analysis-translation-developing-mouse-brain-using-polysome

Analysis of Translation in the Developing Mouse Brain using Polysome Profiling

The proper development of the mammalian brain relies on a fine balance of neural stem cell proliferation and differentiation into different neural cell ...

University of Calgary

The inflammatory bowel diseases (IBDs) are chronic inflammatory disorders with a complex etiology. IBD is thought to arise in genetically susceptible individuals in the context of aberrant interactions with the intestinal microbiota and other environmental risk factors. Recently, the pregnane X receptor (PXR) was identified as a sensor for microbial metabolites, whose activation can regulate the intestinal epithelial barrier. Mutations in NR1I2, the gene that encodes the PXR, have been linked to IBD, and in animal models, PXR deletion leads to barrier dysfunction. In the current study, we sought to assess the mechanism(s) through which the PXR regulates barrier function during inflammation. In Caco-2 intestinal epithelial cell monolayers, tumor necrosis factor-α/interferon-γ exposure disrupted the barrier and triggered zonula occludens-1 relocalization, increased expression of myosin light-chain kinase (MLCK), and activation of c-Jun N-terminal kinase 1/2 (JNK1/2). Activation of the PXR [rifaximin and [[3,5-Bis(1,1-dimethylethyl)-4-hydroxyphenyl]ethenylidene]bis-phosphonic acid tetraethyl ester (SR12813); 10 μM] protected the barrier, an effect that was associated with attenuated MLCK expression and JNK1/2 activation. In vivo, activation of the PXR [pregnenolone 16α-carbonitrile (PCN)] attenuated barrier disruption induced by toll-like receptor 4 activation in wild-type, but not Pxr-/-, mice. Furthermore, PCN treatment protected the barrier in the dextran-sulfate sodium model of experimental colitis, an effect that was associated with reduced expression of mucosal MLCK and phosphorylated JNK1/2. Together, our data suggest that the PXR regulates the intestinal epithelial barrier during inflammation by modulating cytokine-induced MLCK expression and JNK1/2 activation. Thus, targeting the PXR may prove beneficial for the treatment of inflammation-associated barrier disruption in the context of IBD.

Pregnane X Receptor Activation Attenuates Inflammation-Associated Intestinal Epithelial Barrier Dysfunction by Inhibiting Cytokine-Induced Myosin Light-Chain Kinase Expression and c-Jun N-Terminal Kinase 1/2 Activation.

The pathogenesis of the inflammatory bowel diseases (IBD), comprising Crohn's disease (CD) and ulcerative colitis (UC), involves aberrant interactions between a genetically susceptible individual, their microbiota and environmental factors. Alterations in xenobiotic receptor expression and function are associated with increased risk for IBD. Here, we have assessed the role of the constitutive androstane receptor (CAR), a xenobiotic receptor closely related to the pregnane X receptor, in the regulation of intestinal mucosal homeostasis.

Constitutive androstane receptor regulates the intestinal mucosal response to injury.

Clostridioides difficile (formerly Clostridium difficile; C difficile), the leading cause of nosocomial antibiotic-associated colitis and diarrhea in the industrialized world, triggers colonic disease through the release two toxins, toxin A (TcdA) and toxin B (TcdB), glucosyltransferases that modulate monomeric G-protein function and alter cytoskeletal function. The initial degree of the host immune response to C difficile and its pathogenic toxins is a common indicator of disease severity and infection recurrence. Thus, targeting the intestinal inflammatory response during infection could significantly decrease disease morbidity and mortality. In the current study, we sought to interrogate the influence of the pregnane X receptor (PXR), a modulator of xenobiotic and detoxification responses, which can sense and respond to microbial metabolites and modulates inflammatory activity, during exposure to TcdA and TcdB. Following intrarectal exposure to TcdA/B, PXR-deficient mice (Nr1i2 ) exhibited reduced survival, an effect that was associated with increased levels of innate immune cell influx. This exacerbated response was associated with a twofold increase in the expression of Tlr4. Furthermore, while broad-spectrum antibiotic treatment (to deplete the intestinal microbiota) did not alter the responses in Nr1i2 mice, blocking TLR4 signaling significantly reduced TcdA/B-induced disease severity and immune responses in these mice. Lastly, to assess the therapeutic potential of targeting the PXR, we activated the PXR with pregnenolone 16α-carbonitrile (PCN) in wild-type mice, which greatly reduced the severity of TcdA/B-induced damage and intestinal inflammation. Taken together, these data suggest that the PXR plays a role in the host's response to TcdA/B and may provide a novel target to dampen the inflammatory tissue damage in C difficile infections.

The xenobiotic sensing pregnane X receptor regulates tissue damage and inflammation triggered by C difficile toxins.

Gene regulation and metabolism are two fundamental processes that coordinate the self-renewal and differentiation of neural precursor cells (NPCs) in the developing mammalian brain. However, little is known about how metabolic signals instruct gene expression to control NPC homeostasis. Here, we show that methylglyoxal, a glycolytic intermediate metabolite, modulates Notch signalling to regulate NPC fate decision. We find that increased methylglyoxal suppresses the translation of Notch1 receptor mRNA in mouse and human NPCs, which is mediated by binding of the glycolytic enzyme GAPDH to an AU-rich region within Notch1 3'UTR. Interestingly, methylglyoxal inhibits the enzymatic activity of GAPDH and engages it as an RNA-binding protein to suppress Notch1 translation. Reducing GAPDH levels or restoring Notch signalling rescues methylglyoxal-induced NPC depletion and premature differentiation in the developing mouse cortex. Taken together, our data indicates that methylglyoxal couples the metabolic and translational control of Notch signalling to control NPC homeostasis.

Article	Year
Pregnane X Receptor Activation Attenuates Inflammation-Associated Intestinal Epithelial Barrier Dysfunction by Inhibiting Cytokine-Induced Myosin Light-Chain Kinase Expression and c-Jun N-Te... The Journal of pharmacology and experimental therapeutics\| PubMed ID: 27440420	2016
Constitutive androstane receptor regulates the intestinal mucosal response to injury. British journal of pharmacology\| PubMed ID: 28320072	2017
The xenobiotic sensing pregnane X receptor regulates tissue damage and inflammation triggered by C difficile toxins. FASEB journal : official publication of the Federation of American Societies for Experimental Biology\| PubMed ID: 31907988	2020
Methylglyoxal couples metabolic and translational control of Notch signalling in mammalian neural stem cells. Nature communications\| PubMed ID: 32332750	2020

Sarah L. Erickson

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