Catechol estrogens stimulate insulin secretion in pancreatic β-cells via activation of the transient receptor potential A1 (TRPA1) channel

Estrogen hormones play an important role in controlling glucose homeostasis and pancreatic β-cell function. Despite the significance of estrogen hormones for regulation of glucose metabolism, little is known about the roles of endogenous estrogen metabolites in modulating pancreatic β-cell function. In this study, we evaluated the effects of major natural estrogen metabolites, catechol estrogens, on insulin secretion in pancreatic β-cells. We show that catechol estrogens, hydroxylated at positions C2 and C4 of the steroid A ring, rapidly potentiated glucose-induced insulin secretion via a nongenomic mechanism. 2-Hydroxyestrone, the most abundant endogenous estrogen metabolite, was more efficacious in stimulating insulin secretion than any other tested catechol estrogens. In insulin-secreting cells, catechol estrogens produced rapid activation of calcium influx and elevation in cytosolic free calcium. Catechol estrogens also generated sustained elevations in cytosolic free calcium and evoked inward ion current in HEK293 cells expressing the transient receptor potential A1 (TRPA1) cation channel. Calcium influx and insulin secretion stimulated by estrogen metabolites were dependent on the TRPA1 activity and inhibited with the channel-specific pharmacological antagonists or the siRNA. Our results suggest the role of estrogen metabolism in a direct regulation of TRPA1 activity with potential implications for metabolic diseases

The first genome sequence of a metatherian herpesvirus: Macropodid herpesvirus 1

While many placental herpesvirus genomes have been fully sequenced, the complete genome of a marsupial herpesvirus has not been described. Here we present the first genome sequence of a metatherian herpesvirus, Macropodid herpesvirus 1 (MaHV-1)

The first genome sequence of a metatherian herpesvirus: Macropodid herpesvirus 1

While many placental herpesvirus genomes have been fully sequenced, the complete genome of a marsupial herpesvirus has not been described. Here we present the first genome sequence of a metatherian herpesvirus, Macropodid herpesvirus 1 (MaHV-1)

The first genome sequence of a metatherian herpesvirus: Macropodid herpesvirus 1

Background: While many placental herpesvirus genomes have been fully sequenced, the complete genome of a marsupial herpesvirus has not been described. Here we present the first genome sequence of a metatherian herpesvirus, Macropodid herpesvirus 1 (MaHV-1). Results: The MaHV-1 viral genome was sequenced using an Illumina MiSeq sequencer, de novo assembly was performed and the genome was annotated. The MaHV-1 genome was 140 kbp in length and clustered phylogenetically with the primate simplexviruses, sharing 67 % nucleotide sequence identity with Human herpesviruses 1 and 2. The MaHV-1 genome contained 66 predicted open reading frames (ORFs) homologous to those in other herpesvirus genomes, but lacked homologues of UL3, UL4, UL56 and glycoprotein J. This is the first alphaherpesvirus genome that has been found to lack the UL3 and UL4 homologues. We identified six novel ORFs and confirmed their transcription by RT-PCR. Conclusions: This is the first genome sequence of a herpesvirus that infects metatherians, a taxonomically unique mammalian clade. Members of the Simplexvirus genus are remarkably conserved, so the absence of ORFs otherwise retained in eutherian and avian alphaherpesviruses contributes to our understanding of the Alphaherpesvirinae. Further study of metatherian herpesvirus genetics and pathogenesis provides a unique approach to understanding herpesvirus-mammalian interactions

Supplementary Fig. Relative transcript levels for the unique hypothetical MaHV-1 ORFs

<a href="https://static-content.springer.com/esm/art%3A10.1186%2Fs12864-016-2390-2/MediaObjects/12864_2016_2390_MOESM1_ESM.tif">Additional file 1: Figure S1.</a> Relative transcript levels for the unique hypothetical MaHV-1 ORFs PW1 to PW6 at 4 h (grey bars) and 12 h (black bars) post infection (hpi) in wallaby fibroblast cells. Expression was normalised to the host housekeeping gene, GAPDH, and analysed by calculating mean normalised expression values. No viral transcripts were detected in uninfected cells. Error bars indicate standard deviation of three biological replicates

Phylogenetic relationship between macropodid herpesvirus 1 (MaHV1) and other viruses from the Alphaherpesvirinae subfamily.

