Molecular basis for variations in the sensitivity of pathogenic rhodopsin variants to 9-cis-retinal

Roushar, Francis J.; McKee, Andrew G; Kuntz, Charles P.; Ortega, Joseph T.; Penn, Wesley D.; Woods, H. J.; Chamness, Laura M.; Most, Victoria; Meiler, Jens; Jastrzębska, Beata; Schlebach, Jonathan P.

doi:10.1016/j.jbc.2022.102266

Cited by 7 publications

(8 citation statements)

References 46 publications

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“…3A). This observation, which echoes our recent findings on the pharmacochaperone-mediated rescue of misfolded rhodopsin variants, 16 implies that the ability of these compounds to re-balance the folding equilibrium and prevent off-pathway interactions is fundamentally limited, in part, by their binding energy. Our modeling efforts reveal that VX-661 binding generally enforces a common subset of native contacts and provides a similar degree of stabilization for a variety of variants with divergent properties (Fig.…”

Section: Discussionsupporting

confidence: 86%

“…4A, 5A, and 7A). 16 Many variants with intermediate PME exhibit increases in intensity that approach their upper bound in the presence of VX-661 (Fig. 4A), suggesting their shifts in the folding equilibrium are effectively offset by its binding energy.…”

Section: Discussionmentioning

confidence: 99%

“…The increased “growth potential” of CF variants may reflect the intrinsic instability of the WT channel-CFTR exhibits ~30-fold lower PME relative to rhodopsin under similar conditions when adjusted for the number of epitopes. 16 Based on this consideration, we suspect that large, poorly expressed multi-domain proteins may generally constitute favorable targets for correctors given their relative abundance of potential binding sites and low intrinsic folding efficiencies, which render PME highly tunable.…”

Section: Discussionmentioning

confidence: 99%

“…A series of simplifying assumptions was used to derive thermodynamic boundaries for the change in CFTR variant surface immunostaining intensities in the presence of correctors as previously described. 16 Briefly, we first assumed surface immunostaining intensities to be proportional to the equilibrium fraction of folded protein ( f fold ) of CFTR variants. The lowest observed variant immunostaining intensity under each condition was taken as an estimate for the immunostaining intensity of cells expressing variants with an f fold ~ 0.…”

Section: Methodsmentioning

confidence: 99%

“…To survey the effects of these mutations on CFTR PME, we measured their surface immunostaining in parallel by DMS as previously described. [14][15][16] Briefly, we first used our genetic library to generate a pool of recombinant HEK293T cells that each inducibly express a single CFTR variant from a defined genomic locus. The surface immunostaining profile of these recombinant cells reveals that ~30% express variants with comparable immunostaining to WT CFTR (Int.…”

Section: Plasma Membrane Expression Of Rare Cystic Fibrosis Variantsmentioning

confidence: 99%

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Elucidation of Global Trends in the Effects of VX-661 and VX-445 on the Expression of Clinical CFTR Variants

McKee

McDonald

Penn

et al. 2022

Preprint

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Cystic fibrosis (CF) is a chronic genetic disease caused by mutations that compromise the expression and/ or function of the cystic fibrosis transmembrane conductance regulator chloride channel (CFTR). Most people with CF harbor a common misfolded CFTR variant (delF508), which can be rescued by combination therapies containing corrector compounds that restore its expression. Nevertheless, there are over 400 other CF variants that differ in their sensitivity to correctors for reasons that remain unclear. In this work, we utilize deep mutational scanning to quantitatively compare the effects of two FDA-approved correctors on the plasma membrane expression of 129 known CF variants, including 45 that are currently unclassified. Across 67 variants with attenuated expression, we find that VX-661-sensitive variants generally exhibit intermediate expression and feature mutations near its binding pocket within the first membrane spanning domain (MSD1). VX-445 also rescues variants with intermediate expression, though it is uniquely effective for certain mutations that perturb the later stages of CFTR assembly. Structural calculations suggest VX-661 provides similar stabilization to both sensitive and insensitive variants alike. These findings collectively suggest the mutation-specific effects of these compounds depend on the degree of variant destabilization and/ or the timing of cotranslational folding defects. Combining these correctors synergistically rescues variants with deficient and intermediate expression alike, presumably by doubling the total binding energy and suppressing defects at different stages of translation. These results provide an unprecedented overview of the properties of rare CFTR variants and establish new tools for CF pharmacology.

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Section: Discussionsupporting

confidence: 86%

Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Plasma Membrane Expression Of Rare Cystic Fibrosis Variantsmentioning

confidence: 99%

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