Research ResourceBiochemistry

Proximity biotinylation identifies a set of conformation-specific interactions between Merlin and cell junction proteins

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Science Signaling  23 Apr 2019:
Vol. 12, Issue 578, eaau8749
DOI: 10.1126/scisignal.aau8749

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The Merlin interactome identifies cell junction proteins

Merlin (also called NF2) is a tumor suppressor that associates with lipid rafts and cell junctions. To identify potential Merlin binding partners, Hennigan et al. performed proximity biotinylation assays in murine Schwann cells with wild-type and various mutant forms of Merlin that could not associate with the plasma membrane or were locked in different conformations. Fewer proteins interacted with the form of Merlin that could not associate with the plasma membrane, and some proteins interacted specifically with the closed conformation. Most of the Merlin-proximal proteins were components of or associated with cell junctions and focal adhesions. Among the previously unidentified Merlin-interacting proteins was the tumor suppressor ASPP2 (also called Tp53bp2), which bound directly to the closed conformation. This dataset confirms cell junctions as important sites for Merlin activity and reveals additional binding partners.

Abstract

Neurofibromatosis type 2 is an inherited, neoplastic disease associated with schwannomas, meningiomas, and ependymomas and that is caused by inactivation of the tumor suppressor gene NF2. The NF2 gene product, Merlin, has no intrinsic catalytic activity; its tumor suppressor function is mediated through the proteins with which it interacts. We used proximity biotinylation followed by mass spectrometry and direct binding assays to identify proteins that associated with wild-type and various mutant forms of Merlin in immortalized Schwann cells. We defined a set of 52 proteins in close proximity to wild-type Merlin. Most of the Merlin-proximal proteins were components of cell junctional signaling complexes, suggesting that additional potential interaction partners may exist in adherens junctions, tight junctions, and focal adhesions. With mutant forms of Merlin that cannot bind to phosphatidylinositol 4,5-bisphosphate (PIP2) or that constitutively adopt a closed conformation, we confirmed a critical role for PIP2 binding in Merlin function and identified a large cohort of proteins that specifically interacted with Merlin in the closed conformation. Among these proteins, we identified a previously unreported Merlin-binding protein, apoptosis-stimulated p53 protein 2 (ASPP2, also called Tp53bp2), that bound to closed-conformation Merlin predominately through the FERM domain. Our results demonstrate that Merlin is a component of cell junctional mechanosensing complexes and defines a specific set of proteins through which it acts.

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