Akt kinase targets association of CBP with SMAD 3 to regulate TGFβ‐induced expression of plasminogen activator inhibitor‐1

Abstract

Transforming growth factor-beta (TGFbeta) controls expression of plasminogen activator inhibitor type 1 (PAI-1), which regulates degradation of extracellular matrix proteins in fibrotic diseases. The TGFbeta receptor-specific Smad 3 has been implicated in the PAI-1 expression. The mechanism by which non-Smad signaling contributes to this process is not known. We studied the cross-talk between Smad 3 and PI 3 kinase/Akt signaling in TGFbeta-induced PAI-1 expression in renal mesangial cells. Inhibition of PI 3 kinase and Akt kinase blocked TGFbeta- and Smad 3-mediated expression of PAI-1. In contrast, constitutively active PI 3 kinase and Akt kinase increased PAI-1 expression, similar to TGFbeta. Inhibition of PI 3 kinase and Akt kinase had no effect on TGFbeta-induced Smad 3 phosphorylation and its translocation to the nucleus. Notably, inhibition of PI 3 kinase-dependent Akt kinase abrogated TGFbeta-induced PAI-1 transcription, without affecting binding of Smad 3 to the PAI-1 Smad binding DNA element. However, PI 3 kinase inhibition and dominant negative Akt kinase antagonized the association of the transcriptional coactivator CBP with Smad 3 in response to TGFbeta, resulting in inhibition of Smad 3 acetylation. Together our findings identify TGFbeta-induced PI 3 kinase/Akt signaling as a critical regulator of Smad 3-CBP interaction and Smad 3 acetylation, which cause increased PAI-1 expression.

Transforming Growth Factor β Integrates Smad 3 to Mechanistic Target of Rapamycin Complexes to Arrest Deptor Abundance for Glomerular Mesangial Cell Hypertrophy

“…Anti-HA antibody was purchased from Covance, Princeton, NJ. Adenovirus vectors expressing HA-tagged Smad 3 (Ad Smad 3) and FLAG-tagged Smad 7 (Ad Smad 7) have been described previously (9). shRNAs targeting two different regions of deptor (deptor sh1 and deptor sh2), FLAG-deptor, and FLAG-deptor 13X were obtained from Addgene and originally constructed in the laboratory of Dr. David M. Sabatini, Whitehead Institute for Biomedical Research, Boston, MA.…”

“…Phosphorylated Smads dissociate from the type I receptor and the SARA (Smad anchor for receptor activation), a Smad-recruiting protein to the plasma membrane (6). Subsequently, phosphorylated Smad 2 and Smad 3 bind to the co-Smad, Smad 4, and the heterodimer translocates to the nucleus to recruit coactivators or corepressors for target gene regulation (7)(8)(9).…”

Plasminogen activator inhibitor 1: Mechanisms of its synergistic regulation by growth factors

“…In renal mesangial cells, TGF-β-induced PI3 Kinase/Akt signaling enhances Smad-mediated PAI-1 expression by promoting CBP-mediated Smad3 acetylation (Das, Ghosh-Choudhury et al 2008). Matsuzaki reported that Jnk, activated by TGF-β, phosphorylates Smad3 at the linker region (pSmad3L), which increases PAI-1 expression in heptocellular carcinoma (HCC) (Matsuzaki, Murata et al 2007).…”

“…Smad-mediated PAI-1 expression is enhanced by promoting CBP-mediated Smad3 acetylation (Das, Ghosh-Choudhury et al 2008). It will be interest to test whether CBP/P300 has a role in this synergistic regulation.…”

“…kinase/Akt (Das, Ghosh-Choudhury et al 2008) also contribute to increases PAI-1 level ( Figure 2A). Experiments with pharmacologic inhibitors or dominant negative proteins…”