Journal: Journal of Biological Chemistry

Article Title: Chaperone Insufficiency Links TLR4 Protein Signaling to Endoplasmic Reticulum Stress

doi: 10.1074/jbc.m111.315218

Figure Lengend Snippet: FIGURE 6. GRP94 and GRP78 control ERS in THP-1 cells. In A–C, THP-1 cells were maintained in steady state (CTR) or treated with LPS (24 h) or submitted to glucose deprivation without LPS treatment (GLU-R) or submitted to glucose deprivation followed by LPS treatment (GLU-R24 h LPS). Protein extracts were separated by SDS-PAGE and immunoblotted with anti-spliced XBP1 (A), peIF2 (B), or ATF6 (C) antibodies. In D and E, THP-1 cells were treated with Lipo- fectaminealone(Lipo)orinthepresenceofglucosedeprivation(LipoGlu-R)orwithacontrolsiRNA(siCTR),aGRP94(D),oraGRP78(E)siRNA(siGRP94orsiGRP78, respectively), or a GRP94 (D) or a GRP78 (E) siRNA in the presence of glucose deprivation (siGRP94Glu-R or siGRP78Glu-R, respectively). Protein extracts were separated by SDS-PAGE and immunoblotted with anti-GRP94 (D) or GRP78 (E) antibodies. In F–L, THP-1 cells were treated with Lipofectamine alone (Lipo) or in the presence of LPS, glucose deprivation (LipoGlu-R), glucose deprivation plus LPS (GLU-RLPS) in the absence of presence of a GRP94 (F–H and L), or a GRP78 (I–K) siRNA (siGRP94 or siGRP78, respectively). Protein extracts were separated by SDS-PAGE and immunoblotted with anti-spliced XBP1 (F and I), pPERK (G and J), or ATF6 (H and K) antibodies or real-time PCR was used to determine the expression of IL-8 (L). In A–K, all membranes were stripped and reblotted with anti--actin. In all experiments, n 6. *, p 0.05 versus CTR in A–C or versus Lipofectamine alone in D–L. In A and B, #, p 0.05 versus 24-h LPS. asu, arbitrary scanning units.

Article Snippet: In some experiments, cells were deprived of glucose for 6 h or treated with siRNA (GRP94 siRNA, sc35523 and GRP78 siRNA, sc29338 from Santa Cruz Biotechnology) to increase or decrease, respectively, the expression of the chaperones.

Techniques: Control, SDS Page, Real-time Polymerase Chain Reaction, Expressing

Journal: Molecular Metabolism

Article Title: TALK-1 reduces delta-cell endoplasmic reticulum and cytoplasmic calcium levels limiting somatostatin secretion

doi: 10.1016/j.molmet.2018.01.016

Figure Lengend Snippet: TALK-1 channels are expressed in mouse and human δ-cells. (A) Mouse pancreas section stained for TALK-1 (green) and somatostatin (red) (representative of N = 3 mice). (B) Mouse pancreas section stained for TALK-1 (green), ER (GRP94, red), and SST (cyan). (C) Human pancreas section stained for TALK-1 (green) and somatostatin (red) (representative of N = 3 pancreata). (D) Human pancreas section stained for TALK-1 (green), ER (GRP94, red), and somatostatin (E) K2P currents recorded from WT and TALK-1 KO δ-cells ( N = 3 mice per genotype). (F) K2P currents recorded from human δ-cells expressing TALK-1 DN or control mCherry. ( N = 3 islet preparations); * P < 0.05, ** P < 0.005.

Article Snippet: Sections were stained using primary antibodies against somatostatin (Santa Cruz Biotechnology sc-7819: 1:250), TALK-1 (Novus Biologicals #NBP1-83071; 1:175) or TALK-1a (Antibody Verify AAS72353C; 1:250), glucagon (Proteintech #15954-I-AP: 1:500), and the endoplasmic reticulum marker GRP94 (Novus Biologicals #NB300-619; 1:100); secondary antibodies used were Alexa Fluor 488-conjugated donkey anti-rabbit (Jackson Immunoresearch #711-546-152; 1:300), DyLight 650-conjugated donkey anti-goat (Thermo Fisher #SA5-10089; 1:250), and Cy3-conjugated donkey anti-mouse (Jackson Immunoresearch #715-166-150; 1:500).

Techniques: Staining, Expressing, Control