Journal: Cell Host & Microbe

Article Title: Aspergillus fumigatus hijacks human p11 to redirect fungal-containing phagosomes to non-degradative pathway

doi: 10.1016/j.chom.2023.02.002

Figure Lengend Snippet:

Article Snippet: HCP216549-SG01-3 for generation of p11-KO cell line , GeneCopoiea , Cat# HCP216549-SG01-3.

Techniques: Virus, Recombinant, Cell Culture, Protease Inhibitor, Transfection, SYBR Green Assay, Mass Spectrometry, CyQUANT Assay, LDH Cytotoxicity Assay, cDNA Synthesis, Purification, Gene Expression, Construct, Control, Transformation Assay, Cloning, Plasmid Preparation, Expressing, Software, Imaging, Blocking Assay, Red Blood Cell Lysis, Membrane

Journal: eLife

Article Title: TIGAR deficiency enhances skeletal muscle thermogenesis by increasing neuromuscular junction cholinergic signaling

doi: 10.7554/eLife.73360

Figure Lengend Snippet: ( A ) Representative images of α-bungarotoxin immunofluorescent labeling of nicotinic acetylcholine receptor clusters in the extensor digitorium longus (EDL) muscle from wild type (WT) and whole-body Tigar knockout (TKO) mice. ( B ) Quantification of the number of nicotinic acetylcholine receptor clusters following 15 min exposure at 4°C (WT n = 17, TKO n = 13). These data represent the average of over six mice in each group of mean ± SD (unpaired t -test, two-tailed, **p=0.0035). ( C ) Acetylcholine levels in the gastrocnemius muscle of WT and TKO male (n = 7) mice at room temperature or following 1 hr at 4°C. ( D ) Acetylcholine levels in the gastrocnemius muscle of Chat Cre and chTKO male (n = 6) mice at room temperature or following 1 hr at 4°C. ( E ) Representative immunoblots of TIGAR and actin proteins from two scrambled sgRNA and two Tigar sgRNA knockout Sh-SY5Y cell lines. ( F ) Acetylcholine concentrations in the medium of scrambled and TKO SH-SH5Y neuroblastoma cells. ( G ) m2 acetylcholine enrichments in cells labeled with d -glucose-1,2- 13 C 2. ( H ) m2 acetylcholine enrichments in cells labeled with U- 13 C 6 d -glucose. ( I ) m1 glutamate ( m/z 128, C2-C4 fragment) enrichment in the medium of scrambled and TKO SH-SY5Y human neuroblastoma cells labeled with d -glucose-[1,2]- 13 C2 . Statistical analyses are described in ‘Method details,’ and the data are presented as the mean ± SD. *p<0.05, ***p<0.001, ****p<0.0001. Figure 6—source data 1. The culture SH-SY5Y cells were collected from both scrambled and whole-body Tigar knockout (TKO) cells, and 30 μg of the cell lysates were used for TIGAR (30 kDa) immunoblotting analysis as described in the ‘Immunoblotting’ section. The left four lanes of the raw image (lysate from neuroblastoma cells) were used for to confirm the efficiency of TIGAR protein loss in the SH-SY5Y neuroblastoma TKO cells. The right four lanes of the raw image represent the TIGAR immunoblotting of the cell lysates from 7-day differentiated neuroblastoma cells. A detailed description of the raw images is shown in . Figure 6—source data 2. The culture SH-SY5Y cells were collected from both scrambled and whole-body Tigar knockout (TKO) cells, and 30 μg of the cell lysates were used for actin β (42 kDa) immunoblotting analysis as described in the ‘Immunoblotting’ section. The left four lanes of the raw image (lysate from neuroblastoma cells) were used for to show actin β protein in the SH-SY5Y neuroblastoma cells. The right four lanes of the raw image represent the actin β immunoblotting of the cell lysates from 7-day differentiated neuroblastoma cells. A detailed description of the raw images is shown in .

Article Snippet: The CRISPR 3xsgRNA/Cas9 all-in-one expression clone targeting Tigar (NM_020375.2) (Cat# HCP215394-CG04-3) and scrambled sgRNA control for pCRISPR-CG04 (Cat# CCPCTR01-CG04-B) were purchased from GeneCopoeia (Rockville, MD).

Techniques: Labeling, Knock-Out, Two Tailed Test, Western Blot

Journal: eLife

Article Title: TIGAR deficiency enhances skeletal muscle thermogenesis by increasing neuromuscular junction cholinergic signaling

doi: 10.7554/eLife.73360

Figure Lengend Snippet:

Article Snippet: The CRISPR 3xsgRNA/Cas9 all-in-one expression clone targeting Tigar (NM_020375.2) (Cat# HCP215394-CG04-3) and scrambled sgRNA control for pCRISPR-CG04 (Cat# CCPCTR01-CG04-B) were purchased from GeneCopoeia (Rockville, MD).

Techniques: Knock-Out, Recombinant, Expressing, Control, Sequencing, Enzyme-linked Immunosorbent Assay, Plasmid Preparation, Lysis, Blocking Assay, Electron Microscopy, Software