Reduction In Protein Phosphorylation Research Articles

The mechanism underlying pentabromoethylbenzene-induced adipogenesis and the obesogenic outcome in both cell and mouse model

Convergent evidence links traditional brominated flame retardants (BFRs) exposure to weight gain, while the obesogenic potency of new BFRs (NBFRs) remain largely unknown. Aiding by luciferase-reporter gene assay, the present study revealed only pentabromoethylbenzene (PBEB), an alternative for penta-BDEs, binds with retinoid X receptor α (RXRα) but not peroxisomeproliferator receptor γ (PPARγ) among the seven testing NBFRs. An apparent induction of adipogenesis in 3T3-L1 cells was observed at nanomolar of PBEB, much lower than penta-BFRs. Mechanistic research uncovered PBEB initiated the adipogenesis by demethylated CpG sites in the PPARγ promoter region. Specifically, activation RXRα by PBEB strengthened the activity of RXRα/PPARγ heterodimer, tightened the interaction between the heterodimer and PPAR response elements, and further enhanced adipogenesis. RNA sequencing combined with k-means clustering analysis exposed adenosine 5′-monophosphate (AMP)-activated protein kinase and phosphoinositide-3-kinase (PI3K)/protein kinase B (AKT) signaling as two predominant pathways that enriched in PBEB-induced lipogenesis. The obesogenic outcome was further corroborated in offspring mice when the maternal mice exposed to environmental relevant doses of PBEB. We found the male offspring exhibited adipocyte hypertrophy and increased weight gain in the epididymal white adipose tissue (eWAT). Consistent with in vitro findings, the reduction in protein phosphorylation of both AMPK and PI3K/AKT were observed within eWAT. Thus, we posited PBEB disrupts the pathways controlling adipogenesis and adipose tissue maintenance, supporting its potential as an environmental obesogen.

Glutamate receptor signaling is critical for T cell function and antitumor activity

Abstract The interaction between the nervous system and immune system has sparked interest in recent years. Emerging evidence shows an intricate neuroimmune network. We were intrigued by the expression of various neurotransmitter receptors on T lymphocytes. Specifically, following TCR stimulation, glutamate receptors (GluR) were significantly upregulated on both CD4+ and CD8+ T cells, with a peak at 48 h. Concomitant with the upregulation of activation molecules CD69, CD25 and CD44, proliferating CD8+ T cells presented higher levels of GluA3 and mGluR1 when compared with resting cells. By blocking group I metabotropic glutamate receptors and AMPA receptors through antagonists, we show that CD8+ T cells have a delayed activation but their viability is not altered. However, GluR blockade affected CD8+ T cell proliferation and their ability to kill tumor cells in vivo or target cells in vitro. While the frequency of GluR+CD8+ T cells ranges around 30% of total CD8+ T population, the effect of blocking glutamate signaling through receptor antagonists is a dominant effect and it is mediated by transient reduction in protein phosphorylation of TCR mediated pathways as well as calcium modulation. Overall, data suggest that glutamate receptors may have a stimulatory effect on T cell activation and glutamate agonists may boost T cell response in an immunosuppressive setting such as cancer.

Basal Protein Phosphorylation Is Decreased and Phosphatase Activity Increased by an Antioxidant and a Free Radical Trap in Primary Rat Glia

Reversible protein phosphorylation regulates a wide array of cellular functions. Cells respond to cytokines and various stressors via phosphorylation and thus activation of one or more of the mitogen-activated protein kinase (MAPK) pathways. Involvement of these signal transduction pathways has been implicated in numerous pathologies, including inflammation. Using a primary glia cell culture, we show here that the antioxidantN-acetylcysteine (NAC) and the nitrone-based free radical trap, α-phenyl-N-tert-butyl nitrone (PBN), reduce total basal protein phosphorylation in a concentration-dependent manner as assessed by phosphotyrosine analysis and by [γ-32P]ATP transfer radioassay. In addition we show that NAC inhibits H2O2-induced phosphatase inactivation in glia cell lysate. The PBN- and NAC-induced reduction in protein phosphorylation is accompanied by an increase in phosphatase activity, suggesting that PBN and NAC reduce protein phosphorylation by globally augmenting oxidant-sensitive phosphatase activities. These results partly explain why certain antioxidants also possess anti-inflammatory actions.

Purified Mammalian HSP-70 kDa Activates Phosphoprotein Phosphatases in Vitro

We have examined protein phosphorylation in the presence of purified mammalian HSP-70 kDa using the phosphoproteins in the rabbit reticulocyte lysate system as a model. Purified HSP-70 added to the rabbit reticulocyte lysate decreased the general proteinphosphorylation by 50-80% as measured by PAGE analysis of proteins labelled with γ-( 32P)-ATP. Reduction in protein phosphorylation was not due to the ATPase activity of HSP-70 as measured by thin layer chromatography. The reduction in protein phosphorylation was also not due to the reduced activities of the protein kinases. However, using ( 32P)-labelled phosphorylase-α as a substrate in the phosphatase assay system indicated increases in the activity of protein phosphatase 1 (PP-1)and/or 2A(PP-2A) by 20-40% relative to control in the presence of increasing concentrations of HSP-70. Using a variety of activators and inhibitors of the the two major protein phosphatases, PP-1 and PP-2A, we found that Mn 2+ caused a similar pattern of dephosphorylation of proteins as measured by PAGE analysis. Both okadaic acid and microcystin, two protein phosphatase inhibitors, largely counteracted the HSP-70 effect as measured by gel electrophoresis or when ( 32P)-labelled phosphorylase-α was used as a substrate.We conclude that in this system HSP-70 activates specific protein phosphatases.