Discovery of a Cushing's syndrome protein kinase A mutant that biases signaling through type I AKAPs.

Adrenal Cushing's syndrome is a disease of cortisol hypersecretion often caused by mutations in protein kinase A catalytic subunit (PKAc). Using a personalized medicine screening platform, we discovered a Cushing's driver mutation, PKAc-W196G, in ~20% of patient samples analyzed. Proximity proteomics and photokinetic imaging reveal that PKAc is unexpectedly distinct from other described Cushing's variants, exhibiting retained association with type I regulatory subunits (RI) and their corresponding A kinase anchoring proteins (AKAPs).

Contrastive Learning-Based Anomaly Detection for Actual Corporate Environments.

Information systems play an important role in business management, especially in personnel, budget, and financial management. If an anomaly ensues in an information system, all operations are paralyzed until their recovery. In this study, we propose a method for collecting and labeling datasets from actual operating systems in corporate environments for deep learning.

Classification of Cushing's syndrome PKAc mutants based upon their ability to bind PKI.

Cushing's syndrome is an endocrine disorder caused by excess production of the stress hormone cortisol. Precision medicine strategies have identified single allele mutations within the PRKACA gene that drive adrenal Cushing's syndrome. These mutations promote perturbations in the catalytic core of protein kinase A (PKAc) that impair autoinhibition by regulatory subunits and compartmentalization via recruitment into AKAP signaling islands.

Mislocalization of protein kinase A drives pathology in Cushing's syndrome.

Mutations in the catalytic subunit of protein kinase A (PKAc) drive the stress hormone disorder adrenal Cushing's syndrome. We define mechanisms of action for the PKAc-L205R and W196R variants. Proximity proteomic techniques demonstrate that both Cushing's mutants are excluded from A kinase-anchoring protein (AKAP)-signaling islands, whereas live-cell photoactivation microscopy reveals that these kinase mutants indiscriminately diffuse throughout the cell.

Development of a Novel SNAP-Epitope Tag/Near-Infrared Imaging Assay to Quantify G Protein-Coupled Receptor Degradation in Human Cells.

We have developed a novel reporter assay that leverages SNAP-epitope tag/near-infrared (NIR) imaging technology to monitor G protein-coupled receptor (GPCR) degradation in human cell lines. N-terminal SNAP-tagged GPCRs were subcloned and expressed in human embryonic kidney (HEK) 293 cells and then subjected to 24 h of cycloheximide (CHX)-chase degradation assays to quantify receptor degradation half-lives () using LICOR NIR imaging-polyacrylamide gel electrophoresis (PAGE) analysis. Thus far, we have used this method to quantify for all nine adrenergic (ADRA1A, ADRA1B, ADRA1D, ADRA2A, ADRA2B, ADRA2C, ADRB1, ADRB2, ADRB3), five somatostatin (SSTR1, SSTR2, SSTR3, SSTR4, SSTR5), four chemokine (CXCR1, CXCR2, CXCR3, CXCR5), and three 5-HT2 (5HT2A, 5HT2B, 5HT2C) receptor subtypes.

N-glycosylation of α-adrenergic receptor N-terminal domain is required for correct trafficking, function, and biogenesis.

G protein-coupled receptor (GPCR) biogenesis, trafficking, and function are regulated by post-translational modifications, including N-glycosylation of asparagine residues. α-adrenergic receptors (α-ARs) - key regulators of central and autonomic nervous system function - contain two putative N-glycosylation sites within the large N-terminal domain at N65 and N82. However, determining the glycosylation state of this receptor has proven challenging.

Scribble co-operatively binds multiple α-adrenergic receptor C-terminal PDZ ligands.

Many G protein-coupled receptors (GPCRs) are organized as dynamic macromolecular complexes in human cells. Unraveling the structural determinants of unique GPCR complexes may identify unique protein:protein interfaces to be exploited for drug development. We previously reported α-adrenergic receptors (α-ARs) - key regulators of cardiovascular and central nervous system function - form homodimeric, modular PDZ protein complexes with cell-type specificity.

Convergence of Canonical and Non-Canonical Wnt Signal: Differential Kat3 Coactivator Usage.

Background: The ancient and highly evolutionarily conserved Wnt signaling pathway is critical in nearly all tissues and organs for an organism to develop normally from embryo through adult. Wnt signaling is generally parsed into "canonical" or Wnt-β-catenin-dependent or "non-canonical" β-catenin-independent signaling. Even though designating Wnt signaling as either canonical or noncanonical allows for easier conceptual discourse about this signaling pathway, in fact canonical and non-canonical Wnt crosstalk regulates complex nonlinear networks.

