Selective inhibition of protein secretion by abrogating receptor-coat interactions during ER export

Gómez-Navarro, Natalia; Maldutyte, Julija; Poljak, Kristina; Peak‐Chew, Sew‐Yeu; Orme, Jonathon P.; Bisnett, Brittany; Lamb, Caitlin H.; Boyce, Michael; Gianni, Davide; