Identification of α-Tocopherol succinate as an RFFL-substrate interaction inhibitor inducing peripheral CFTR stabilization and apoptosis

Taniguchi, Sachiho; Ono, Yuji; Doi, Yukako; Taniguchi, Shogo; Matsuura, Yuta; Iwasaki, Ayuka; Hirata, Noriaki; Fukuda, Ryosuke; Inoue, Keitaro; Yamaguchi, Miho; Tashiro, Anju; Egami, Daichi; Aoki, Shunsuke; Kondoh, Yasumitsu; Honda, Kaori; Osada, Hiroyuki; Kumeta, Hiroyuki; Saio, Tomohide; Okiyoneda, Tsukasa

doi:10.1016/j.bcp.2023.115730

Cited by 7 publications

(9 citation statements)

References 66 publications

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“…The following antibodies were used: mouse anti-HA (16B12, BioLegend, San Diego, CA, USA, Cat# 901515), anti-K48-Ub (Apu2 ZooMAb, Sigma-Aldrich, Cat# ZRB2150), anti-UBE3C (Abcepta, Inc., San Diego, CA, USA, Cat# AP20457c), anti-GST (clone 5A7, FUJIFILM Wako Pure Chemical Corporation, Cat# 013-21851), anti-Ub (P4D1, Santa Cruz Biotechnology, Santa Cruz, CA, USA, Cat# sc-8017), anti-DYKDDDDK (anti-FLAG, clone 1E6, FUJIFILM Wako Pure Chemical Corporation, Cat# 014-22383), anti-Myc (clone 9E10, JI-FILM Wako Pure Chemical Corporation, Cat# 017-21871), RNF185 antiserum (anti-RNF185) from rabbit [25], anti-HiBiT (Promega, Madison, WI, USA, Cat# N7200), Peroxidase AffiniPure Goat Anti-Mouse IgG (H+L) Secondary Antibody (Jackson Immuno Research, West Grove, PA, USA, Cat# 115-035-166), Peroxidase AffiniPure Donkey Anti-Rabbit IgG (H+L) Secondary Antibody (Jackson Immuno Research, Cat# 711-035-152).…”

Section: Reagents and Antibodiesmentioning

confidence: 99%

Section: Plasmidsmentioning

confidence: 99%

“…∆F508-CFTR-HiBiT(CT), N1303K-CFTR-HiBiT(CT), Insig-1-HiBiT(CT), ∆Y490-ABCB1-HiBiT(CT), and ∆F508-CFTR-Nluc(Ex) were constructed previously [23][24][25]. pCold-GST-4xTR-TUBE (GST-TUBE) was constructed by in-fusion cloning (Takara Bio, Kusatsu, Japan) using the pCold-GST plasmid [25] and pRSET-4xGST-TR-TUBE (Addgene #110312) as templates. pNUT-∆F508-CFTR-3HA [19] and pCold-His-sumo-USP21 (196-565) were kindly provided by Dr. Gergely Lukacs (McGill University) and Dr. Yusuke Sato (Tottori University), respectively.…”

Section: Plasmidsmentioning

confidence: 99%

“…pNUT-∆F508-CFTR-3HA [19] and pCold-His-sumo-USP21 (196-565) were kindly provided by Dr. Gergely Lukacs (McGill University) and Dr. Yusuke Sato (Tottori University), respectively. T70-CFTR-HiBiT(Ex) was constructed by replacing the 3xHA tag with the HiBiT tag in the 4th extracellular loop of CFTR as previously [25]. ∆Y490-ABCB1-HiBiT(Ex) was constructed by inserting the HiBiT tag in the 1st extracellular loop by PCR-based mutagenesis and inserted to pLX304 (Addgene #25890) via LR reaction using LR clonase II enzyme mix (ThermoFisher Scientific, Waltham, MA, USA).…”

Section: Plasmidsmentioning

confidence: 99%

“…BHK and BHK cells stably expressing HBH-∆F508-CFTR were cultured as previously [22]. BEAS-2B cells stably expressing ∆F508-CFTR-Nluc(Ex) [25], ∆F508-CFTR-Nluc(CT) [24], T70-CFTR-HiBiT(Ex), or ∆Y490-ABCB1-HiBiT(Ex) were grown in DMEM supplemented with 10% FBS, and 10 µg/mL blasticidin S. The parental BEAS-2B cells (Cat# 95102433) were obtained from the European Collection of Authenticated Cell Cultures (ECACC). Penicillin-streptomycin solution (FUJI-FILM Wako Pure Chemical Corporation) was added to all cell culture mediums.…”

Section: Cell Lines and Cell Culturementioning

confidence: 99%

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UBE3C Facilitates the ER-Associated and Peripheral Degradation of Misfolded CFTR

