The Lnx Family Proteins Function as Molecular Scaffolds for Numb Family Proteins

Rice, Dennis S.; Northcutt, Grace M.; Kurschner, Cornelia

doi:10.1006/mcne.2001.1024

Cited by 61 publications

(109 citation statements)

References 57 publications

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“…Moreover, we also obtained LNX2 from the original yeast two-hybrid screening, suggesting that LNX2 is likewise a binding partner for JAM4. The molecular structure of LNX2 is similar to LNXp80 and has a RING finger domain (Rice et al, 2001). In the case that JAM4 is anchored to LNXp80 or LNX2, JAM4 may be degraded after endocytosis.…”

Section: Discussionmentioning

confidence: 99%

Ligand-of-Numb protein X is an endocytic scaffold for junctional adhesion molecule 4

et al. 2006

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Junctional adhesion molecule 4 (JAM4) is a cell adhesion molecule that interacts with a tight junction protein, membrane-associated guanylate kinase inverted 1 (MAGI-1). Our previous studies suggest that JAM4 is implicated in the regulation of paracellular permeability and the signalings of hepatocyte growth factor. In this study, we performed yeast two-hybrid screening to search for an unidentified JAM4-binding protein and obtained one isoform of Ligand-of-Numb protein X1 (LNX1), LNXp70, that is an interactor of Numb. Ligand-of-Numb protein X1 is expressed in kidney glomeruli and intestinal epithelial cells, where JAM4 is also detected. Immunoprecipitation from kidney lysates supports the in vivo interaction of proteins. Biochemical studies reveal that JAM4 directly binds the second PDZ domain of LNX1 through its carboxyl terminus. Junctional adhesion molecule 4, LNX1 and Numb form a tripartite complex in vitro and are partially colocalized in heterologous cells. Ligand-of-Numb protein X1 facilitates endocytosis of JAM4 and is involved in transforming growth factor b -induced redistribution of JAM4 in mammary epithelial cells. Experiments using dominant-negative constructs and RNA interference insure that Numb is necessary for the LNX1-mediated endocytosis of JAM4. All these findings indicate that LNX1 provides an endocytic scaffold for JAM4 that is implicated in the reorganization of cell junctions.

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Section: Discussionmentioning

confidence: 99%

Ligand-of-Numb protein X is an endocytic scaffold for junctional adhesion molecule 4

et al. 2006

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“…We have previously shown that Numb is an LNX substrate and that Numb binding to LNX promotes its ubiquitination and proteasomal degradation. A highly related protein called LNX2 has also been characterized (17) that is capable of binding to the PTB domain of mNumb, although its potential function as an E3 ligase was not tested. In addition to mNumb, the coxsackievirus and adenovirus receptor was identified as an LNX binding partner (18).…”

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confidence: 99%

A Novel PTB-PDZ Domain Interaction Mediates Isoform-specific Ubiquitylation of Mammalian Numb

Nie

McGlade

2004

Journal of Biological Chemistry

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LNX was originally cloned as a Numb PTB-binding molecule, and it was subsequently found to act as a RING finger-type E3 ubiquitin ligase for the ubiquitylation and degradation of mNumb. Numb is a PTB domaincontaining protein that functions as an intrinsic determinant of cell fate in asymmetric cell division. In mammals, four protein isoforms arise from alternative mRNA splicing. Here we report that while all four protein isoforms bind to LNX, only p72 and p66 Numb isoforms are ubiquitylated and degraded. The p72 and p66 Numb proteins differ from the other two isoforms by the presence of an 11-amino acid sequence insert in the PTB domain (PTBi). We demonstrate that the isoform-specific ubiquitylation of mNumb is due to a novel interaction between the first PDZ domain (PDZ1) of LNX and the PTBi variant. Deletion of LNX PDZ1 domain resulted in loss of ubiquitylation and subsequent degradation of the PTBi form of Numb. Interestingly efficient PTBi ubiquitylation not only depends on association with the LNX PDZ1 domain but also requires binding to the canonical PTB-binding motif NPAY in LNX. Thus two distinct modes of PTBi-mediated interaction with LNX work in concert to allow the effective and specific degradation of the p72 and p66 isoforms of mNumb.Controlled protein degradation mediated by ubiquitin-dependent proteolysis underlies the regulation of many critical biological events during the life and death of a cell as well as in health and disease (1). Ubiquitylation of a target protein is achieved through a cascade involving at least three enzymes, namely E1, 1 E2, and E3, which activate and transfer ubiquitin to the substrate in a sequential manner. Polyubiquitylated proteins are recognized by the 26 S proteasome and are subsequently degraded into small peptides (2, 3).E3 ubiquitin ligases are responsible for the specific recognition of a multitude of substrates. Two major classes of E3 have been identified to date characterized by the presence of either a HECT (homologous to E6-AP carboxyl terminus) domain or a RING finger domain. HECT-type E3s catalyze ubiquitylation of a substrate by first forming an E3-ubiquitin thioester intermediate followed by the direct transfer of ubiquitin onto the substrate. On the other hand, RING-type E3s function as scaffolds that bring together the E2 and the target substrate to facilitate efficient transfer of the activated ubiquitin moiety from the E2 to the substrate protein (4). In addition to HECT or RING finger domains, E3 ligases also contain protein interaction modules for substrate recognition. Diversity in the repertoire of E3 substrate selection is achieved by linking a variety of protein interaction modules to the HECT or RING domains. For example, many HECT domain-containing E3s share the WW domain, which is involved in protein-protein interactions and has a role in targeting substrates containing a PY motif for ubiquitylation (5, 6). The RING-type E3 family is composed of two distinct groups, single and multisubunit proteins. Single subunit E3s such as c-Cbl contain...

