Journal: Journal of Virology

Article Title: Specific Akt Family Members Impair Stress-Mediated Transactivation of Viral Promoters and Enhance Neuronal Differentiation: Important Functions for Maintaining Latency

doi: 10.1128/jvi.00901-20

Figure Lengend Snippet: Figure 4. Akt3 efficiently promotes neurite formation in Neuro-2A cells. 730

Article Snippet: A plasmid that expresses Akt3 was a gift from William Sellers (1236 320 pcDNA3 Myr HA Akt3; plasmid 9017; Addgene).

Techniques:

Journal: Journal of Ovarian Research

Article Title: MYB/AKT3 axis is a key driver of ovarian cancer growth, aggressiveness, and chemoresistance

doi: 10.1186/s13048-025-01761-9

Figure Lengend Snippet: AKT3 is a direct transcriptional target of MYB in ovarian cancer cells. A . AKT3 mRNA expression was analyzed in a panel of MYB-modulated OC cell lines by qPCR. Actin Ct was used to normalize the data. B . In silico analysis of AKT3 promoter was performed using PROMO- ALGGEN to identify putative MYB binding sites (MBS). Seven putative MBS (P1-P7) were identified. C . Chromatin immunoprecipitation was performed using anti-MYB antibody or IgG control, followed by qPCR analysis using site-specific primers in SKOV3 and SKOV3-ip cells. The highest binding was recorded at the P4 site. The direct MYB binding in the P4 region of the AKT3 promoter increased in MYB-overexpressing cells and decreased in MYB-silenced OC cells. D . MYB-modulated cells were also subjected to ChIP-qPCR analysis, and enhanced pull-down of the P4 MYB-binding sequence in the AKT3 promoter was observed in MYB-overexpressing SKOV3 cells, while it was reduced in MYB-silenced SKOV3-ip cells compared to their respective control cells

Article Snippet: Primary antibodies used were anti-MYB (cat #12319), AKT3 (cat #14982), pAKT (#4060), E-cadherin (cat #3195), N-Cadherin (cat #13116), anti-GSK3β (cat #9315), -pGSK3β (cat #5558), normal Rabbit IgG (cat #2729), all obtained from (Cell Signaling Technology, Beverly, MA); Rabbit IgG peroxidase-conjugated secondary antibody (cat #HAF008) and Mouse IgG horseradish peroxidase-conjugated antibody (cat # HAF007) (R&D Systems, Inc., Minneapolis, MN, USA).

Techniques: Expressing, In Silico, Binding Assay, Chromatin Immunoprecipitation, Control, ChIP-qPCR, Sequencing

Journal: Journal of Ovarian Research

Article Title: MYB/AKT3 axis is a key driver of ovarian cancer growth, aggressiveness, and chemoresistance

doi: 10.1186/s13048-025-01761-9

Figure Lengend Snippet: PI3K/AKT signaling is induced in MYB-overexpressing ovarian cancer cells. A & B . MYB-regulated AKT3 expression was confirmed in OC cell lines with modulated MYB expression by immunoblotting, which correlated with relative increase or decrease in the phosphorylation of AKT and its downstream target, GSK3β. β-actin was used as a loading control. C . KEGG pathway analysis using the DEGs identified PI3K/AKT as the most activated pathway in MYB-overexpressing cells. The x-axis represents the significance of fold enrichment while the y-axis shows the pathway categories

Article Snippet: Primary antibodies used were anti-MYB (cat #12319), AKT3 (cat #14982), pAKT (#4060), E-cadherin (cat #3195), N-Cadherin (cat #13116), anti-GSK3β (cat #9315), -pGSK3β (cat #5558), normal Rabbit IgG (cat #2729), all obtained from (Cell Signaling Technology, Beverly, MA); Rabbit IgG peroxidase-conjugated secondary antibody (cat #HAF008) and Mouse IgG horseradish peroxidase-conjugated antibody (cat # HAF007) (R&D Systems, Inc., Minneapolis, MN, USA).

Techniques: Expressing, Western Blot, Phospho-proteomics, Control

Journal: Journal of Ovarian Research

Article Title: MYB/AKT3 axis is a key driver of ovarian cancer growth, aggressiveness, and chemoresistance

doi: 10.1186/s13048-025-01761-9

Figure Lengend Snippet: AKT3 mediates the actions of MYB on ovarian cancer cell growth, survival, and chemoresistance. A . OC cells with forced (SKOV3-MYB) or endogenously high expression of MYB (SKOV3-ip) were treated with 50 nM of AKT3 siRNA for 48 h as described in the materials and methods. Subsequently, cell lysates were prepared and analyzed by immunoblotting. B & C . OC cell lines were seeded in 96-well culture plates and treated with 50 nM of AKT3 siRNA. After 72 h, cells were subjected to WST assay for assessment of cell growth. Bars represent the relative differences in the growth, * p < 0.05. D & E . SKOV3 and SKOV3-ip cells were transfected with either AKT3 siRNA (50nM) or scrambled siRNA (NT-Scr) and incubated for 24 h. After 24 h, the cells were washed and cultured for 72 h in serum-deprived media. Following this, LIVE/DEAD kit reagents (Ethidium homodimer and Calcein-AM) were added to the culture, which were then incubated for 20 min, rinsed 3 times with PBS. Live cells display green fluorescence while dead cells fluoresce Red/orange. The graph depicts the count of dead cells per well. (* p < 0.05). F & G . OC cells seeded in 96-well plates and transfected with 50 nM of AKT3 or scrambled siRNA were incubated for 24 h. Following that, these cells were treated with different concentrations of cisplatin for 72 h. After 72 h, percent viability was measured by WST-1 assay, and IC50 values were obtained in GraphPad Prism version 10 (GraphPad Software, San Diego, CA, USA), through the equation: log(inhibitor) vs. normalized response, using a Variable slope model. Effect of MYB modulation on the migration and invasion of OC cells. H & I . SKOV3-MYB and SKOV3-ip cells were transfected with 50 nM of either non-targeting scrambled siRNA (NT-Scr) or AKT3 siRNA for 48 h and subsequently were plated in the non-coated (for migration) or Matrigel-coated (for invasion) transwell chambers and allowed to migrate for 16 h under a chemotactic drive. Cells that had migrated through the membrane were fixed, stained, and counted in 10 random view fields. Bars represent the mean ± standard deviation. * p < 0.05

