👤 Wouter de Laat

Enhancer RNAs (eRNAs) are a pivotal class of enhancer-derived non-coding RNAs that drive gene expression. Here we identify the SNAI1 enhancer RNA (SNAI1e; SCREEM2) as a key activator of SNAI1 expression and a potent enforcer of transforming growth factor-β (TGF-β)/SMAD signaling in cancer cells. SNAI1e depletion impairs TGF-β-induced epithelial-mesenchymal transition (EMT), migration, in vivo extravasation, stemness, and chemotherapy resistance in breast cancer cells. SNAI1e functions as an eRNA to cis-regulate SNAI1 enhancer activity by binding to and strengthening the enrichment of the transcriptional co-activator bromodomain containing protein 4 (BRD4) at the local enhancer. SNAI1e selectively promotes the expression of SNAI1, which encodes the EMT transcription factor SNAI1. Furthermore, we reveal that SNAI1 interacts with and anchors the inhibitory SMAD7 in the nucleus, and thereby prevents TGF-β type I receptor (TβRI) polyubiquitination and proteasomal degradation. Our findings establish SNAI1e as a critical driver of SNAI1 expression and TGF-β-induced cell plasticity. Show less

Genetics play a significant role in coagulation phenotype and venous thromboembolism risk. Resistance to the anticoagulant activated protein C (APC) is an established risk for thrombosis. Herein, we explored the genetic determinants of thrombin generation (TG) and thrombomodulin (TM)-modulated TG using plasma from the Human Functional Genomics Project. Calibrated TG was measured both in absence and presence of TM using tissue factor as trigger. Genetic determinants of TG parameters and protein C pathway function were assessed using genome-wide single-nucleotide polymorphism (SNP) genotyping. Plasma samples were supplemented with purified apolipoprotein A-IV, prekallikrein, or kallikrein to test their influence on the anticoagulant function of TM and APC in TG. Thrombin generation data from 392 individuals were analyzed. Genotyping showed that the KLKB1 gene (top SNP: rs4241819) on chromosome 4 was associated with the normalized sensitivity ratio of endogenous thrombin potential to TM at genome-wide level (nETP-TMsr, P = 4.27 × 10 Our results suggest that kallikrein plays a role in the regulation of the anticoagulant protein C pathway in TG, which may provide a novel mechanism for the previously observed association between the KLKB1 gene and venous thrombosis. Show less

Neurotrophins, essential regulators of many aspects of neuronal differentiation and function, signal via four receptors, p75, TrkA, TrkB and TrkC. The three Trk paralogs are members of the LIG superfamily of membrane proteins, which share extracellular domains consisting of leucine-rich repeat and C2 Ig domains. Another LIG protein, LINGO-1 has been reported to bind and influence signaling of p75 as well as TrkA, TrkB and TrkC. Here we examine the manner in which LINGO-1 influences the function of TrkA, TrkB and TrkC. We report that Trk activation promotes Trk association with LINGO-1, and that this association promotes Trk degradation by a lysosomal mechanism. This mechanism resembles the mechanism by which another LIG protein, LRIG1, promotes lysosomal degradation of receptor tyrosine kinases such as the EGF receptor. We present evidence indicating that the Trk/LINGO-1 interaction occurs, in part, within recycling endosomes. We show that a mutant form of LINGO-1, with much of the extracellular domain deleted, has the capacity to enhance TrkA signaling in PC12 cells, possibly by acting as an inhibitor of Trk down-regulation by full length LINGO-1. We propose that LINGO-1 functions as a negative feedback regulator of signaling by cognate receptor tyrosine kinases including TrkA, TrkB and TrkC. Show less

Sequential proteolytic cleavages of amyloid-β protein precursor (AβPP) by β-secretase and γ-secretase generate amyloid β (Aβ) peptides, which are thought to contribute to Alzheimer's disease (AD). Much of this processing occurs in endosomes following endocytosis of AβPP from the plasma membrane. However, this pathogenic mode of processing AβPP may occur in competition with lysosomal degradation of AβPP, a common fate of membrane proteins trafficking through the endosomal system. Following up on published reports that LINGO-1 binds and promotes the amyloidogenic processing of AβPP we have examined the consequences of LINGO-1/AβPP interactions. We report that LINGO-1 and its paralogs, LINGO-2 and LINGO-3, decrease processing of AβPP in the amyloidogenic pathway by promoting lysosomal degradation of AβPP. We also report that LINGO-1 levels are reduced in AD brain, representing a possible pathogenic mechanism stimulating the generation of Aβ peptides in AD. Show less

Axon outgrowth inhibition in response to trauma is thought to be mediated via the binding of myelin-associated inhibitory factors (e.g. Nogo-66, myelin-associated glycoprotein, oligodendrocyte myelin glycoprotein, and myelin basic protein) to a putative tripartite LINGO-1·p75(NTR)·Nogo-66 receptor (NgR) complex at the cell surface. We found that endogenous LINGO-1 expression in neurons in the cortex and cerebellum is intracellular. Mutation or truncation of the highly conserved LINGO-1 C terminus altered this intracellular localization, causing poor intracellular retention and increased plasma membrane expression. p75(NTR) associated predominantly with natively expressed LINGO-1 containing immature N-glycans, characteristic of protein that has not completed trans-Golgi-mediated processing, whereas mutant forms of LINGO-1 with enhanced plasma membrane expression did not associate with p75(NTR). Co-immunoprecipitation experiments demonstrated that LINGO-1 and NgR competed for binding to p75(NTR) in a manner that is difficult to reconcile with the existence of a LINGO-1·p75(NTR)·NgR ternary complex. These findings contradict models postulating functional LINGO-1·p75(NTR)·NgR complexes in the plasma membrane. Show less