Label-Free Dynamic Mass Redistribution Reveals Low-Density, Prosurvival -Adrenergic Receptors in Human SW480 Colon Carcinoma Cells.

Small molecules that target the adrenergic family of G protein-coupled receptors (GPCRs) show promising therapeutic efficacy for the treatment of various cancers. In this study, we report that human colon cancer cell line SW480 expresses low-density functional -adrenergic receptors (ARs) as revealed by label-free dynamic mass redistribution (DMR) signaling technology and confirmed by quantitative reverse-transcriptase polymerase chain reaction analysis. Remarkably, although endogenous -ARs are not detectable via either [H]-prazosin-binding analysis or phosphoinositol hydrolysis assays, their activation leads to robust DMR and enhanced cell viability.

Causality patterns and machine learning for the extraction of problem-action relations in discharge summaries.

Clinical narrative text includes information related to a patient's medical history such as chronological progression of medical problems and clinical treatments. A chronological view of a patient's history makes clinical audits easier and improves quality of care. In this paper, we propose a clinical Problem-Action relation extraction method, based on clinical semantic units and event causality patterns, to present a chronological view of a patient's problem and a doctor's action.

Endogenous N-terminal Domain Cleavage Modulates α1D-Adrenergic Receptor Pharmacodynamics.

The α1D-adrenergic receptor (ADRA1D) is a key regulator of cardiovascular, prostate, and central nervous system functions. This clinically relevant G protein-coupled receptor has proven difficult to study, as it must form an obligate modular homodimer containing the PDZ proteins scribble and syntrophin or become retained in the endoplasmic reticulum as non-functional protein. We previously determined that targeted removal of the N-terminal (NT) 79 amino acids facilitates ADRA1D plasma membrane expression and agonist-stimulated functional responses.

Dynamic mass redistribution reveals diverging importance of PDZ-ligands for G protein-coupled receptor pharmacodynamics.

G protein-coupled receptors (GPCRs) are essential membrane proteins that facilitate cell-to-cell communication and co-ordinate physiological processes. At least 30 human GPCRs contain a Type I PSD-95/DLG/Zo-1 (PDZ) ligand in their distal C-terminal domain; this four amino acid motif of X-[S/T]-X-[φ] sequence facilitates interactions with PDZ domain-containing proteins. Because PDZ protein interactions have profound effects on GPCR ligand pharmacology, cellular localization, signal-transduction effector coupling and duration of activity, we analyzed the importance of Type I PDZ ligands for the function of 23 full-length and PDZ-ligand truncated (ΔPDZ) human GPCRs in cultured human cells.

Individual protomers of a G protein-coupled receptor dimer integrate distinct functional modules.

Recent advances in proteomic technology reveal G-protein-coupled receptors (GPCRs) are organized as large, macromolecular protein complexes in cell membranes, adding a new layer of intricacy to GPCR signaling. We previously reported the α-adrenergic receptor (ADRA1D)-a key regulator of cardiovascular, urinary and CNS function-binds the syntrophin family of PDZ domain proteins (SNTA, SNTB1, and SNTB2) through a C-terminal PDZ ligand interaction, ensuring receptor plasma membrane localization and G-protein coupling. To assess the uniqueness of this novel GPCR complex, 23 human GPCRs containing Type I PDZ ligands were subjected to TAP/MS proteomic analysis.

The effects of bodyweight-based exercise with blood flow restriction on isokinetic knee muscular function and thigh circumference in college students.

[Purpose] The purpose of this study was to investigate the effects of bodyweight-based exercise with blood flow restriction on isokinetic muscular function and thigh circumference in college students. [Subjects and Methods] The subjects were 17 college students who were recruited and randomly assigned to bodyweight-based exercise with blood flow restriction and bodyweight-based exercise groups. Participants performed front lunges and squats at ratings of perceived exertion of 11-13 three times a week during a 6-week training period.

Effect of posterior pelvic tilt taping in women with sacroiliac joint pain during active straight leg raising who habitually wore high-heeled shoes: a preliminary study.

Objective: The purpose of this study was to assess whether a 1-day application of posterior pelvic tilt taping (PPTT) using a kinesiology tape would decrease anterior pelvic tilt and active straight leg raising test scores in women with sacroiliac joint who habitually wore high-heeled shoes.

