Humans have approximately 400 intact odorant receptors, but each individual has a unique set of genetic variations that lead to variation in olfactory perception. We used a heterologous assay to determine how often genetic polymorphisms in odorant receptors alter receptor function. We identified agonists for 18 odorant receptors and found that 63% of the odorant receptors we examined had polymorphisms that altered in vitro function. On average, two individuals differ functionally at over 30% of their odorant receptor alleles. To show that these in vitro results are relevant to olfactory perception, we verified that variations in OR10G4 genotype explain over 15% of the observed variation in perceived intensity and over 10% of the observed variation in perceived valence for the high affinity in vitro agonist guaiacol, but do not explain phenotypic variation for the lower affinity agonists vanillin and ethyl vanillin.
Summary Zika virus (ZIKV) infects fetal and adult human brain, and is associated with serious neurological complications. To date, no therapeutic treatment is available to treat ZIKV infected patients. We performed a high content chemical screen using human embryonic stem cell derived cortical neuron progenitor cells (hNPCs) and found that hippeastrine hydrobromide (HH) and amodiaquine dihydrochloride dihydrate (AQ), can inhibit ZIKV infection in hNPCs. Further validation showed that HH also rescues ZIKV-induced growth and differentiation defects in hNPCs and human fetal-like forebrain organoids. Finally, HH and AQ inhibit ZIKV infection in adult mouse brain in vivo. Strikingly, HH suppresses viral propagation when administered to adult mice with active ZIKV infection, highlighting its therapeutic potential. Our approach highlights the power of stem cell-based screens and validation in human forebrain organoids and mouse models in identifying drug candidates for treating ZIKV infection and related neurological complications in fetal and adult patients.
Although the human olfactory system is capable of discriminating a vast number of odors, we do not currently understand what chemical features are encoded by olfactory receptors. In large part this is due to a paucity of data in a search space covering the interactions of hundreds of receptors with billions of odorous molecules. Of the approximately 400 intact human odorant receptors, only 10% have a published ligand. Here we used a heterologous luciferase assay to screen 73 odorants against a clone library of 511 human olfactory receptors. This dataset will allow other researchers to interrogate the combinatorial nature of olfactory coding.
Key pointsr Carotid body (CB) glomus cells mediate acute oxygen sensing and the initiation of the hypoxic ventilatory response, yet the gene expression profile of these cells is not available.r We demonstrate that the single cell RNA-Seq method is a powerful tool for identifying highly expressed genes in CB glomus cells.r Our single cell RNA-Seq results characterized novel CB glomus cell genes, including members of the G protein-coupled receptor signalling pathway, ion channels and atypical mitochondrial electron transport chain subunits.r A heterologous cell-based screening identified acetate (which is known to affect CB glomus cell activity) as an agonist for the most highly abundant G protein-coupled receptor (Olfr78) in CB glomus cells.r These data established the first transcriptome profile of CB glomus cells, highlighting genes with potential implications in CB chemosensory function. AbstractThe carotid body (CB) is a major arterial chemoreceptor containing glomus cells whose activities are regulated by changes in arterial blood content, including oxygen. Despite significant advancements in the characterization of their physiological properties, our understanding of the underlying molecular machinery and signalling pathway in CB glomus cells is still limited. To overcome this, we employed the single cell RNA-Seq method by performing next-generation sequencing on single glomus cell-derived cDNAs to eliminate contamination of genes derived from other cell types present in the CB. Using this method, we identified a set of genes abundantly expressed in glomus cells, which contained novel glomus cell-specific genes. Transcriptome and subsequent in situ hybridization and immunohistochemistry analyses identified abundant G protein-coupled receptor signalling pathway components and various types of ion channels, as well as members of the hypoxia-inducible factors pathway. A short-chain fatty acid olfactory receptor Olfr78, recently implicated in CB function, was the most abundant G protein-coupled receptor. Two atypical mitochondrial electron transport chain subunits (Ndufa4l2 and Cox4i2) were among the most specifically expressed genes in CB glomus cells, highlighting their potential roles in mitochondria-mediated oxygen sensing. The wealth of information provided by the present study offers a valuable foundation for identifying molecules functioning in the CB.
