Journal: Journal of Virology

Article Title: Identification of a Feline Leukemia Virus Variant That Can Use THTR1, FLVCR1, and FLVCR2 for Infection

doi: 10.1128/jvi.02317-08

Figure Lengend Snippet: FIG. 2. LacZ-encoding virus bearing either FY981, FeLV-C, or FeLV-A Env was generated by cotransfection of HEK293 cells with MLV Gag-Pol, LacZ, and Env expression constructs. Supernatants were harvested, and infection titers were tested on a panel of cell lines. Titers are represented as the number of CFU per milliliter of virus supernatant, and values are averages of three independent infection studies. The cell lines tested are feline kidney H06T1, guinea pig 104C1, mink lung Mv1Lu, human cervical carcinoma HeLa, porcine testes ST-IOWA, and murine NIH 3T3. Arrows indicate zero infection titers. Standard deviation is shown for each infection.

Article Snippet: Human embryonic kidney cells, HEK293 (ATCC CRl-1573), and Phoenix ampho retroviral packaging cells (provided by Garry Nolan, Stanford University, Stanford, CA) were maintained in Dulbecco’s minimal essential medium with high glucose and 10% FBS.

Techniques: Virus, Generated, Cotransfection, Expressing, Construct, Infection, Standard Deviation

Journal: Cancers

Article Title: The YAP/TEAD Axis as a New Therapeutic Target in Osteosarcoma: Effect of Verteporfin and CA3 on Primary Tumor Growth

doi: 10.3390/cancers12123847

Figure Lengend Snippet: Role of TEAD in YAP-driven transcriptional activity. ( A ) Localization of endogenous YAP/TEAD1 complexes by in situ PLA in HOS cells. The red signal was obtained using Alexa555-labeled hybridization oligo nucleotides targeting amplified in situ PLA products. DAPI (blue) staining was used for nuclear visualization ( left panel ). Bars indicate means ± SD of three independent experiments (* p < 0.05, ** p < 0.01, right panel ). ( B HEK293 were transiently co-transfected with the YAP-S94A, YAP-S127A, TEAD1 or empty vector as indicated. 48 h after transfection lysates were subjected to immunoprecipitation (IP) with anti-Flag antibody followed by Western blotting (WB) by anti-Flag and anti-HA antibodies as indicated. ( C ) HOS cells were co-transfected with the TEAD-specific construct (TEAD)8-lux with or without empty, YAPS94A and YAPS127A expression vectors. Bars indicate means ± SD of four independent experiments, each performed in triplicate (** p < 0.01). ( D ) YAP mRNA steady-state levels were quantified by RT-q-PCR analysis in mock-, YAPS94A- and YAPS127A-transfected K-HOS cells. Bars indicate means ± SD of four independent experiments, each performed in duplicate (* p < 0.05, left panel ). YAP production was detected by Western blot analysis in mock-, YAPS94A- and YAPS127A-transfected K-HOS cells. Results shown are representative of two independent experiments ( right panel ). ( E ) Localization of YAP/TEAD1 complexes by in situ PLA experiments in mock-, YAPS94A- and YAPS127A-transfected K-HOS cells. The red signal was obtained using Alexa555-labeled hybridization oligo nucleotides targeting amplified in situ PLA products. DAPI (blue) staining was used for nuclear visualization ( left panel ). Bars indicate means ± S.D. of three independent experiments (** p < 0.01, right panel ). ( F ) Mock-, YAPS94A- and YAPS127A-transfected cells were transiently transfected with the TEAD-specific construct (TEAD)8-lux. Bars indicate means ± SD of four independent experiments, each performed in duplicate (* p < 0.05).

Article Snippet: To generate mutant YAP-S94A and YAP-S127A expressing cells, retrovirus infection was performed by transfecting 293 Phoenix retrovirus packaging cells with empty vector, pQCXIH-Myc-YAP-S94A and pQCXIH-Flag-YAP-S127A. pQCXIH-Myc-YAP-S94A and pQCXIH-Flag-YAP-S127A were gifts from Kunliang Guan (respectively Addgene plasmid #33094 and #33092; http://n2t.net/addgene:33094 and 33092; RRID:Addgene-33094 and -33092). pQCXIH CMV/TO DEST (w382-1) was a gift from Eric Campeau and Paul Kaufman (Addgene plasmid #17394; http://n2t.net/addgene:17394 ; RRID:Addgene-17394).

Techniques: Activity Assay, In Situ, Labeling, Hybridization, Amplification, Staining, Transfection, Plasmid Preparation, Immunoprecipitation, Western Blot, Construct, Expressing