SMAD-proteins

Activation of TGPp is accompanied by multiple phosphorylation on cytoplasmic parts of the receptor this includes Tyr phosphorylation in addition to Ser/Thr phosphorylation. It is assumed that these autophosphorylations occur in trans, i.e., between the protomers of the receptor. Prom the activated TGPp receptor, signals are directed to the transcription level. A set of proteins, known as Smad proteins, are involved in this signal conduction (see Pig. 12 review Heldin et al., 1997, Massague, 1998). Besides Smad proteins, G-proteins and the MAPK cascade are also involved in the downstream signaling of TGPp family members. 

At least nine different Smad proteins have been identified in higher organisms, and 

Structure-function investigations have shown that the Smads have a sequence-specific DNA binding domain and two regulatory domains known as the MHl and MH2 domains. It is assiuned that the pathway-restricted Smads are kept in an inactive form by intramolecular interactions between the MHl and MH2 domains. The inactive conformation is lifted by phosphorylation at the C-terminus upon activation of the TGPp receptor, and an interaction with the conunon mediator Smad 4 becomes possible. According to these observations, the MHl domain functions as a negative regulator of the MH2 domain. 

DNA binding of Smad proteins takes place via a DNA binding motif, which differs substantially from classical DNA binding motives. The DNA binding domain of the Smad proteins contains a P-sheet motif, which positions itself in the large groove of the DNA (Shi et al 1998). 

In total, TGPP-Smad signal conduction has distinct similarities to signal conduction in the Jak-Stat pathway (see 11.1.3). In both pathways, cytosolic transcription factors are activated by phosphorylation and are translocated in oligomeric complexes to the nucleus and the DNA. Common to both pathways is the short distance from the extracellular signal to the transcription level. 

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The phosphorylation state of the transcription factor NF-AT has a different effect on translocation. The phosphorylated form of this protein is localized in the cytosol and requires dephosphorylation by the protein phosphatase calcineiuln in order to be translocated to the nucleus (see also 7.5.2). Other examples for phosphorylation-dependent nuclear translocation include the STAT-proteins (see 11.1.3.2) and the SMAD-proteins (see 12.1.2). 

Receptors with Intrinsic Ser/Thr Kinase Activity the TGFP Receptor and the Smad Proteins... 

Pathway-restricted Smad proteins (Smad 1, 2, 3) These Smad proteins are effector molecules directly downstream of type I TGPp receptor. They bind to TGPp receptor I and are phosphorylated and activated for further signal conduction by the receptor. Phosphorylation takes place on a C-terminal SSXS motif... 

Inhibitory Smads Smad 6 and Smad 7 function as inhibitors of TGPp signaling. They bind to type I receptors and interfere with phosphorylation of the pathway-restricted Smads. These Smad proteins may be used to modulate and weaken the TGPp receptor. Such transmodulation has been described for the interferon y pathway. Activation of the IFNy pathway leads to increased transcription of Smad 7, which diminishes signal conduction in the TGPp pathway (Ulloa et al., 1999). 

Heldin, C.H., Miyazono, K. and ten Dijke, P. TGF-beta signalling from cell membrane to nucleus through SMAD proteins (1997) Nature 390, 465-671... 

The R-Smad proteins interact with type I receptor and co-regulatory Smad (co-Smad) proteins in the signal cascade by virtue of conserved MH1 and MH2 domains at their N- and C-termini, which are separated by a variable, non-conserved region. These domains possess DNA binding and transcriptional activation functions, respectively. The inactive MH1 domain interacts with the MH2 domain in an auto-inhibitory manner. However, once C-terminal phosphorylation by the type I receptor occurs, this inhibitory association is alleviated and the R-Smad is then able to interact with its appropriate co-Smad. The phosphorylated R-Smad/co-Smad complex is capable of translocation to the cell nucleus where, in coordination with nuclear factors, it is able to exert its transcriptional activation function. 

Zavala-Gongora, R., Kroner, A., Wittek, B., Knaus, P. and Brehm, K. (2003) Identification and characterisation of two distinct Smad proteins from the fox-tapeworm Echinococcus multilocularis. International journal for Parasitology 33, 1 665-1677. 

Figure 3.4 Factors affecting foreign body reaction and potential points of intervention at the level of the myofibroblast (1) inhibit synthesis or release of TGF-P (2) block stimulation by TGF-P of its membrane receptors on the activated fibroblast (3) inhibit the Smad proteins, which transfer the TGF-P effect to the nucleus (4) inhibit transcription of procollagen mRNA (5) inhibit translation of the message to form procollagen (6) inhibit prolyl-4-hydroxylase, which creates hydroxyproline and facilitates helix formation (7) inhibit lysyl oxidase, which cross-links the collagen (8) enhance the function of MMPs, which degrade collagen, or inhibit TIMPs, which degrade MMPs.

Fig. 6.7 Proposed model for TGF 31-linked mediation of apoptosis by angiotensin II. In one car-diomyocyte, angiotensin II promotes the upregulation of TGF 31, which is released into the intracellular space where it interacts with the TGFpl receptor of a second cardiomyocyte. Levels of Smad proteins rise in the second cardiomyocyte, causing it to undergo apoptosis.

Christian, J.L., and Nakayama, T. 1999. Can t get no SMADisfaction Smad proteins as positive and negative regulators of TGF-beta family signals. Bioessays 21 382-390. 

Evans, R.A., Tian, Y.C., Steadman, R., and Phillips, A.O. 2003. TGF-betal-mediated fibroblast-myofibroblast terminal differentiation—the role of smad proteins. Exp. Cell Res. 282 90-100. 

Hao, J., Wang, B., Jones, S.C., Jassal, D.S., and Dixon, I.M.C. 2000. Interaction between angiotensin II and Smad proteins in fibroblasts in failing heart and in vitro. Am. J. Physiol. 279 H3020-H3030. 

Lagna, G., Hata, A., Hemmati-Brivanlou, A., and Massague, J. 1996. Partnership between DPC4 and SMAD proteins in TGF-beta signalling pathways. Nature 383 832-836. 

Luo, K., Stroschein, S.L., Wang, W., Chen, D., Martens, E., Zhou, S., and Zhou, Q. 1999. The Ski oncoprotein interacts with the Smad proteins to repress TGFbeta signaling. Genes Dev. 13 2196-2206. 

It is known that SMAD proteins, identified as signaling mediators of the TGF-P superfamily, are involved in cell growth, morphogenesis, development, and... 

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