not known. Here, by screening the expression of force-sensitive adhesion GPCRs in vestibular hair cells and phenotype profiling in animal models, we identified that a seven-transmembrane receptor, GPR133, was able to sense force in utricle hair cells and was required for maintenance of normal equilibrioception. Notably, GPR133 converted mechanical stimuli into changes in intracellular cAMP levels through Gi engagement and then modulated plasma membrane excitability and mediated MET by coupling to CNGA3 activity changes in approximately 30% of GPR133-expressing utricle hair cells. GPR133-mediated MET and its coupling with CNGA3 were recapitulated by an in vitro reconstitution system. Further chemical labeling, mass spectrometry and cryo-EM analysis provided potential structural information on force-induced GPR133 activation and Gi3 engagement. Collectively, our findings reveal the essential role of GPR133 in the maintenance of normal equilibrioception and suggest that GPCR family members can participate in the MET process in utricle hair cells through modulation of intracellular second messenger levels and ion channel coupling.

physiology↗

bioRxiv2023

Force sensation by a Gs-coupled adhesion GPCR mediates equilibrioception by increasing vestibular hair cell membrane excitability and CNGA3 coupling

AbstractEquilibrioception is essential for the perception and navigation of mammals in the three-dimensional world. A rapid mechanoelectrical transduction (MET) response in vestibular hair cells plays a critical role in positional and motional perception. Here, we identified that the G protein-coupled receptor LPHN2/ADGRL2, which is expressed in the apical membrane of utricular hair cells, is required for maintenance of normal balance. Hair cell-specific Lphn2 deficiency in mice impaired both balance behaviors and MET responses. Functional analyses using Pou4f3-CreER+/-; Lphn2fl/fl mice and LPHN2-specific inhibitors revealed that LPHN2 regulated the tip link-independent MET current at the apical surface of the utricular hair cell by converting force stimuli into transmembrane channel-like protein 1 (TMC1) activity. Force sensation by LPHN2 also induced glutamate release and calcium signaling in utricular hair cells. Reintroduction of LPHN2 into the hair cells of Lphn2-deficient mice restored the vestibular functions and MET responses. Our data suggest an indispensable role for a mechanosensitive GPCR in equilibrioception.

physiology↗

bioRxiv2021

Detection of Multiple Types of Cancer Driver Mutations Us-ing Targeted RNA Sequencing in NSCLC

Currently, DNA and RNA are used separately to capture different types of gene mutations. DNA is commonly used for the detection of SNVs, indels and CNVs; RNA is used for analysis of gene fusion and gene expression. To perform both DNA sequencing (DNA-seq) and RNA-seq, material is divided into two copies, and two different procedures are required for sequencing. Due to overconsumption of samples and experimental process complexity, it is necessary to create an experimental method capable of analyzing SNVs, indels, fusions and expression. We developed an RNA-based hybridization capture panel targeting actionable driver oncogenes in solid tumors and corresponding sample preparation and bioinformatics workflows. Analytical validation with an RNA standard reference containing 16 known fusion mutations and 6 SNV mutations demonstrated a detection specificity of 100.0% [95% CI 88.7%~100.0%] for SNVs and 100.0% [95% CI 95.4%~100.0%] for fusions. The targeted RNA panel achieved a 0.73-2.63 copies/ng RNA lower limit of detection (LOD) for SNVs and 0.21-6.48 copies/ng RNA for fusions. Gene expression analysis revealed a correlation greater than 0.9 across all 15 cancer-related genes between the RNA-seq results and targeted RNA panel. Among 1253 NSCLC FFPE tumor samples, multiple mutation types were called from DNA- and RNA-seq data and compared between the two assays. The DNA panel detected 103 fusions and 21 METex14 skipping events; 124 fusions and 26 METex14 skipping events were detected by the target RNA panel; 21 fusions and 4 METex14 skipping events were only detected by the target RNA panel. Among the 173 NSCLC samples negative for targetable mutations by DNA-seq, 15 (15/173, 8.67%) showed targetable gene fusions that may change clinical decisions with RNA-seq. In total, 226 tier I and tier II missense variants for NSCLC were analyzed at genomic (DNA-seq) and transcriptomic (RNA-seq) levels. The positive percent agreement (PPA) was 97.8%, and the positive predictive value (PPV) was 98.6%. Interestingly, variant allele frequencies were generally higher at the RNA level than at the DNA level, suggesting relatively dominant expression of mutant alleles. PPA was 97.6% and PPV 99.38% for EGFR 19del and 20ins variants. We also explored the relationship of RNA expression with gene copy number and protein expression. The RPKM of EGFR transcripts assessed by the RNA panel showed a linear relationship with copy number quantified by the DNA panel, with an R of 0.8 in 1253 samples. In contrast, MET gene expression is regulated in a more complex manner. In IHC analysis, all 3+ samples exhibited higher RPKM levels; IHC level of 2+ and below showed lower RNA expression. Parallel DNA- and RNA-seq and systematic analysis demonstrated the accuracy and robustness of the RNA sequencing panel in identifying multiple types of variants for cancer therapy. Contact: zhaojia0327@126.com

molecular biology↗

bioRxiv2019

The Structural Basis for Glycerol Permeation by human AQP7

Human glycerol channel AQP7 conducts glycerol release from adipocyte and entry into the cells in pancreatic islets, muscles and kidney tubule, and thus regulate glycerol metabolism in those tissues. Compared with other human aquaglyceroporins, AQP7 shows a less conserved "NPA" motif in the center cavity, and a pair of aromatic residues at Ar/R selectivity filter. To understand the structural basis for the glycerol conductance, we crystallized the human AQP7 and determined the structure at 3.7 [A]. A substrate binding pocket was found near to the Ar/R filter and the bound glycerol molecule stabilized by R229. In vivo functional assay on human AQP7 as well as AQP3 and AQP10 demonstrated strong glycerol transportation activities at physiological condition. The human AQP7 structure reveals a fully closed conformation with its permeation pathway strictly confined by Ar/R filter at the exoplasmic side and the gate at the cytoplasmic side, and the dislocation of the residues at narrowest parts of glycerol pathway in AQP7 play a critical role in controlling the glycerol flux.

molecular biology↗