Swift residue‐screening identifies key N‐glycosylated asparagines sufficient for surface expression of neuroglycoprotein Lingo‐1

Zhong, Xiaotian; Pocas, Jennifer; Liu, Yan; Wu, Paul W.; Mosyak, Lidia; Somers, Will; Kriz, Ron

doi:10.1016/j.febslet.2009.02.034

Cited by 6 publications

(5 citation statements)

References 15 publications

(33 reference statements)

Supporting

Mentioning

Contrasting

Order By: Relevance

“…For this purpose, we used an anti‐LINGO‐1 antibody, and its specificity was first evaluated by Western blotting on HEK‐293 cell lysates overexpressing LINGO‐1‐HA and compared with the results obtained with the anti‐HA antibody. Both antibodies labelled two prominent immunoreactive species at approximatively 80 and 100 kDa (Figure A), corresponding to different states of LINGO‐1 glycosylation (Zhong et al ., ; Supporting Information http://onlinelibrary.wiley.com/doi/10.1111/bph.12945/suppinfo) and thereby demonstrating the specificity of the anti‐LINGO‐1 antibody used here. Following chemical cross‐linking, cell lysates were subjected to SDS/PAGE under reducing conditions.…”

Section: Resultsmentioning

confidence: 67%

Targeting the cis‐dimerization of LINGO‐1 with low MW compounds affects its downstream signalling

Cobret

Tauzia

Ferent

et al. 2014

British J Pharmacology

View full text Add to dashboard Cite

BACKGROUND AND PURPOSEThe transmembrane protein LINGO-1 is a negative regulator in the nervous system mainly affecting axonal regeneration, neuronal survival, oligodendrocyte differentiation and myelination. However, the molecular mechanisms regulating its functions are poorly understood. In the present study, we investigated the formation and the role of LINGO-1 cis-dimers in the regulation of its biological activity. EXPERIMENTAL APPROACHLINGO-1 homodimers were identified in both HEK293 and SH-SY5Y cells using co-immunoprecipitation experiments and BRET saturation analysis. We performed a hypothesis-driven screen for identification of small-molecule protein-protein interaction modulators of LINGO-1 using a BRET-based assay, adapted for screening. The compound identified was further assessed for effects on LINGO-1 downstream signalling pathways using Western blotting analysis and AlphaScreen technology. KEY RESULTSLINGO-1 was present as homodimers in primary neuronal cultures. LINGO-1 interacted homotypically in cis-orientation and LINGO-1 cis-dimers were formed early during LINGO-1 biosynthesis. A BRET-based assay allowed us to identify phenoxybenzamine as the first conformational modulator of LINGO-1 dimers. In HEK-293 cells, phenoxybenzamine was a positive modulator of LINGO-1 function, increasing the LINGO-1-mediated inhibition of EGF receptor signalling and Erk phosphorylation. CONCLUSIONS AND IMPLICATIONSOur data suggest that LINGO-1 forms constitutive cis-dimers at the plasma membrane and that low MW compounds affecting the conformational state of these dimers can regulate LINGO-1 downstream signalling pathways. We propose that targeting the LINGO-1 dimerization interface opens a new pharmacological approach to the modulation of its function and provides a new strategy for drug discovery.

show abstract

Section: Resultsmentioning

confidence: 67%

Targeting the cis‐dimerization of LINGO‐1 with low MW compounds affects its downstream signalling

Cobret

Tauzia

Ferent

et al. 2014

British J Pharmacology

View full text Add to dashboard Cite

show abstract

“…Indeed, galectin‐4 has also been demonstrated to bind to N‐glycans on human colon cancer cells (Ideo et al, 2005; Morelle et al, 2009; Wu et al, 2004). Interestingly, LINGO‐1 and Notch are N‐glycosylated OLG cell surface receptors that have been shown to negatively regulate OLG maturation (Mi etal., 2005; Wang et al, 1998; Zhong et al, 2009). Accordingly, galectin‐4 might be a potential agonist for these receptors.…”