Article Snippet: Primary antibodies used were anti-MYB (cat #12319), AKT3 (cat #14982), pAKT (#4060), E-cadherin (cat #3195), N-Cadherin (cat #13116), anti-GSK3β (cat #9315), -pGSK3β (cat #5558), normal Rabbit IgG (cat #2729), all obtained from (Cell Signaling Technology, Beverly, MA); Rabbit IgG peroxidase-conjugated secondary antibody (cat #HAF008) and Mouse IgG horseradish peroxidase-conjugated antibody (cat # HAF007) (R&D Systems, Inc., Minneapolis, MN, USA).

Techniques: Expressing, Western Blot, WST Assay, Transfection, Incubation, Cell Culture, Fluorescence, WST-1 Assay, Software, Migration, Membrane, Staining, Standard Deviation

Journal: Oncotarget

Article Title: SIRT1 at the crossroads of AKT1 and ERβ in malignant pleural mesothelioma cells

doi: 10.18632/oncotarget.7321

Figure Lengend Snippet: A. Kaplan-Meier survival curves of malignant pleural mesothelioma patients stratified for AKT1 and B. AKT3 high or low expression levels.

Article Snippet: Anti-AKT1 and AKT3 monoclonal antibodies were from Rockland Immunochemicals Inc. (Gilbertsville, PA, USA).

Techniques: Expressing

Journal: Oncotarget

Article Title: SIRT1 at the crossroads of AKT1 and ERβ in malignant pleural mesothelioma cells

doi: 10.18632/oncotarget.7321

Figure Lengend Snippet: A. Phase contrast images (200X magnification) of MSTO-211H cells transfected with non-specific control siRNA (NS siRNA) or AKT1 siRNA (siRNA AKT1), AKT3 siRNA (siRNA AKT3) or both specific siRNAs (siRNA AKT1/AKT3). B. Representative RT-PCR analyses and relative densitometry of AKT1 , AKT3 , and CDH1 in AKT1 and AKT3 silenced MSTO-211H cells compared to their controls. 18S rRNA was used as housekeeping gene. C. Phase contrast images (200X magnification) of MSTO-211H cells transfected with non-specific control siRNA or AKT isoform specific siRNAs grown on Matrigel coated dishes for 24 hours. D. Soft agar colony counts in non-specific control siRNA or AKT isoform specific siRNAs transfected MSTO-211H cells. Columns represent the percentage of the mean number of colonies versus control ± s.d.; * p≤0.05.

Article Snippet: Anti-AKT1 and AKT3 monoclonal antibodies were from Rockland Immunochemicals Inc. (Gilbertsville, PA, USA).

Techniques: Transfection, Control, Reverse Transcription Polymerase Chain Reaction

Journal: Oncotarget

Article Title: SIRT1 at the crossroads of AKT1 and ERβ in malignant pleural mesothelioma cells

doi: 10.18632/oncotarget.7321

Figure Lengend Snippet: A. Representative RT-PCR analyses of AKT1 , AKT3 and SIRT1 in MSTO-211H cells transfected with non-specific control siRNA (NS siRNA), and siRNAs of AKT1 (siRNA AKT1) or AKT3 (siRNA AKT3). B. MSTO-211H cells treated for 24 hours ± 8 nM MK2206. C. Mock or AKT1-HA transfected REN cells. 18S rRNA was used as housekeeping gene. D. Real time and representative Western blot analyses of ERβ expression in MSTO-211H cells transfected with non-specific control siRNA (NS siRNA) or SIRT1 siRNA (siRNA SIRT1). Tubulin was used as loading control. E. Representative RT-PCR analyses of AKT1, AKT3 and FOXM1 in MSTO-211H cells transfected with non-specific control siRNA (NS siRNA), and siRNAs of AKT1 (siRNA AKT1) or AKT3 (siRNA AKT3) or treated for 24 hours ± 8 nM MK2206. F. Representative RT-PCR analyses of AKT1 and FOXM1 in Mock, AKT1-HA and AKTΔN transfected REN cells. 18S rRNA was used as housekeeping gene. G. Representative RT-PCR analyses of SIRT1 , FOXM1 and AKT1 in MSTO-211H cells transfected with non-specific control siRNA (NS siRNA) or SIRT1 siRNA (siRNA SIRT1) or FOXM1 siRNA (siRNA FOXM1). 18S rRNA was used as housekeeping gene. H. Real time and representative Western blot analyses of ERβ expression in MSTO-211H cells transfected with non-specific control siRNA (NS siRNA) or FOXM1 siRNA (siRNA FOXM1). Tubulin was used as loading control.

Article Snippet: Anti-AKT1 and AKT3 monoclonal antibodies were from Rockland Immunochemicals Inc. (Gilbertsville, PA, USA).

Techniques: Reverse Transcription Polymerase Chain Reaction, Transfection, Control, Western Blot, Expressing