Differential effects of Best disease causing missense mutations on bestrophin-1 trafficking.

Abstract

Mutations in bestrophin-1 (Best1) cause Best vitelliform macular dystrophy (BVMD), a dominantly inherited retinal degenerative disease. Best1 is a homo-oligomeric anion channel localized to the basolateral surface of retinal pigment epithelial (RPE) cells. A number of Best1 mutants mislocalize in Madin-Darby canine kidney (MDCK) cells. However, many proteins traffic differently in MDCK and RPE cells, and MDCK cells do not express endogenous Best1. Thus, effects of Best1 mutations on localization in MDCK cells may not translate to RPE cells. To determine whether BVMD causing mutations affect Best1 localization, we compared localization and oligomerization of Best1 with Best1 mutants V9M, W93C, and R218C. In MDCK cells, Best1 and Best1(R218C) were basolaterally localized. Best1(W93C) and Best1(V9M) accumulated in cells. In cultured fetal human retinal pigment epithelium cells (fhRPE) expressing endogenous Best1, Best1(R218C) and Best1(W93C) were basolateral. Best1(V9M) was intracellular. All three mutants exhibited similar fluorescence resonance energy transfer (FRET) efficiencies to, and co-immunoprecipitated with Best1, indicating unimpaired oligomerization. When human Best1 was expressed in RPE in mouse eyes it was basolaterally localized. However, Best1(V9M) accumulated in intracellular compartments in mouse RPE. Co-expression of Best1 and Best1(W93C) in MDCK cells resulted in basolateral localization of both Best1 and Best1(W93C), but co-expression of Best1 with Best1(V9M) resulted in mislocalization of both proteins. We conclude that different mutations in Best1 cause differential effects on its localization and that this effect varies with the presence or absence of wild-type (WT) Best1. Furthermore, MDCK cells can substitute for RPE when examining the effects of BVMD causing mutations on Best1 localization if co-expressed with WT Best1.

• Authors: Adiv A Johnson, Yong-Suk Lee, J Brett Stanton, Kuai Yu, Criss H Hartzell, Lihua Y Marmorstein, Alan D Marmorstein
• Journal: Human molecular genetics (Hum Mol Genet) Vol. 22 Issue 23 Pg. 4688-97 (Dec 01 2013) ISSN: 1460-2083 [Electronic] England
• PMID: 23825107 (Publication Type: Journal Article, Research Support, N.I.H., Extramural, Research Support, Non-U.S. Gov't)

Chemical References

• BEST1 protein, human
• Best1 protein, mouse
• Bestrophins
• Chloride Channels
• Eye Proteins
• Ion Channels
• Mutant Proteins

Topics

• Animals
• Bestrophins
• Calcium Signaling
• Cell Membrane (metabolism)
• Cells, Cultured
• Chloride Channels (genetics, metabolism)
• Eye (metabolism)
• Eye Proteins (genetics, metabolism)
• Gene Expression Regulation
• Humans
• Ion Channels (genetics, metabolism)
• Mice
• Mice, Inbred BALB C
• Mice, Transgenic
• Mutagenesis, Site-Directed
• Mutant Proteins (genetics, metabolism)
• Mutation, Missense
• Protein Transport (genetics)
• Vitelliform Macular Dystrophy (genetics, pathology)