Kamada,

Tateishi,

Nakayamada

et al. 2023

Cells

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The ubiquitin E3 ligase UBE3C promotes the proteasomal degradation of cytosolic proteins and endoplasmic reticulum (ER) membrane proteins. UBE3C is proposed to function downstream of the RNF185/MBRL ER-associated degradation (ERAD) branch, contributing to the ERAD of select membrane proteins. Here, we report that UBE3C facilitates the ERAD of misfolded CFTR, even in the absence of both RNF185 and its functional ortholog RNF5 (RNF5/185). Unlike RNF5/185, UBE3C had a limited impact on the ubiquitination of misfolded CFTR. UBE3C knockdown (KD) resulted in an additional increase in the functional ∆F508-CFTR channels on the plasma membrane when combined with the RNF5/185 ablation, particularly in the presence of clinically used CFTR modulators. Interestingly, although UBE3C KD failed to attenuate the ERAD of insig-1, it reduced the ERAD of misfolded ∆Y490-ABCB1 and increased cell surface expression. UBE3C KD also stabilized the mature form of ∆F508-CFTR and increased the cell surface level of T70-CFTR, a class VI CFTR mutant. These results suggest that UBE3C plays a vital role in the ERAD of misfolded CFTR and ABCB1, even within the RNF5/185-independent ERAD pathway, and it may also be involved in maintaining the peripheral quality control of CFTR.

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Section: Reagents and Antibodiesmentioning

confidence: 99%

Section: Plasmidsmentioning

confidence: 99%

Section: Plasmidsmentioning

confidence: 99%

Section: Plasmidsmentioning

confidence: 99%

Section: Cell Lines and Cell Culturementioning

confidence: 99%

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UBE3C Facilitates the ER-Associated and Peripheral Degradation of Misfolded CFTR

Kamada,

Tateishi,

Nakayamada

et al. 2023

Cells

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Cystic Fibrosis Modulator Therapies: Bridging Insights from CF to other Membrane Protein Misfolding Diseases

Kim,

Plate

2024

Israel Journal of Chemistry

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Cystic Fibrosis (CF) is a genetic disorder resulting from mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene, leading to a faulty CFTR protein. Dysfunctional CFTR causes chloride ion imbalance, resulting in dense mucus accumulation in various organs, particularly the lungs. CF treatments focus on symptom management and addressing CFTR′s functional defects. Notably, development of CFTR modulator therapies has significantly advanced CF treatment. These drugs target CFTR protein structural defects induced by mutations, restoring its function and improving CF symptoms. VX‐770, a CFTR potentiator, and CFTR correctors like VX‐809, VX‐661, and VX‐445, have gained FDA approval and widespread clinical use, greatly enhancing the health and survival of many CF patients. However, some CFTR mutations lack effective targeted therapies, leaving approximately 6 % of CF patients without suitable options. CFTR modulator therapies have proven essential for combating the underlying causes of protein misfolding diseases, serving as a blueprint for similar treatments in other membrane protein misfolding diseases. This review explores current and future CFTR modulator therapies, and applications of established paradigms to membrane protein misfolding diseases. Ongoing research and innovation hold the potential for further improvements in CF management and the treatment of protein misfolding diseases.

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Targeting ubiquitination machinery in cystic fibrosis: Where do we stand?

Okiyoneda,

Borgo,

Bosello Travain

et al. 2024

Cell. Mol. Life Sci.

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Cystic Fibrosis (CF) is a genetic disease caused by mutations in CFTR gene expressing the anion selective channel CFTR located at the plasma membrane of different epithelial cells. The most commonly investigated variant causing CF is F508del. This mutation leads to structural defects in the CFTR protein, which are recognized by the endoplasmic reticulum (ER) quality control system. As a result, the protein is retained in the ER and degraded via the ubiquitin–proteasome pathway. Although blocking ubiquitination to stabilize the CFTR protein has long been considered a potential pharmacological approach in CF, progress in this area has been relatively slow. Currently, no compounds targeting this pathway have entered clinical trials for CF. On the other hand, the emergence of Orkambi initially, and notably the subsequent introduction of Trikafta/Kaftrio, have demonstrated the effectiveness of molecular chaperone-based therapies for patients carrying the F508del variant and even showed efficacy against other variants. These treatments directly target the CFTR variant protein without interfering with cell signaling pathways. This review discusses the limits and potential future of targeting protein ubiquitination in CF.

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Identification of α-Tocopherol succinate as an RFFL-substrate interaction inhibitor inducing peripheral CFTR stabilization and apoptosis

Cited by 7 publications

References 66 publications

UBE3C Facilitates the ER-Associated and Peripheral Degradation of Misfolded CFTR

UBE3C Facilitates the ER-Associated and Peripheral Degradation of Misfolded CFTR

Cystic Fibrosis Modulator Therapies: Bridging Insights from CF to other Membrane Protein Misfolding Diseases

Targeting ubiquitination machinery in cystic fibrosis: Where do we stand?

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