Journal: Molecular Vision

Article Title: NF-κB feedback control of JNK1 activation modulates TRPV1-induced increases in IL-6 and IL-8 release by human corneal epithelial cells

doi:

Figure Lengend Snippet: Positive feedback control of JNK1 phosphorylation by NF-κB through DUSP1. A : Time-dependent increases in p- IκBα. Scrambled shRNA subline was exposed to CAP (20 µM) for up to 90 min. Western blots reveal the time course of changes in p-IκBα formation, which serves as a readout of NF-κB activation. B : Contribution by JNK1 to IκBα phosphorylation. Western blots compare CAP (20 µM)-induced IκBα phosphorylation in scrambled shRNA and JNK1 sublines at 60 min. Preincubation with either 5z-OX (0.1 µM), CPZ (10 µM) or PDTC (50 µM) for 60 min suppressed CAP-induced IκBα phosphorylation. C : Positive feedback control by NF-κB of JNK1/2 activation. Loss of NF-κB activation reduces transient JNK1/2, p38, and ERK1/2 MAPK activation induced by CAP (20 µM) for up to 90 min. Summary plots contrast time-dependent patterns of MAPK activation in the scrambled shRNA subline (left) with those in NF-κB1 subline (right). D : Inverse relationship between changes in PKCδ and DUSP1 expression. Scrambled shRNA and NF-κB1 sublines were exposed to CAP (20 µM) as described in B . Summary plot (left) indicates that in the scrambled shRNA subline CAP-induced increases in PKCδ expression whereas DUSP1 remained invariant (left). Summary plot (right) reveals inverse responses by PKCδ and DUSP1 to CAP in NF-κB1 subline.

Article Snippet: The anti- DUSP1 antibody was obtained from ABNOVA (Walnut Creek, CA).

Techniques: Control, Phospho-proteomics, shRNA, Western Blot, Activation Assay, Expressing

Journal: Molecular Vision

Article Title: NF-κB feedback control of JNK1 activation modulates TRPV1-induced increases in IL-6 and IL-8 release by human corneal epithelial cells

doi:

Figure Lengend Snippet: Changes in MAPK activation patterns and IL-6/8 release in DUSP1 subline. A : Western blot analysis of total DUSP1 protein expression in resting scrambled HCEC and DUSP1 subline. B : Comparison of time dependent changes in MAPK phosphorylation in scrambled shRNA and DUSP1 subline. Cells were exposed to CAP (20 µM) for indicated times. Changes are compared in p-ERK1/2, p-JNK1/2, and p38 MAPK in two different sublines. Protein loading equivalence validated based on invariant ERK1/2 expression levels. C : DUSP1 gene knockdown enhances CAP-induced IL-6 and IL-8 release. ELISA was performed on scrambled shRNA and DUSP1 subline after 24 h exposure to CAP (20 µM). Three independent experiments each performed in triplicate.

Article Snippet: The anti- DUSP1 antibody was obtained from ABNOVA (Walnut Creek, CA).

Techniques: Activation Assay, Western Blot, Expressing, Comparison, Phospho-proteomics, shRNA, Knockdown, Enzyme-linked Immunosorbent Assay

Journal: Molecular Vision

Article Title: NF-κB feedback control of JNK1 activation modulates TRPV1-induced increases in IL-6 and IL-8 release by human corneal epithelial cells

doi:

Figure Lengend Snippet: TRPV1-linked signaling pathways mediating IL-6 and IL-8 release. TRPV1 stimulation leads to TAK1-dependent JNK1 and NF-κB activation. Activated JNK1 potentiates NF-κB activation by TAK1. NF-κB provides a positive feedback control of JNK1 phosphorylation through inhibition of DUSP1 expression, which is possibly controlled by PKCδ. Activated NF-κB translocates to nucleus, along with other transcription factors activated by p-JNK1 (e.g., AP-1) to promote IL-6 and IL-8 mRNA expression. Solid line indicates established interaction. Broken line represents unproven interaction. Arrowhead means stimulation whereas hammerhead represents inhibition. Arrow pointing from TAK1 to NF-κB is broken since it is not yet known if interaction is direct or there are signaling intermediates mediating NF-κB activation by TAK1.

Article Snippet: The anti- DUSP1 antibody was obtained from ABNOVA (Walnut Creek, CA).

Techniques: Protein-Protein interactions, Activation Assay, Control, Phospho-proteomics, Inhibition, Expressing