Journal: Proceedings of the National Academy of Sciences of the United States of America

Article Title: Endothelial deletion of EPH receptor A4 alters single-cell profile and Tie2/Akap12 signaling to preserve blood–brain barrier integrity

doi: 10.1073/pnas.2204700120

Figure Lengend Snippet: Raw average gene expression values from all capillary-derived endothelial cells

Article Snippet: Rabbit anti-EphA4 (Proteintech), goat anti-Cd31 (BD Biosciences), rabbit anti-CD45 (Cell Signaling Technology), rat anti-CD68 (Invitrogen), rabbit anti-Akap12 (Alomone, Jerusalem), and goat anti-Tie2 (R&D Systems).

Techniques: Expressing, Activation Assay, Shear, Membrane, Control

Journal: Journal of immunology (Baltimore, Md. : 1950)

Article Title: Systemic lupus erythematosus immune complexes increase the expression of SLAM family members CD319 (CRACC) and CD229 (LY-9) on plasmacytoid dendritic cells and CD319 on CD56(dim) NK cells.

doi: 10.4049/jimmunol.1301022

Figure Lengend Snippet: FIGURE 5. pDCs lack the activating adaptor molecules but express inhibitory adaptor molecules of SLAM receptors. (A) IFN-a production by RNA-IC–stimulated pDCs after cross-linking of CD319, CD229, or BDCA-2 by plate-bound mAbs. IFN-a levels were determined in 20-h cell culture supernatants by an immunoassay and normalized to the IFN-a production in cultures without plate-bound mAb. The dashed line indicates 100%. Data (mean + SD) are from three donors from two independent experiments. Expression of EAT2 and SAP (B) and SHIP-1, SHP-1, SHP- 2, and CSK (C) in cell lysates from isolated pDCs or NK cells from two healthy donors was determined by Western blot. Expression of b-actin was used as control.

Article Snippet: Rabbit polyclonal Abs to Ewing’s sarcoma-activated transcript 2 (EAT2), SHIP-1, SHP-2, CSK (Proteintech Group, Chicago, IL), and a rabbit mAb to SHP-1 (EPR5519; Epitomics, Burlingame, CA), followed by HRP-conjugated goat anti-rabbit IgG (H+L) (Invitrogen), were used to detect the indicated signaling molecules.

Techniques: Cell Culture, Expressing, Isolation, Western Blot, Control

Journal: Cell Death & Disease

Article Title: A GSTP1-mediated lactic acid signaling promotes tumorigenesis through the PPP oxidative branch

doi: 10.1038/s41419-023-05998-4

Figure Lengend Snippet: A Flag-tagged GSTP1 was overexpressed in cell lines (MCF-7, Bcap37, and 293 T) for 24 h. Cells were then incubated in glc medium (0.5 mM glucose) or lac medium (0.5 mM glucose with 20 mM lactic acid) for 12 h, and Flag-tagged GSTP1 protein immuno-purified for interaction detection. B MCF-7 cells/HeLa cells/HepG2 cells were incubated as in ( A ) for 12 h. Lysates were immuno-precipitated with anti-G6PD antibodies for endogenous interactions with GSTP1 and SRC. C Relative abundances of GSTP1 and SRC were obtained from 3 independent replicate experiments and normalized to immunoprecipitated G6PD endogenous protein in ( B ). * p < 0.05, ** p < 0.01. D GST-SRC pull down to detect the GSTP1-G6PD-SRC interaction with or without ATP in vitro, as stained by CBB. E GST-SRC pull down to detect the GSTP1-G6PD-SRC interaction with or without lactic acid in vitro. F A modeled spatial arrangement of the GSTP1-SRC-G6PD complex. Green: GSTP1 (PDB ID: 3GUS); blue: SRC (PDB ID: 2H8H); Orange: G6PD(PDB ID: 2bhl). (G/H) Knockdown of GSTP1 decreases the interaction between G6PD and SRC ( G ) and overall G6PD phosphorylation level ( H ). I , J The interactions between GSTP1 (SRC) and G6PD decreased in G6PD mutants (2YF/2YA) in cells. HA-SRC ( I ), HA-GSTP1 ( J ), or HA-tagged empty control plasmids were co-transfected with WT (G6PD) or mutants (2YF/2YA) constructs for 24 h, and Flag-tagged G6PD was immunopurified and immunoblotted with indicated antibodies. K , L Modeled spatial arrangement of the GSTP1-SRC ( L ) or GSTP1-G6PD ( K ) complex. Orange: G6PD; Green: SRC; Pink: the 1-70AA region of GSTP1; Yellow: the 141-210AA region of GSTP1. M GFP, GFP-fused full length or indicated domains of GSTP1 were overexpressed in MCF-7 cells and immunoprecipitated by anti-GFP beads and examined through Western blots to check the interaction among G6PD, GSTP1 and SRC. N Proteins as indicated in ( M ) were overexpressed in MCF-7 cells, and cells treated with glucose-deprived (0.5 mM) media without or with 20 mM lactic acid to check the dynamic change of the tri-complex formation through Western Blots.

Article Snippet: GST tagged SRC protein was purchased from Sino Biological.

Techniques: Incubation, Purification, Immunoprecipitation, In Vitro, Staining, Transfection, Construct, Western Blot