Section: Discussionmentioning

confidence: 99%

Galectin‐4, a novel neuronal regulator of myelination

Stancic

Slijepćević

Nomden

et al. 2012

Glia

View full text Add to dashboard Cite

Myelination of axons by oligodendrocytes (OLGs) is essential for proper saltatory nerve conduction, i.e., rapid transmission of nerve impulses. Among others, extracellular matrix (ECM) molecules, neuronal signaling, and axonal adhesion regulate the biogenesis and maintenance of myelin membranes, driven by polarized transport of myelin-specific proteins and lipids. Galectin-4, a tandem-repeat-type lectin with affinity to sulfatide and nonsialylated termini of N-glycans, has the ability to regulate adhesion of cells to ECM components and is also involved in polarized membrane trafficking. We, therefore, anticipated that galectin-4 might play a role in myelination. Here, we show that in developing postnatal rat brains galectin-4 expression is downregulated just before the onset of myelination. Intriguingly, when immature OLGs were treated with galectin-4, OLG maturation was retarded, while a subset of the immature OLGs reverted to a morphologically less complex progenitor stage, displaying concomitantly an increase in proliferation. Similarly, myelination was inhibited when galectin-4 or anti-galectin-4 antibodies were added to co-cultures of dorsal root ganglion neurons and OLGs. Neurons and OLGs were identified as a possible source of galectin-4, both in vitro and in vivo. In culture, neurons but not OLGs released galectin-4. Interestingly, in co-cultures, a reduced release of endogenous galectin-4 correlated with the onset of myelination. Moreover, galectin-4-reactive sites are transiently expressed on processes of premyelinating primary OLGs, but not on neurons. Taken together, these results identify neuronal galectin-4 as a candidate for a soluble regulator of OLG differentiation and, hence, myelination. © 2012 Wiley Periodicals, Inc.

show abstract

“…In particular, methods used for production and purification of secreted recombinant proteins could affect their biological activity, including their ability to interact with other proteins because of differences in posttranslational modifications introduced by artificial cell expression systems (23). In fact, LINGO-1 contains several N-glycosylation sites that could negatively and unpredictably affect its capacity for self-interaction either in cis or in trans (24,25), for interaction with its other partners, or with gangliosides that modulate those interactions (23,26,27). Instead, the use of gene expression vectors or transgenic animals to drive expression in primary cells would be expected to be less prone to this kind of modification in biological activity.…”

Section: Discussionmentioning

confidence: 99%

LINGO-1, a Transmembrane Signaling Protein, Inhibits Oligodendrocyte Differentiation and Myelination through Intercellular Self-interactions

Jepson

Vought

Gross

et al. 2012

Journal of Biological Chemistry

View full text Add to dashboard Cite

Background: Myelination of axons by oligodendrocytes is critical for neuronal function. LINGO-1, a cell receptor, inhibits myelination. Results: An extracellular fragment of LINGO-1 binds to LINGO-1 displayed on a cell surface and also inhibits oligodendrocyte maturation and myelination. Conclusion: LINGO-1 in axons and oligodendrocytes can self-interact to inhibit myelination. Significance: Understanding mechanisms governing myelination may lead to the treatment of demyelinating diseases like multiple sclerosis.

show abstract

Swift residue‐screening identifies key N‐glycosylated asparagines sufficient for surface expression of neuroglycoprotein Lingo‐1

Cited by 6 publications

References 15 publications

Targeting the cis‐dimerization of LINGO‐1 with low MW compounds affects its downstream signalling

Targeting the cis‐dimerization of LINGO‐1 with low MW compounds affects its downstream signalling

Galectin‐4, a novel neuronal regulator of myelination

LINGO-1, a Transmembrane Signaling Protein, Inhibits Oligodendrocyte Differentiation and Myelination through Intercellular Self-interactions

Contact Info

Product

Resources

About