Methods: Sixteen women (mean age, 23.63 ± 3.

Bioderived polyelectrolyte nanogels for robust antigen loading and vaccine adjuvant effects.

An easy but robust strategy for the synthesis of bioderived polyelectrolyte nanogels for protein antigen loading and vaccine adjuvant systems that can improve both humoral (Th2) and cellular immunity (Th1) is presented. The synthesized polyelectrolyte nanogels promote the uptake of antigens into antigen-presenting cells and strongly induce ovalbumin-specific INF-γ producing cells, cytotoxic T cell activity, and antibody production.

Syntrophin isoforms play specific functional roles in the α1D-adrenergic receptor/DAPC signalosome.

α(1D)-Adrenergic receptors, key regulators of cardiovascular system function, are organized as a multi-protein complex in the plasma membrane. Using a Type-I PDZ-binding motif in their distal C-terminal domain, α(1D)-ARs associate with syntrophins and dystrophin-associated protein complex (DAPC) members utrophin, dystrobrevin and α-catulin. Three of the five syntrophin isoforms (α, β(1) and β(2)) interact with α(1D)-ARs and our previous studies suggest multiple isoforms are required for proper α(1D)-AR function in vivo.

Monoclonal antibodies against Muscleblind-like 3, a protein with punctate nuclear localization.

Muscleblind-like 3 (MBNL3) belongs to a family of RNA binding proteins that regulate alternative splicing. We have generated a set of monoclonal antibodies (MAbs) against mouse MBNL3, three of which do not cross-react with the other muscleblind-like (MBNL) proteins, MBNL1 and MBNL2. Epitope mapping revealed that MAbs P1C7, P1E7, SP1C2, and P2E6 recognize distinct, non-overlapping segments of the MBNL3 polypeptide sequence.

Alpha-dystrobrevin-1 recruits alpha-catulin to the alpha1D-adrenergic receptor/dystrophin-associated protein complex signalosome.

α(1D)-Adrenergic receptors (ARs) are key regulators of cardiovascular system function that increase blood pressure and promote vascular remodeling. Unfortunately, little information exists about the signaling pathways used by this important G protein-coupled receptor (GPCR). We recently discovered that α(1D)-ARs form a "signalosome" with multiple members of the dystrophin-associated protein complex (DAPC) to become functionally expressed at the plasma membrane and bind ligands.

RNA-binding protein Muscleblind-like 3 (MBNL3) disrupts myocyte enhancer factor 2 (Mef2) {beta}-exon splicing.

Mammalian MBNL (muscleblind-like) proteins are regulators of alternative splicing and have been implicated in myotonic dystrophy, the most common form of adult onset muscular dystrophy. MBNL3 functions as an inhibitor of muscle differentiation and is expressed in proliferating muscle precursor cells but not in differentiated skeletal muscle. Here we demonstrate that MBNL3 regulates the splicing pattern of the muscle transcription factor myocyte enhancer factor 2 (Mef2) by promoting exclusion of the alternatively spliced β-exon.

Human papillomavirus type 16 E6-specific antitumor immunity is induced by oral administration of HPV16 E6-expressing Lactobacillus casei in C57BL/6 mice.

Given that local cell-mediated immunity (CMI) against the human papillomavirus type 16 E6 (HPV16 E6) protein is important for eradication of HPV16 E6-expressing cancer cells in the cervical mucosa, the HPV16 E6 protein may be a target for the mucosal immunotherapy of cervical cancer. Here, we expressed the HPV16 E6 antigen on Lactobacillus casei (L. casei) and investigated E6-specific CMI following oral administration of the L.

Lactic acid bacteria affect serum cholesterol levels, harmful fecal enzyme activity, and fecal water content.

Background: Lactic acid bacteria (LAB) are beneficial probiotic organisms that contribute to improved nutrition, microbial balance, and immuno-enhancement of the intestinal tract, as well as lower cholesterol. Although present in many foods, most trials have been in spreads or dairy products. Here we tested whether Bifidobacteria isolates could lower cholesterol, inhibit harmful enzyme activities, and control fecal water content.

MBNL3/CHCR prevents myogenic differentiation by inhibiting MyoD-dependent gene transcription.

Muscle differentiation is controlled by positive and negative signals. While much attention has been placed on proteins that promote muscle formation, the importance of negative regulators has been underemphasized. MBNL3/CHCR belongs to the muscleblind family of Cys3His zinc finger proteins implicated in myotonic dystrophy.