An integrated approach identifies the molecular underpinnings of murine anterior visceral endoderm migration.

The anterior visceral endoderm (AVE) differs from the surrounding visceral endoderm (VE) in its migratory behavior and ability to restrict primitive streak formation to the opposite side of the mouse embryo. To characterize the molecular bases for the unique properties of the AVE, we combined single-cell RNA sequencing of the VE prior to and during AVE migration with phosphoproteomics, high-resolution live-imaging, and short-term lineage labeling and intervention. This identified the transient nature of the AVE with attenuation of "anteriorizing" gene expression as cells migrate and the emergence of heterogeneities in transcriptional states relative to the AVE's position.

A small-molecule PI3Kα activator for cardioprotection and neuroregeneration.

Harnessing the potential beneficial effects of kinase signalling through the generation of direct kinase activators remains an underexplored area of drug development. This also applies to the PI3K signalling pathway, which has been extensively targeted by inhibitors for conditions with PI3K overactivation, such as cancer and immune dysregulation. Here we report the discovery of UCL-TRO-1938 (referred to as 1938 hereon), a small-molecule activator of the PI3Kα isoform, a crucial effector of growth factor signalling.

Transient Inhibition of PI3Kδ Enhances the Therapeutic Effect of Intravenous Delivery of Oncolytic Vaccinia Virus.

Tumor-targeting oncolytic viruses such as vaccinia virus (VV) are attractive cancer therapeutic agents that act through multiple mechanisms to provoke both tumor lysis and anti-tumor immune responses. However, delivery of these agents remains restricted to intra-tumoral administration, which prevents effective targeting of inaccessible and disseminated tumor cells. In the present study we have identified transient pharmacological inhibition of the leukocyte-enriched phosphoinositide 3-kinase δ (PI3Kδ) as a novel mechanism to potentiate intravenous delivery of oncolytic VV to tumors.

Vps34 PI 3-kinase inactivation enhances insulin sensitivity through reprogramming of mitochondrial metabolism.

Vps34 PI3K is thought to be the main producer of phosphatidylinositol-3-monophosphate, a lipid that controls intracellular vesicular trafficking. The organismal impact of systemic inhibition of Vps34 kinase activity is not completely understood. Here we show that heterozygous Vps34 kinase-dead mice are healthy and display a robustly enhanced insulin sensitivity and glucose tolerance, phenotypes mimicked by a selective Vps34 inhibitor in wild-type mice.

Molecules in medicine mini-review: isoforms of PI3K in biology and disease.

The PI3K lipid kinases are involved in signal transduction and intracellular vesicular traffic, endowing these enzymes with multiple cellular functions and important roles in normal physiology and disease. In this mini-review, we aim to distill from the vast PI3K literature the key relevant concepts for successful targeting of this pathway in disease. Of the eight isoforms of PI3K, the class I PI3Ks have been implicated in the aetiology and maintenance of various diseases, most prominently cancer, overgrowth syndromes, inflammation and autoimmunity, with emerging potential roles in metabolic and cardiovascular disorders.

Novel Role for p110β PI 3-Kinase in Male Fertility through Regulation of Androgen Receptor Activity in Sertoli Cells.

The organismal roles of the ubiquitously expressed class I PI3K isoform p110β remain largely unknown. Using a new kinase-dead knockin mouse model that mimics constitutive pharmacological inactivation of p110β, we document that full inactivation of p110β leads to embryonic lethality in a substantial fraction of mice. Interestingly, the homozygous p110β kinase-dead mice that survive into adulthood (maximum ~26% on a mixed genetic background) have no apparent phenotypes, other than subfertility in females and complete infertility in males.

Essential role of class II PI3K-C2α in platelet membrane morphology.

The physiologic roles of the class II phosphoinositide 3-kinases (PI3Ks) and their contributions to phosphatidylinositol 3-monophosphate (PI3P) and PI(3,4)P2 production remain elusive. Here we report that mice heterozygous for a constitutively kinase-dead PI3K-C2α display aberrant platelet morphology with an elevated number of barbell-shaped proplatelets, a recently discovered intermediate stage in the final process of platelet production. Platelets with heterozygous PI3K-C2α inactivation have critical defects in α-granules and membrane structure that are associated with modifications in megakaryocytes.

The p110δ isoform of the kinase PI(3)K controls the subcellular compartmentalization of TLR4 signaling and protects from endotoxic shock.

Lipopolysaccharide activates plasma-membrane signaling and endosomal signaling by Toll-like receptor 4 (TLR4) through the TIRAP-MyD88 and TRAM-TRIF adaptor complexes, respectively, but it is unclear how the signaling switch between these cell compartments is coordinated. In dendritic cells, we found that the p110δ isoform of phosphatidylinositol-3-OH kinase (PI(3)K) induced internalization of TLR4 and dissociation of TIRAP from the plasma membrane, followed by calpain-mediated degradation of TIRAP. Accordingly, inactivation of p110δ prolonged TIRAP-mediated signaling from the plasma membrane, which augmented proinflammatory cytokine production while decreasing TRAM-dependent endosomal signaling that generated anti-inflammatory cytokines (interleukin 10 and interferon-β).

Isoform-selective induction of human p110δ PI3K expression by TNFα: identification of a new and inducible PIK3CD promoter.

PI3Ks (phosphoinositide 3-kinases) are signalling molecules and drug targets with important biological functions, yet the regulation of PI3K gene expression is poorly understood. Key PI3Ks are the class IA PI3Ks that consist of a catalytic subunit (p110α, p110β and p110δ) in complex with a p85 regulatory subunit. Whereas p110α and p110β are ubiquitously expressed, high levels of p110δ are mainly found in white blood cells, with most non-leucocytes expressing low levels of p110δ.

Apparent tolerance of turkey vultures (Cathartes aura) to the non-steroidal anti-inflammatory drug diclofenac.

The nonsteroidal anti-inflammatory drug diclofenac is extremely toxic to Old World Gyps vultures (median lethal dose -0.1-0.2 mg/kg), evoking visceral gout, renal necrosis, and mortality within a few days of exposure.

Small carboxyl-terminal domain phosphatase 2 attenuates androgen-dependent transcription.

Small carboxyl-terminal domain (CTD) phosphatase 2 (SCP2) was identified and verified as a protein that interacts with the androgen receptor (AR). Ectopic expression of SCP2 or two other family members, SCP1 and SCP3, attenuated AR transcriptional activity in LNCaP cells and were recruited in an androgen- and AR-dependent fashion onto the prostate-specific antigen (PSA) promoter. Silencing SCP2 and SCP1 by short hairpin RNAs increased androgen-dependent transcription of the PSA gene and augmented AR loading onto the PSA promoter and enhancer.

Expression and purification of the mitochondrial serine protease LACTB as an N-terminal GST fusion protein in Escherichia coli.

LACTB is a mammalian mitochondrial protein sharing sequence similarity to the beta-lactamase/penicillin-binding protein family of serine proteases that are involved in bacterial cell wall metabolism. The physiological role of LACTB is unclear. In this study we have subcloned the cDNA of mouse LACTB (mLACTB) and produced recombinant mLACTB protein in Escherichia coli.