Cell Signaling Technology

Product Pathways - Translational Control

Phospho-p70 S6 Kinase (Thr389) (1A5) Mouse mAb #9206

Applications Reactivity Sensitivity MW (kDa) Isotype
W H M R Mk Dm Endogenous 70, 85 Mouse IgG2a

Applications Key:  W=Western Blotting
Reactivity Key:  H=Human  M=Mouse  R=Rat  Mk=Monkey  Dm=D. melanogaster
Species cross-reactivity is determined by western blot. Species enclosed in parentheses are predicted to react based on 100% sequence homology.

Protocols

Specificity / Sensitivity

Phospho-p70 S6 Kinase (Thr389) (1A5) Mouse mAb detects endogenous levels of p70 S6 kinase only when phosphorylated at Thr389. This antibody also detects p85 S6 kinase when phosphorylated at the analogous site (Thr412) and possibly S6KII phosphorylated at Thr401.

Source / Purification

Monoclonal antibody is produced by immunizing animals with a synthetic phosphopeptide corresponding to residues surrounding Thr389 of human p70 S6 kinase.

Western Blotting

Western Blotting

Western blot analysis of extracts from NIH/3T3 cells, serum-starved or serum-treated (20%) using Phospho-p70 S6 Kinase (Thr389) (1A5) Mouse mAb (upper) and p70 S6 Kinase Antibody #9202 (lower).

Background

p70 S6 kinase is a mitogen activated Ser/Thr protein kinase that is required for cell growth and G1 cell cycle progression (1,2). p70 S6 kinase phosphorylates the S6 protein of the 40S ribosomal subunit and is involved in translational control of 5' oligopyrimidine tract mRNAs (1). A second isoform, p85 S6 kinase, is derived from the same gene and is identical to p70 S6 kinase except for 23 extra residues at the amino terminus, which encode a nuclear localizing signal (1). Both isoforms lie on a mitogen activated signaling pathway downstream of phosphoinositide-3 kinase (PI-3K) and the target of rapamycin, FRAP/mTOR, a pathway distinct from the Ras/MAP kinase cascade (1). The activity of p70 S6 kinase is controlled by multiple phosphorylation events located within the catalytic, linker and pseudosubstrate domains (1). Phosphorylation of Thr229 in the catalytic domain and Thr389 in the linker domain are most critical for kinase function (1). Phosphorylation of Thr389, however, most closely correlates with p70 kinase activity in vivo (3). Prior phosphorylation of Thr389 is required for the action of phosphoinositide 3-dependent protein kinase 1 (PDK1) on Thr229 (4,5). Phosphorylation of this site is stimulated by growth factors such as insulin, EGF and FGF, as well as by serum and some G-protein-coupled receptor ligands, and is blocked by wortmannin, LY294002 (PI-3K inhibitor) and rapamycin (FRAP/mTOR inhibitor) (1,6,7). Ser411, Thr421 and Ser424 lie within a Ser-Pro-rich region located in the pseudosubstrate region (1). Phosphorylation at these sites is thought to activate p70 S6 kinase via relief of pseudosubstrate suppression (1,2). Another LY294002 and rapamycin sensitive phosphorylation site, Ser371, is an in vitro substrate for mTOR and correlates well with the activity of a partially rapamycin resistant mutant p70 S6 kinase (8).

  1. Pullen, N. and Thomas, G. (1997) FEBS Lett. 410, 78-82.
  2. Dufner, A. and Thomas, G. (1999) Exp. Cell Res. 253, 100-109.
  3. Weng, Q.P. et al. (1998) J. Biol. Chem. 273, 16621-16629.
  4. Pullen, N. et al. (1998) Science 279, 707-710.
  5. Alessi, D.R. et al. (1998) Curr. Biol. 8, 69-81.
  6. Polakiewicz, R.D. et al. (1998) J. Biol. Chem. 273, 23534-23541.
  7. Fingar, D.C. et al. (2002) Genes Dev. 16, 1472-1487.
  8. Saitoh, M. et al. (2002) J. Biol. Chem. 277, 20104-20112.

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