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“…Lnx1 and Lnx2 mRNAs are expressed prominently in the CNS during embryonic development, with strong Lnx2 mRNA expression in the forebrain being particularly noteworthy [ 2 ]. Here, we show that both LNX1 and LNX2 proteins are present in the juvenile murine brain, albeit at low levels.…”

Section: Discussionmentioning

confidence: 57%

“…Ligand of NUMB protein X (LNX) proteins were first characterized based on their ability to bind to NUMB and NUMBLIKE [ 1,2 ]. LNX1 and LNX2 are closely related E3 ubiquitin ligases that can ubiquitinate specific isoforms of NUMB, and LNXmediated ubiquitination, at least in the case of LNX1, has been shown to target NUMB for proteasomal degradation [ 3,4,5 ].…”

Section: Introductionmentioning

confidence: 99%

“…However, LNX2 knockdown in colorectal cancer cell lines caused a decrease in NUMB levels, a result that does not fit with the notion of LNX2 targeting NUMB for degradation [ 6 ]. Developmentally, expression of both Lnx1 and Lnx2 messenger RNA (mRNA) is prominent in the embryonic and adult central nervous system (CNS) [ 1,2 ]. This observation suggests a possible role for LNX1 and LNX2 in modulating neural development through their interaction with NUMB and/or its paralogue NUMBLIKE-key regulators of mammalian neurogenesis and neuronal differentiation [ 7 ].…”

Section: Introductionmentioning

confidence: 99%

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Decreased Anxiety-Related Behaviour but Apparently Unperturbed NUMB Function in Ligand of NUMB Protein-X (LNX) 1/2 Double Knockout Mice

et al. 2016

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Type of publicationArticle (peer-reviewed) AbstractNUMB is a key regulator of neurogenesis and neuronal differentiation that can be ubiquitinated and targeted for proteasomal degradation by ligand of numb proteinX (LNX) family E3 ubiquitin ligases. However, our understanding of LNX protein function in vivo is very limited. To examine the role of LNX proteins in regulating NUMB function in vivo, we generated mice lacking both LNX1 and LNX2 expression in the brain. Surprisingly, these mice are healthy, exhibit unaltered levels of NUMB protein and do not display any neuroanatomical defects indicative of aberrant NUMB function. Behavioural analysis of LNX1/LNX2 double knockout mice revealed decreased anxiety related behaviour, as assessed in the open field and elevated plus maze paradigms. By contrast, no major defects in learning, motor or sensory function were observed. Given the apparent absence of major NUMB dysfunction in LNX null animals, we performed a proteomic analysis to identify neuronal LNXinteracting proteins other than NUMB that might contribute to the anxiolytic phenotype observed. We identified and/or confirmed interactions of LNX1 and LNX2 with proteins known to have presynaptic and neuronal signalling functions, including the presynaptic active zone constituents ERC1, ERC2 and LIPRINαs (PPFIA1, PPFIA3), as well as the FBAR domain proteins FCHSD2 (nervous wreck homologue) and SRGAP2. These and other novel LNXinteracting proteins identified are promising candidates to mediate LNX functions in the central nervous system, including their role in modulating anxietyrelated behaviour.

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The Lnx Family Proteins Function as Molecular Scaffolds for Numb Family Proteins

Cited by 61 publications

References 57 publications

Ligand-of-Numb protein X is an endocytic scaffold for junctional adhesion molecule 4

Ligand-of-Numb protein X is an endocytic scaffold for junctional adhesion molecule 4

A Novel PTB-PDZ Domain Interaction Mediates Isoform-specific Ubiquitylation of Mammalian Numb

Decreased Anxiety-Related Behaviour but Apparently Unperturbed NUMB Function in Ligand of NUMB Protein-X (LNX) 1/2 Double Knockout Mice

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