Non-Smad Signaling Pathways of the TGF-β Family

Abstract

Transforming growth factor β (TGF-β) and structurally related factors use several intracellular signaling pathways in addition to Smad signaling to regulate a wide array of cellular functions. These non-Smad signaling pathways are activated directly by ligand-occupied receptors to reinforce, attenuate, or otherwise modulate downstream cellular responses. This review summarizes the current knowledge of the mechanisms by which non-Smad signaling pathways are directly activated in response to ligand binding, how activation of these pathways impinges on Smads and non-Smad targets, and how final cellular responses are affected in response to these noncanonical signaling modes.

Figure 1.

Transforming growth factor β (TGF-β)-induced activation of the extracellular signal-regulated kinase (Erk) mitogen-activated protein kinase (MAPK) pathway. On binding of TGF-β to its receptor complex, the constitutively active type II receptors phosphorylate the type I receptors at Ser/Thr residues, and induce tyrosine phosphorylation of both the type I and II receptors and ShcA. The phosphorylated tyrosines then recruit Grb2/Sos to activate Erk1/2 MAPK through activation of Ras, Raf, and MEK1/2 in a Smad-independent manner. ShcA directs the TGF-β receptors to caveolin-1-containing lipid raft for Erk MAPK activation. Clathrin-dependent internalization of TGF-β receptors into the Smad anchor for receptor activation (SARA)-containing early endosome is required for Smad activation. Activated Erk MAPK contributes to epithelial-to-mesenchymal transition (EMT) by phosphorylating targeted transcription factors, which in turn control transcription of EMT-related genes without or in cooperation with activated Smad complexes. Erk MAPK also directly phosphorylates R-Smads, thus controlling their activity.

Figure 2.

Transforming growth factor β (TGF-β)-induced activation of c-Jun amino terminal kinase (JNK) and p38 mitogen-activated protein kinase (MAPK) signaling, and the IκB kinase (IKK) pathway. Ligand-bound TGF-β receptors interact with TRAF6 and TGF-β-activated kinase 1 (TAK1), and induce Lys63-linked polyubiquitylation of TRAF6, which then activates TAK1 and downstream kinases, such as JNK, p38 MAPK, and IKK. Smad6 inhibits the TGF-β-induced JNK and p38 MAPK activation by recruiting a deubiquitylase A20 to deubiquitylate TRAF6, whereas Smad7 promotes TGF-β-induced JNK and p38 MAPK activation by directing the TGF-β receptors to caveolin-1-containing lipid rafts through interaction with Smurf2. Activated JNK and p38 MAPK then phosphorylate their targeted transcription factors, and IKK phosphorylates nuclear factor-κB (NF-κB), and these transcription factors cooperate with activated Smads to regulate apoptosis and epithelial-to-mesenchymal transition (EMT). In addition, JNK directly phosphorylates receptor-activated Smad (R-Smad) to regulate Smad activity. TRAF6 can also stimulate proteolytic cleavage of TβRI dependent on presenillin 1 and Lys63 ubiquitylation of TRAF6 to generate the intracellular domain (ICD) of TβRI, which then translocates into the nucleus to regulate cell invasion. SARA, Smad anchor for receptor activation; MKK, MAP kinase kinase; TRAF6, tumor necrosis factor receptor–associated factor 6.

Figure 3.

Transforming growth factor β (TGF-β)-induced activation of the PI3K-Akt pathway. TGF-β can induce PI3K and Akt activation, possibly through interaction of the p85 subunit of PI3K with the TGF-β receptors. Activated Akt then controls translational responses through mTOR1 and S6K, or directly acts on the translational responses by inducing phosphorylation of hnRNPE. TGF-β also induces activation of mTORC2, which contributes to enhanced Akt activation and forms a reinforcing feedback loop in PI3K-Akt activation. These non-Smad-mediated translational responses collaborate with Smad-mediated transcriptional responses during epithelial-to-mesenchymal transition (EMT), but can antagonize Smad-mediated transcription responses during growth arrest or apoptosis. Akt can regulate Smad3 activity by sequestering Smad3 in the cytoplasm or inhibiting GSK3β-mediated Smad3 phosphorylation and degradation. Akt can also regulate apoptosis or EMT by directly phosphorylating FoxO or Twist1 or inducing Snail phosphorylation by GSK3β.

Figure 4.

Transforming growth factor β (TGF-β)-induced regulation of the Rho family of small GTPases. RhoA and Rho-associated protein kinase (ROCK) can be activated by TGF-β by either Smad-dependent or -independent mechanisms to induce actin polymerization and stress fiber formation during epithelial-to-mesenchymal transition (EMT). Additionally, the TβRII receptor can directly phosphorylate Par6 by recruiting Smurf1, thus targeting RhoA for degradation, which leads to tight junction dissociation. TGF-β can also induce tight junction dissociation and cell migration during EMT by recruiting Cdc42 and/or Rac1 to the receptor complex, and activate p21-activated kinase (PAK) signaling. PKC, Protein kinase C.