<div>The relationship between macropodid herpesvirus 1 (MaHV1) and other viruses from the Alphaherpesvirinae subfamily. Neighbour-joining distance trees were generated using the translated protein sequences of conserved herpesvirus ORFs: a glycoprotein B, gB (UL27); b DNA polymerase, DPOL (UL30); and c glycoprotein D, gD (US6). Bootstrap values (10,000 replicates) are shown for each branch, with Gallid herpesvirus 2 (GaHV2) as an outgroup. The abbreviations and GenBank accession details are: macropodid herpesvirus 2 (MaHV2) [GenBank:AAD11961, GenBank:AAL13143, gB and gD], macropodid herpesvirus 4 (MaHV4) [GenBank:AGC54689, gB], macacine herpesvirus 1 (HBV) [GenBank:AAA85652, GenBank:NP_851890, GenBank:AAB24129, gB, DPOL and gD], fruit bat herpesvirus 1 (FbHV1) [GenBank:YP_009042089, GenBank:YP_009042092, GenBank:YP_009042126, gB, DPOL and gD], gallid herpesvirus 1 (GaHV1) [GenBank:AEB97319 , GenBank:AEB97322, GenBank:AEB97368, gB, DPOL and gD], gallid herpesvirus 2 (GaHV2) [GenBank:CAA63039, GenBank:YP_001033959, GenBank:AAA64967, gB, DPOL and gD], human herpesvirus 1 (HHV1) [GenBank:NP_044629, GenBank:NP_044632, GenBank:CAA32283, gB, DPOL and gD], human herpesvirus 2 (HHV2) [GenBank:NP_044497, GenBank:NP_044500, GenBank:AAB60553, gB, DPOL and gD], human herpesvirus 3 (HHV3) [GenBank:NP_040154.2, GenBank:NP_040151, gB and DPOL], saimiriine herpesvirus 1 (SaHV1) [GenBank:YP_003933812, GenBank:YP_003933809, GenBank:ADO13836, gB, DPOL and gD], bovine herpesvirus 1 (BoHV1) [GenBank:AAA46013, GenBank:NP_045328, GenBank:CAA80604, gB, DPOL and gD], bovine herpesvirus 2 (BoHV2) [GenBank:P12641.2, GenBank:AAD55134:, gB and DPOL], bovine herpesvirus 5 (BoHV5) [GenBank:YP_003662497, GenBank:YP_003662494, GenBank:AAA67359, gB, DPOL and gD], canine herpesvirus 1 (CHV) [GenBank:AAK51052, GenBank:AAX47050, GenBank:AAB67058, gB, DPOL and gD], felid herpesvirus 1 (FeHV1) [GenBank:AAB28559, GenBank:YP_003331549, GenBank:BAA06442, gB, DPOL and gD], equine herpesvirus 1 (EHV1) [GenBank:Q6DLH8, GenBank:YP_053075, GenBank:AAA46073, gB, DPOL and gD], equine herpesvirus 4 (EHV4) [GenBank:NP_045250, GenBank:NP_045247, GenBank:NP_045289, gB, DPOL and gD], suid herpesvirus 1 (PRV) [GenBank:YP_068330, GenBank:YP_068333, GenBank:AAC36717, gB, DPOL and gD].</div

A Novel Humanized GLP-1 Receptor Model Enables Both Affinity Purification and Cre-LoxP Deletion of the Receptor

<div><p>Class B G protein-coupled receptors (GPCRs) are important regulators of endocrine physiology, and peptide-based therapeutics targeting some of these receptors have proven effective at treating disorders such as hypercalcemia, osteoporosis, and type 2 diabetes mellitus (T2DM). As next generation efforts attempt to develop novel non-peptide, orally available molecules for these GPCRs, new animal models expressing human receptor orthologs may be required because small molecule ligands make fewer receptor contacts, and thus, the impact of amino acid differences across species may be substantially greater. The objective of this report was to generate and characterize a new mouse model of the human glucagon-like peptide-1 receptor (hGLP-1R), a class B GPCR for which established peptide therapeutics exist for the treatment of T2DM. <i>hGLP-1R</i> knock-in mice express the receptor from the murine <i>Glp-1r</i> locus. Glucose tolerance tests and gastric emptying studies show <i>hGLP-1R</i> mice and their wild-type littermates display similar physiological responses for glucose metabolism, insulin secretion, and gastric transit, and treatment with the GLP-1R agonist, exendin-4, elicits similar responses in both groups. Further, ex vivo assays show insulin secretion from humanized islets is glucose-dependent and enhanced by GLP-1R agonists. To enable additional utility, the targeting construct of the knock-in line was engineered to contain both flanking LoxP sites and a C-terminal FLAG epitope. Anti-FLAG affinity purification shows strong expression of hGLP-1R in islets, lung, and stomach. We crossed the <i>hGLP-1R</i> line with <i>Rosa26Cre</i> mice and generated global <i>Glp-1r^−/−</i> animals. Immunohistochemistry of pancreas from humanized and knock-out mice identified a human GLP-1R-specific antibody that detects the GLP-1R in human pancreas as well as in the pancreas of <i>hGLP-1r</i> knock-in mice. This new <i>hGLP-1R</i> model will allow tissue-specific deletion of the <i>GLP-1R</i>, purification of potential GLP-1R partner proteins, and testing of novel therapeutic agents targeting the hGLP-1R.</p></div