ArticleInvestigative ophthalmology & visual science2016
A Novel Luciferase Assay For Sensitively Monitoring Myocilin Variants in Cell Culture.
Article in Investigative ophthalmology & visual science, 2016. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 13 papers.
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Who cites it
13 citing papers in PubMed, 17 citations in OpenAlex.
- Monitoring mutant myocilin secretion and localization in trabecular meshwork cell cultures using a protein complementation-based luminescence assay.Experimental eye research · 2025Article
- Monitoring mutant myocilin secretion and localization in trabecular meshwork cell cultures using a protein complementation-based luminescence assay.bioRxiv : the preprint server for biology · 2025Article
- Protein misfolding and mitochondrial dysfunction in glaucoma.Frontiers in cell and developmental biology · 2025Review
- Quantitative differentiation of benign and misfolded glaucoma-causing myocilin variants on the basis of protein thermal stability.Disease models & mechanisms · 2023Article
- Specifications of the ACMG/AMP variant curation guidelines for myocilin: Recommendations from the clingen glaucoma expert panel.Human mutation · 2022Article
- A Simple Secretion Assay for Assessing New and Existing Myocilin Variants.Current eye research · 2022Article
- Common and rare myocilin variants: Predicting glaucoma pathogenicity based on genetics, clinical, and laboratory misfolding data.Human mutation · 2021Review
- Clinically-identified C-terminal mutations in fibulin-3 are prone to misfolding and destabilization.Scientific reports · 2021Article
- Article
- Non-antibiotic Small-Molecule Regulation of DHFR-Based Destabilizing DomainsMolecular therapy. Methods & clinical development · 2019Article
- Physiological function of myocilin and its role in the pathogenesis of glaucoma in the trabecular meshwork (Review).International journal of molecular medicine · 2019Review
- Endoplasmic Reticulum Stress Response of Trabecular Meshwork Stem Cells and Trabecular Meshwork Cells and Protective Effects of Activated PERK Pathway.Investigative ophthalmology & visual science · 2019Article
- A High-Throughput Assay for Collagen Secretion Suggests an Unanticipated Role for Hsp90 in Collagen Production.Biochemistry · 2018Article
Corrections and comments
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Authors and funding
4 authors at 2 institutions in 1 country.
Funding
Abstract
purposePrimary open angle glaucoma-associated mutations in myocilin (MYOC) cause protein "nonsecretion," rendering secreted MYOC difficult to detect using conventional techniques. This study focused on developing and using an assay that can quickly and easily detect mutant MYOC secretion.
methodsWe fused Gaussia luciferase (eGLuc2) to MYOC variants and expressed the constructs in HEK-293T and NTM-5 cells. Secreted and intracellular levels of MYOC eGLuc2 variants were evaluated by Western blotting and compared to untagged and FLAG-tagged MYOC constructs. Secreted and soluble intracellular MYOC eGLuc2 were measured by a GLuc assay. The secretion of nine additional MYOC mutants was assayed in conditioned media from transfected cells to test the applicability of the assay for monitoring other MYOC variants.
resultsMyocilin eGLuc2 behaved similarly to untagged and FLAG-tagged MYOC with respect to secretion, soluble intracellular levels, and in response to drug treatment. The GLuc assay could sensitively detect Y437H MYOC secretion 30 minutes after media change. Gaussia luciferase fused variants followed anticipated trends; nonpathogenic variants (D208E, G244V) were secreted at wild-type (WT) levels, whereas predicted disease-causing variants (C245Y, G246R, E300K, Y437H, I477N) demonstrated substantial secretion defects. Secretion defects caused by the C245Y, G246R, and Y437H mutations were partially rescued by permissive growth temperature. Interestingly, however, this increase in secretion was independent of newly synthesized protein.
conclusionsFusion of eGLuc2 to MYOC does not significantly change the behavior of MYOC. This newly developed MYOC reporter system can be used to study engineered MYOC variants and potentially to identify modulators of MYOC secretion and function.
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Read under generation 80e0d062 · epoch 390. Bibliography from PubMed, PubMed Central and OpenAlex; grants from NIH RePORTER; trial links from ClinicalTrials.gov; estimates, votes and beliefs from the OpenQuestion graph.