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Image Search Results
Journal: Journal of Biological Chemistry
Article Title: BOK and NOXA Are Essential Mediators of p53-dependent Apoptosis
doi: 10.1074/jbc.m313526200
Figure Lengend Snippet: FIG. 5. mRNA expression profiles of proapoptotic BCL-2 family members. A, cell cultures were treated with etoposide for the indicated times. RT-PCR analysis of the abundance of mRNAs encoding specified members of the BCL-2 family in wild type, p53 DN-transfected, and MCF7 cells. Total cellular RNA was subjected to RT and PCR with gene-specific primers or glyceraldehyde-3-phosphate dehydrogenase (GAPDH) primers. PCR products were analyzed by agarose gel electrophoresis and staining with ethidium bromide. B, results of real time multiplex RT-PCR of selected mRNA demonstrate minor changes in BAX and PUMA expression but marked induction of BOK and NOXA genes. Data present mean values of three independent experiments. SH, SH-SY5Y.
Article Snippet: A polyclonal rabbit antibody to active caspase-3 was obtained from Cell Signaling Technology (New England Biolabs, 9661); a monoclonal mouse antibody to caspase-9 was from Medical and Biological Laboratories (MBL, clone 5B4); a monoclonal rat antibody to APAF-1 was from Alexis Biochemicals (804-348- C100); a monoclonal mouse antibody to -actin was from Sigma (A5441), polyclonal rabbit antibodies to BAK and BAX were from Upstate Biotechnology (06-536 and 06-499); polyclonal rabbit antibodies to procaspase-3, BCL-2, and BCL-xL were from Santa Cruz Biotechnology (sc-7148, sc-783, and sc-1690); a polyclonal rabbit antibody to BOK was from Abgent (AP1310a); a mouse monoclonal antibody to Cyto-c was from BD Biosciences (556433); a mouse monoclonal antibody to
Techniques: Expressing, Reverse Transcription Polymerase Chain Reaction, Transfection, Agarose Gel Electrophoresis, Staining, Multiplex Assay
Journal: Journal of Biological Chemistry
Article Title: BOK and NOXA Are Essential Mediators of p53-dependent Apoptosis
doi: 10.1074/jbc.m313526200
Figure Lengend Snippet: FIG. 6. BAX, BOK, and NOXA protein expression profiling. A, SH-SY5Y cells were treated with etoposide for the indicated times. Total protein extracts were probed with the antibodies against BAX, BOK, NOXA, and -actin. The release of Cyto-c was evaluated in cytosolic fractions and shown for comparison with BOK and NOXA expression. B, expression levels of BAK, BCL-xL, and -actin proteins were analyzed in total protein fractions from control and etoposide- treated SH-SY5Y cells.
Article Snippet: A polyclonal rabbit antibody to active caspase-3 was obtained from Cell Signaling Technology (New England Biolabs, 9661); a monoclonal mouse antibody to caspase-9 was from Medical and Biological Laboratories (MBL, clone 5B4); a monoclonal rat antibody to APAF-1 was from Alexis Biochemicals (804-348- C100); a monoclonal mouse antibody to -actin was from Sigma (A5441), polyclonal rabbit antibodies to BAK and BAX were from Upstate Biotechnology (06-536 and 06-499); polyclonal rabbit antibodies to procaspase-3, BCL-2, and BCL-xL were from Santa Cruz Biotechnology (sc-7148, sc-783, and sc-1690); a polyclonal rabbit antibody to BOK was from Abgent (AP1310a); a mouse monoclonal antibody to Cyto-c was from BD Biosciences (556433); a mouse monoclonal antibody to
Techniques: Expressing, Comparison, Control
Journal: Journal of Biological Chemistry
Article Title: BOK and NOXA Are Essential Mediators of p53-dependent Apoptosis
doi: 10.1074/jbc.m313526200
Figure Lengend Snippet: FIG. 7. BOK and NOXA are essential mediators of DNA damage-induced apoptosis. A, immunostaining for BAX, BOK, NOXA, and mitochondrial markers cytochrome. B, SH-SY5Y and MCF7 cells were treated with 50 M etoposide (ETOP) or 0.5 M staurosporine (STS) for 5 h. Cytosolic protein fractions were probed with the antibodies against BAX, Cyto-c, and -actin. C, MCF7 cells were transiently transfected with BAX, BOK, or NOXA expression plasmids or the empty pcDNA3.1 vector (Control). The release of Cyto-c was evaluated in cytosolic extracts 18 h after transfection. D, MCF7 cells were transiently co-transfected with the expression vectors encoding BAX, BOK, or NOXA and the empty pSilencer 3.1-H1 () or the pSilencer vector expressing the indicated siRNAs. Expression of the indicated proteins was analyzed in total protein extracts by immunoblotting 48 h after transfection. Immunostaining of -actin served as a loading control. E, SH-SY5Y cells were co-transfected with green fluorescent protein (GFP) expression vector and each of the indicated siRNA (si)-expressing constructs or the empty pSilencer 3.1-H1. Forty-eight hours after transfection cells were treated with etoposide for 18 h. Untreated green fluorescent protein cells served as a control. Cell viability was analyzed by counting viable green fluorescent cells. At least 100 cells were counted in each of three independent experiments. Data are expressed as a percentage of the value for etoposide-treated green fluorescent protein-transfected (control) cells S.D. *, p 0.05 damage-induced apoptosis. A, SH-SY5Y cells were treated with 50 M etoposide for 5 h; untreated cells served as a control.
