Figure 5. Signaling pathways involving TGFβ, Smad4, and Lkb1.

The solid lines represent the plasma membrane, and the discontinuous lines show the nuclear membrane. TGFβ binds to two distinct receptor types, known as type I and type II. Both type I and type II receptors contain serine/threonine kinase domains in their intracellular portions. The type II receptor kinases are constitutively active. When they bind to the ligand, they form hetero-tetrameric complexes composed of two molecules each of type I and type II receptors. In the hetero-tetrameric receptor I and II complexes, type II receptor kinases transphosphorylate the GS-domain of type I receptor, which, in turn, phosphorylate intracellular substrates. Smad proteins are major signaling molecules acting downstream of the serine/threonine kinase TGFβ receptors. Smads are classified into three subclasses, i.e. receptor-regulated Smads (R-Smads), common-partner Smad (co-Smad: Smad 4), and inhibitory Smads (I-Smads).

After activation of TGFβRI, the signal activates Smad2 and Smad3 proteins bound to the receptors, by phosphorylation of their C-terminal (SXS motif) residues. The next step is formation of a functional trimeric complex by phosphorylated R-Smad and Smad4, and then this complex is translocated to the nucleus, where it regulates the transcription of TGFβ1-dependent genes; thus Smads have the activity of transcription factors. The activity of TGFβ signaling pathway is regulated by a negative regulatory feedback loop mediated by I-Smads (Smads inhibitors): Smad 6 and 7. They are able to interact with membrane receptors by forming stable complex with activated TGFβRI, and thus impairing their interaction with the R-Smad (inhibition of their phosphorylation). Smad7 expression is induced by TGFβ, leading to inhibition of the cellular response to this cytokine. Smad7 has been shown to promote recruitment of E3 ubiquitin ligases, including Smad ubiquitin regulatory factors (Smurf1/2) into the receptor complex. Binding of Smad7 and Smurf to the receptor complex also results in competitive inhibition of Smad2/3 binding to TGFβRI. Direct Interaction of Ski with either Smad3 or Smad4 is necessary and sufficient for Ski-mediated repression of TGFβ signaling (133).

Lkb1 is a well-studied tumor suppressor kinase that regulates cell growth and polarity. It encodes a serine-threonine kinase that directly phosphorylates and activates AMPK, a central metabolic sensor (134). Lkb1 is capable of phosphorylating Smad4 on its DNA-binding domain, inhibiting Smad4 from binding to either TGFβ- or bone morphogenetic protein-specific promoter sequences, thereby Smad4-dependent transcription (135). Lkb1 activates AMPK and send signals to Foxo3, Tsc1/2, p53, fatty acid synthase, and other molecules, which enforces metabolic checkpoints for regulating cell growth and metabolism.