Laboratory for signal transduction mechanisms of TGF-ß

TGF-β signal transduction

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. Our long term aim is to translate our findings towards the development of novel treatment modalities. 

2017 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 (CCl 4 ) 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.

USP4 inhibits SMAD4 monoubiquitination and promotes activin and BMP signaling. 

SMAD4 is a common intracellular effector for TGF-β family cytokines, but the mechanism by which its activity is dynamically regulated is unclear. We demonstrated that ubiquitin-specific protease (USP) 4 strongly induces activin/BMP signaling by removing the inhibitory monoubiquitination from SMAD4. This modification was triggered by the recruitment of the E3 ligase, SMURF2, to SMAD4 following ligand-induced regulatory (R)-SMAD-SMAD4 complex formation. Whereas the interaction of the negative regulator c-SKI inhibits SMAD4 monoubiquitination, the ligand stimulates the recruitment of SMURF2 to the c-SKI-SMAD2 complex and triggers c-SKI ubiquitination and degradation. Thus, SMURF2 has a role in termination and initiation of TGF-β family signaling. An increase in monoubiquitinated SMAD4 in USP4-depleted mouse embryonic stem cells (mESCs) decreased both the BMP- and activin-induced changes in the embryonic stem cell fate. USP4 sustained SMAD4 activity during activin- and BMP-mediated morphogenic events in early zebrafish embryos. Moreover, zebrafish depleted of USP4 exhibited defective cell migration and slower coordinated cell movement known as epiboly, both of which could be rescued by SMAD4. Therefore, USP4 is a critical determinant of SMAD4 activity.

JUNB governs a feed-forward network of TGFβ signaling that aggravates breast cancer invasion.

It is well established that TGF-β switches its function from being a tumor suppressor to a tumor promoter during the course of tumorigenesis, which involves both cell-intrinsic and environment-mediated mechanisms. We are interested in breast cancer cells, in which SMAD mutations are rare and interactions between SMAD and other transcription factors define pro-oncogenic events. Here, we have performed chromatin immunoprecipitation (ChIP)-sequencing analyses which indicate that the genome-wide landscape of SMAD2/3 binding is altered after prolonged TGFβ stimulation. De novo motif analyses of the SMAD2/3 binding regions predict enrichment of binding motifs for activator protein (AP)1 in addition to SMAD motifs. TGFβ-induced expression of the AP1 component JUNB was required for expression of many late invasion-mediating genes, creating a feed-forward regulatory network. Moreover, we found that several components in the WNT pathway were enriched among the late TGF-β-target genes, including the invasion-inducing WNT7 proteins. Consistently, overexpression of WNT7A or WNT7B enhanced and potentiated TGF-β-induced breast cancer cell invasion, while inhibition of the WNT pathway reduced this process. Our study thereby helps to explain how accumulation of pro-oncogenic stimuli switches and stabilizes TGF-β-induced cellular phenotypes of epithelial cells.