Article Snippet: A polyclonal rabbit antibody to active caspase-3 was obtained from Cell Signaling Technology (New England Biolabs, 9661); a monoclonal mouse antibody to caspase-9 was from Medical and Biological Laboratories (MBL, clone 5B4); a monoclonal rat antibody to APAF-1 was from Alexis Biochemicals (804-348- C100); a monoclonal mouse antibody to -actin was from Sigma (A5441), polyclonal rabbit antibodies to BAK and BAX were from Upstate Biotechnology (06-536 and 06-499); polyclonal rabbit antibodies to procaspase-3, BCL-2, and BCL-xL were from Santa Cruz Biotechnology (sc-7148, sc-783, and sc-1690); a polyclonal rabbit antibody to BOK was from Abgent (AP1310a); a mouse monoclonal antibody to Cyto-c was from BD Biosciences (556433); a mouse monoclonal antibody to
Techniques: Immunostaining, Transfection, Expressing, Plasmid Preparation, Control, Western Blot, Construct
Journal: Cellular physiology and biochemistry : international journal of experimental cellular physiology, biochemistry, and pharmacology
Article Title: A New Perspective for Osteosarcoma Therapy: Proteasome Inhibition by MLN9708/2238 Successfully Induces Apoptosis and Cell Cycle Arrest and Attenuates the Invasion Ability of Osteosarcoma Cells in Vitro.
doi: 10.1159/000456598
Figure Lengend Snippet: Fig. 4. MLN2238 treatment increases the levels of pro-apoptotic proteins and down regulates the anti-apop totic proteins that control MOMP. A, MG-63 and Saos-2 cells were treated with MLN2238 at the IC50 doses in the 24h treatment, for the indicated times and then harvested. Whole-cell lysates were subjected to western blot analysis with BAX and BCL-2 antibodies. The data for the band intensity ratio of BAX/BCL-2 presented as means ± SD (n=3; *P< 0.05; **P< 0.01; ***P<0.001). Actin was monitored to control for equal loading. B, MG-63 and Saos-2 cells were treated for 24h at the indicated doses (+: 0.40µM for MG-63; +: 0.80µM for Saos-2). The whole-cell lysates were subjected to western blot analysis to detect BID, p-BAD, BAD (total), P53, NOXA and PUMA. *: nonspecific band. Tubulin was monitored to control for equal loading. The blots shown in the figure are representative of 3 independent experiments.
Article Snippet: Antibodies used were Caspase3/8/9 (9662/9746/9502), PARP (9542), MMP2/9 (4022/3852), APAF-1 (5088), BCL-2 (2872), Rb (9309), BID (2002) and COX IV (4844; all from Cell Signalling Technology; Beverly, MA, USA); P53 (ab1101), P21 (ab109199), BAX (ab32503), BAD (ab32445), p-BAD (Ser136, ab28824), P-CHK1 (Ser345, ab58567), Cytc (ab13575), Smac (ab32023) and OMI (ab75982; all from Abcam; Cambridge, UK); CDK1 (A0220) and p-CDK1 (Tyr15, AP0016; both from Abclonal; Woburn, MA, USA); WEE1 (sc-5285), E2F1 (sc137059), XIAP (sc-55550),
Techniques: Control, Western Blot