Laboratory for signal transduction mechanisms of TGF-ß

TGF-β signal transduction

Transforming growth factor- β (TGF- β ) is a secreted cytokine that regulates numerous responses, such as proliferation, differentiation, migration and apoptosis. TGF- β is the prototypic member of a family of 33 structurally related pleiotropic proteins. Members of the TGF- β superfamily have crucial roles in development and tissue homeostasis. We are interested in unraveling the mechanisms by which transforming growth factor- β (TGF- β ) family members elicit their multifunctional cellular effects and how perturbation in their signal transduction pathways contribute to human diseases. Previous research from us and other laboratories have now firmly established the intracellular signaling cascade of TGF- β via serine/threonine kinase receptors and SMAD transcriptional effectors. However, how this pathway is (mis)regulated in cancer and other diseases, remains not well understood. The present focus within our group is on 1) the identification of novel critical regulators of TGF- β family signaling pathways using functional genetic screens, 2) how TGF- β specificity and bioavailability is regulated via known (co-)receptors and ligand binding proteins and 3) chemical biology approaches to normalize deregulated TGF- β signalling. In particular, we study the involvement of TGF- β family members in the metastasis of cancer cells, tumor angiogenesis and cardiovascular diseases. We use 3-dimensional cell culture systems that allow for a functional analysis of homo- and heterotypic cell-cell interactions in structures that resemble those found in vivo. Moreover, mouse and zebra fish models are used that mimic diseases with misregulated TGF- β signaling. Our long term aim is to translate our findings towards the development of novel treatment modalities. 

2016 highlights:

BMP-9 interferes with liver regeneration and promotes liver fibrosis.

BMP9 is constitutively produced in the liver. Systemic levels act on many organs and tissues including bone and endothelium, but little is known about its hepatic functions in health and disease. Levels of BMP-9 and its receptors were analysed in primary liver cells. We investigated direct effects of BMP-9 on hepatic stellate cells (HSCs) and hepatocytes in vitro, and in acute and chronic liver injury models in mice. Quiescent and activated HSCs were identified as major BMP-9 producing liver cell type. BMP-9 stimulation of cultured hepatocytes inhibited proliferation, epithelial to mesenchymal transition and preserved expression of important metabolic enzymes such as cytochrome P450. Acute liver injury caused by partial hepatectomy or single injections of carbon tetrachloride (CCl4) or lipopolysaccharide (LPS) into mice resulted in transient downregulation of hepatic BMP-9 mRNA expression. Correspondingly, LPS stimulation led to downregulation of BMP-9 expression in cultured HSCs. Application of BMP-9 after partial hepatectomy significantly enhanced liver damage and disturbed the proliferative response. Chronic liver damage in BMP-9-deficient mice or in mice adenovirally overexpressing the selective BMP-9 antagonist activin-like kinase 1-Fc resulted in reduced deposition of collagen and subsequent fibrosis. Constitutive expression of low levels of BMP-9 stabilises hepatocyte function in the healthy liver. Upon HSC activation, endogenous BMP-9 levels increase in vitro and in vivo and high levels of BMP-9 cause enhanced damage upon acute or chronic injury.

TGFβ signaling in colorectal cancer cells with microsatellite instability despite biallelic mutations in TGFBR2. 

Most colorectal cancer (CRC) cells with high levels of microsatellite instability (MSI-H) accumulate mutations at a microsatellite sequence in the gene encoding TGFBR2. TGFβ signaling therefore is believed to be defective in these tumors, although CRC cells with TGFBR2 mutations have been reported to remain sensitive to TGFβ. We investigated how TGFβ signaling might continue in MSI-H CRC cells. SMAD2 was phosphorylated in most MSI-H CRC tissues (strong detection in 44% and weak detection in 34% of MSI-H tumors). Phosphorylation of SMAD2 in MSI-H cells required TGFBR2—even the form encoding a frameshift mutation. Transcription and translation of TGFBR2 with a 1-nucleotide deletion at its microsatellite sequence still produced a full-length TGFBR2 protein. However, protein expression required preservation of the TGFBR2 microsatellite sequence; cells in which this sequence was replaced with a synonymous nonmicrosatellite sequence did not produce functional TGFBR2 protein. Thus, TGFβ signaling remains active in some MSI-H CRC cells despite the presence of frameshift mutations in the TGFBR2 gene because the mutated gene still expresses a functional protein.

FAF1 phosphorylation by AKT accumulates TGF-β type II receptor and drives breast cancer metastasis.

TGF-β is pro-metastatic for the late-stage breast cancer cells. Despite recent progress, the regulation of TGF-β type II receptor remains uncertain. Here we report that FAF1 destabilizes TβRII on the cell surface by recruiting the VCP/E3 ligase complex, thereby limiting excessive TGF-β response. Importantly, activated AKT directly phosphorylates FAF1 at Ser 582, which disrupts the FAF1-VCP complex and reduces FAF1 at the plasma membrane. The latter results in an increase in TβRII at the cell surface that promotes both TGF-β-induced SMAD and non-SMAD signalling. We uncovered a metastasis suppressing role for FAF1 through analyses of FAF1-knockout animals, various in vitro and in vivo models of epithelial-to-mesenchymal transition and metastasis, an MMTV-PyMT transgenic mouse model of mammary tumour progression and clinical breast cancer samples. These findings describe a previously uncharacterized mechanism by which TβRII is tightly controlled. Together, we reveal how SMAD and AKT pathways interact to confer pro-oncogenic responses to TGF-β.