anti ps6 ribosomal protein s235 236  (Cell Signaling Technology Inc)

Journal: bioRxiv

Article Title: L-pentahomoserine correlates with therapy outcome in esophageal cancer and promotes metabolic adaptations that support cell survival under nutrient-deprived conditions

doi: 10.64898/2026.01.15.699640

Figure Lengend Snippet: a ) Chemical structure of L-serine and L-2A5HPA. Representative volcano plot of untargeted analysis comparing SK-GT-4 ( b ) and OA-P4C cells ( c ) treated with or without L-2A5HPA (50 µM). The grey circles represent metabolites detected, and L-2A5HPA is highlighted in red. Intracellular levels of L-serine and L-2A5HPA are shown in SK-GT-4 cells ( d ) and OA-P4C cells ( e ). f ) Correlation between the plasma levels of serine and L-2A5HPA (n=11). The Pearson r value and P value are shown. g ) Representative image of SW982 cells that were nutrient-deprived in the absence or presence of L-2A5HPA (0, 50, 200 and 2,000 µM) for 4 days, recovered in complete media for 8 days and stained with sulforhodamine B. h ) Quantification of sulforhodamine B absorbance at 510 nm. i ) Immunoblot analysis of U2OS cells nutrient-deprived in the presence or absence of L-2A5HPA for 5 days. mTORC1 activation was measured by the levels of phosphor-S6 (S235-236). Total S6 is shown as a loading control. j ) Representative images from U2OS cells starved and treated with vehicle, L-Serine (2mM) or L-2A5HPA (2mM) for 5 days. k ) Representative images from cells treated as in j ), then exposed to complete media for 5 days and stained with sulforhodamine B. l ) Quantification of sulforhodamine B absorbance at 510 nm from three biological replicates. m ) Representative images of starved cells treated with either L-2A5HPA (2mM) or D-2A5HPA (2mM) for 5 days. n ) Representative images from cells stained with sulforhodamine B after being treated as in m ), then exposed to complete media for 5 days. o ) Quantification of sulforhodamine B absorbance at 510 nm of three biological replicates. p ) Relative intracellular levels of L-2A5HPA or D-2A5HPA in U2OS cells after being exposed to 50 µM of each of the enantiomers for 1 min. Bars represent the average ± SEM of three independent replicates. Data were analyzed using a one-way ANOVA, Student’s t-test or One-sample t-test (p). Circles represent metabolites, independent experiments or individual patients.

Article Snippet: The primary antibodies used: Cleaved Caspase 3 (Cell Signaling, 9662 dilution 1:500), Cleaved PARP (Cell Signaling, 9532 dilution 1:1000), P62 (Cell Signaling, 397495 dilution 1:1000), LC3BII (Cell Signaling, 3868 dilution 1:1000), P-S6 S235/236 (Cell Signaling 4858 dilution 1:1000), S6 (Cell Signaling, 2317 dilution 1:1000), β-actin (Cell Signaling 4970S dilution 1:1000), Vinculin (Merck, MAB3574 dilution 1:1000), PCB (ab126707 dilution 1:1000).

Techniques: Clinical Proteomics, Staining, Western Blot, Activation Assay, Control