Figure 1
Figure 1. K1 encodes a transmembrane protein with an immunoreceptor tyrosine-based activation motif (ITAM). (A) Schematic of K1 protein with the ITAM consensus amino acid sequence. The K1 gene encodes for a single transmembrane (TM) protein with a short cytoplasmic (CY) sequence that contains an ITAM. The mutant K1 gene (K1m) has a truncated ITAM. Above the bars are the amino acid numbers. The arrows below the bars indicate the locations and names of the oligonucleotide primers used for PCR. (B) K1myc expression in transfected HEK 293 cell extracts subjected to immunoprecipitation and immunoblotting with an anti-myc antibody. A unique band with an estimated size of approximately 50 kDa was present in lysates of HEK 293 cells transfected with pSG5K1myc. (C) PCR analysis of K1 expression in the BJAB cells stably transfected with pLXSN vectors expressing K1 (BJABK1) or K1m (BJABK1m) or with pLXSN alone (BJABXS) after selection of cells with G-418 antibiotic. The sizes of the bands are consistent with the predicted DNA sizes. (D) Expression of K1-encoded protein in BJAB cells. Cells were stained with the primary anti-K1 antibody 2H5 and a secondary anti–mouse IgG–FITC antibody. Images were visualized using an Olympus BX51 TF fluorescence microscope (Olympus Optical, Tokyo, Japan) equipped with a 40×/0.75 numerical aperture (NA) objective. Images were captured using an Olympus DP21 camera system. (E) Flow-cytometry analysis of the expression of K1 on the surface of BJAB transfectants. BJAB transfectants were stained with 2H5 and the anti–mouse IgG–FITC antibody. As a control, each experimental histogram is overlaid with mouse isotype IgG-stained BJAB transfectants. (F) HHV-8–infected cells show the presence of K1 in monomer and aggregate forms by immunoprecipitation-immunoblotting analysis. (G) Mice transfected with K1-Flag show the presence of K1 (arrowheads). Images were visualized using a Zeiss Axioskop 2 plus (Zeiss Microimaging, Thornwood, NY) equipped with a 20×/0.50 NA objective. Images were captured using a Zeiss MC 80 DX camera, and were scanned using an HP Scanjet 8200 (Hewlett-Packard, Palo Alto, CA). (H) Total RNA was isolated from transfected mouse liver and cells. RNA from equal numbers of BJAB cells, cells from the HBL6 cell line, primary human PEL cells, and mouse liver cells was subjected to RT-PCR. β-actin mRNA was amplified as an internal control for input mRNA.

K1 encodes a transmembrane protein with an immunoreceptor tyrosine-based activation motif (ITAM). (A) Schematic of K1 protein with the ITAM consensus amino acid sequence. The K1 gene encodes for a single transmembrane (TM) protein with a short cytoplasmic (CY) sequence that contains an ITAM. The mutant K1 gene (K1m) has a truncated ITAM. Above the bars are the amino acid numbers. The arrows below the bars indicate the locations and names of the oligonucleotide primers used for PCR. (B) K1myc expression in transfected HEK 293 cell extracts subjected to immunoprecipitation and immunoblotting with an anti-myc antibody. A unique band with an estimated size of approximately 50 kDa was present in lysates of HEK 293 cells transfected with pSG5K1myc. (C) PCR analysis of K1 expression in the BJAB cells stably transfected with pLXSN vectors expressing K1 (BJABK1) or K1m (BJABK1m) or with pLXSN alone (BJABXS) after selection of cells with G-418 antibiotic. The sizes of the bands are consistent with the predicted DNA sizes. (D) Expression of K1-encoded protein in BJAB cells. Cells were stained with the primary anti-K1 antibody 2H5 and a secondary anti–mouse IgG–FITC antibody. Images were visualized using an Olympus BX51 TF fluorescence microscope (Olympus Optical, Tokyo, Japan) equipped with a 40×/0.75 numerical aperture (NA) objective. Images were captured using an Olympus DP21 camera system. (E) Flow-cytometry analysis of the expression of K1 on the surface of BJAB transfectants. BJAB transfectants were stained with 2H5 and the anti–mouse IgG–FITC antibody. As a control, each experimental histogram is overlaid with mouse isotype IgG-stained BJAB transfectants. (F) HHV-8–infected cells show the presence of K1 in monomer and aggregate forms by immunoprecipitation-immunoblotting analysis. (G) Mice transfected with K1-Flag show the presence of K1 (arrowheads). Images were visualized using a Zeiss Axioskop 2 plus (Zeiss Microimaging, Thornwood, NY) equipped with a 20×/0.50 NA objective. Images were captured using a Zeiss MC 80 DX camera, and were scanned using an HP Scanjet 8200 (Hewlett-Packard, Palo Alto, CA). (H) Total RNA was isolated from transfected mouse liver and cells. RNA from equal numbers of BJAB cells, cells from the HBL6 cell line, primary human PEL cells, and mouse liver cells was subjected to RT-PCR. β-actin mRNA was amplified as an internal control for input mRNA.

Close Modal
This Feature Is Available To Subscribers Only

or Create an Account

Close Modal
Close Modal