In Brief E et al. reveal an unexpected function of antimicrobial peptides as the signal molecule to trigger aging-and infectionassociated dendrite degeneration and show that an epidermally expressed antimicrobial peptide can activate a conserved neuronal GPCR to cause dendrite degeneration through autophagic machinery. E et al., 2018, Neuron 97, 125- SUMMARYInfections have been identified as possible risk factors for aging-related neurodegenerative diseases, but it remains unclear whether infection-related immune molecules have a causative role in neurodegeneration during aging. Here, we reveal an unexpected role of an epidermally expressed antimicrobial peptide, NLP-29 (neuropeptide-like protein 29), in triggering aging-associated dendrite degeneration in C. elegans. The age-dependent increase of nlp-29 expression is regulated by the epidermal tir-1/ SARM-pmk-1/p38 MAPK innate immunity pathway. We further identify an orphan G protein-coupled receptor NPR-12 (neuropeptide receptor 12) acting in neurons as a receptor for NLP-29 and demonstrate that the autophagic machinery is involved cell autonomously downstream of NPR-12 to transduce degeneration signals. Finally, we show that fungal infections cause dendrite degeneration using a similar mechanism as in aging, through NLP-29, NPR-12, and autophagy. Our findings reveal an important causative role of antimicrobial peptides, their neuronal receptors, and the autophagy pathway in aging-and infection-associated dendrite degeneration.
SUMMARY Genome wide association studies (GWAS) have increased our knowledge of loci associated with a range of human diseases. However, applying such findings to elucidate pathophysiology and promote drug discovery remains challenging. Here, we created isogenic human embryonic stem cells (hESCs) with mutations in GWAS-identified susceptibility genes for type 2 diabetes. In pancreatic betalike cells differentiated from these lines, we found that mutations in CDKAL1, KCNQ1 and KCNJ11 led to impaired glucose secretion in vitro and in vivo, coinciding with defective glucose homeostasis. CDKAL1 mutant insulin+ cells were also hypersensitive to glucolipotoxicity. A high-content chemical screen identified a candidate drug that rescued CDKAL1 specific defects in vitro and in vivo by inhibiting the FOS/JUN pathway. Our approach of a proof-of-principle platform, which uses isogenic hESCs for functional evaluation of GWAS-identified loci and identification of a drug candidate that rescues gene-specific defects, paves the way for precision therapy of metabolic diseases.
Highlights d hPSC-based multiplex platform for interrogation of autismassociated mutations d Prefrontal cortex paradigm in hPSCs identifies autism-related neurogenesis defects d Abnormal WNT/bcatenin responses in class of autism genes with neurogenesis defects d Endophenotypes in hPSCs correlate with clinical data in autism patients
Diabetes is linked to loss of pancreatic beta-cells. Pluripotent stem cells offer a valuable source of human beta-cells for basic studies of their biology and translational applications. However, the signalling pathways that regulate beta-cell development and functional maturation are not fully understood. Here we report a high content chemical screen, revealing that H1152, a ROCK inhibitor, promotes the robust generation of insulin-expressing cells from multiple hPSC lines. The insulin expressing cells obtained after H1152 treatment show increased expression of mature beta cell markers and improved glucose stimulated insulin secretion. Moreover, the H1152-treated beta-like cells show enhanced glucose stimulated insulin secretion and increased capacity to maintain glucose homeostasis after transplantation. Conditional gene knockdown reveals that inhibition of ROCKII promotes the generation and maturation of glucose-responding cells. This study provides a strategy to promote human beta-cell maturation and identifies an unexpected role for the ROCKII pathway in the development and maturation of beta-like cells.
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