Angiogenesis is coordinated by VEGF and Notch signaling. DLL4-induced Notch signaling inhibits tip cell formation and vessel branching. To ensure proper Notch signaling, receptors and ligands are clustered at adherens junctions. However, little is known about factors that control Notch activity by influencing the cellular localization of Notch ligands. Here we show that the multiple PDZ domain protein (MPDZ) enhances Notch signaling activity. MPDZ physically interacts with the intracellular carboxyterminus of DLL1 and DLL4 and enables their interaction with the adherens junction protein Nectin-2. Inactivation of the MPDZ gene leads to impaired Notch signaling activity and increased blood vessel sprouting in cellular models and the embryonic mouse hindbrain. Tumor angiogenesis was enhanced upon endothelial-specific inactivation of MPDZ leading to an excessively branched and poorly functional vessel network resulting in tumor hypoxia. As such, we identified MPDZ as a novel modulator of Notch signaling by controlling ligand recruitment to adherens junctions.
- Andreas Fischer
- Andreas Fischer
- Fabian Tetzlaff
- Amitai Menuchin
- David Sprinzak
- Andreas Fischer
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Animal experimentation: Mice were kept under pathogen-free barrier conditions. All animal procedures were performed in accordance with the institutional and national regulations and approved by the local committees for animal experimentation (Heidelberg University and DKFZ) and the local government (Regierungspräsidium Karlsruhe, Germany).(reference number: 35-9185.81/G-30/14 and 35-9185.81/G-259/12).
- Elisabetta Dejana, FIRC Institute of Molecular Oncology, Italy
© 2018, Tetzlaff et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
Chemical manipulation of estrogen receptor alpha ligand binding domain structural mobility tunes receptor lifetime and influences breast cancer therapeutic activities. Selective estrogen receptor modulators (SERMs) extend ERα cellular lifetime/accumulation. They are antagonists in the breast but agonists in the uterine epithelium and/or in bone. Selective estrogen receptor degraders/downregulators (SERDs) reduce ERα cellular lifetime/accumulation and are pure antagonists. Activating somatic ESR1 mutations Y537S and D538G enable resistance to first-line endocrine therapies. SERDs have shown significant activities in ESR1 mutant setting while few SERMs have been studied. To understand whether chemical manipulation of ERα cellular lifetime and accumulation influences antagonistic activity, we studied a series of methylpyrollidine lasofoxifene derivatives that maintained the drug's antagonistic activities while uniquely tuning ERα cellular accumulation. These molecules were examined alongside a panel of antiestrogens in live cell assays of ERα cellular accumulation, lifetime, SUMOylation, and transcriptional antagonism. High-resolution x-ray crystal structures of WT and Y537S ERα ligand binding domain in complex with the methylated lasofoxifene derivatives or representative SERMs and SERDs show that molecules that favor a highly buried helix 12 antagonist conformation achieve the greatest transcriptional suppression activities in breast cancer cells harboring WT/Y537S ESR1. Together these results show that chemical reduction of ERα cellular lifetime is not necessarily the most crucial parameter for transcriptional antagonism in ESR1 mutated breast cancer cells. Importantly, our studies show how small chemical differences within a scaffold series can provide compounds with similar antagonistic activities, but with greatly different effects of the cellular lifetime of the ERα, which is crucial for achieving desired SERM or SERD profiles.
Tyrosine phosphorylation, orchestrated by tyrosine kinases and phosphatases, modulates a multi-layered signaling network in a time- and space-dependent manner. Dysregulation of this post-translational modification is inevitably associated with pathological diseases. Our previous work has demonstrated that non-receptor tyrosine kinase FER is upregulated in ovarian cancer, knocking down which attenuates metastatic phenotypes. However, due to the limited number of known substrates in the ovarian cancer context, the molecular basis for its pro-proliferation activity remains enigmatic. Here, we employed mass spectrometry and biochemical approaches to identify insulin receptor substrate 4 (IRS4) as a novel substrate of FER. FER engaged its kinase domain to associate with the PH and PTB domains of IRS4. Using a proximity-based tagging system in ovarian carcinoma-derived OVCAR-5 cells, we determined that FER-mediated phosphorylation of Tyr779 enables IRS4 to recruit PIK3R2/p85β, the regulatory subunit of PI3K, and activate the PI3K-AKT pathway. Rescuing IRS4-null ovarian tumor cells with phosphorylation-defective mutant, but not WT IRS4 delayed ovarian tumor cell proliferation both in vitro and in vivo. Overall, we revealed a kinase-substrate mode between FER and IRS4, and the pharmacological inhibition of FER kinase may be beneficial for ovarian cancer patients with PI3K-AKT hyperactivation.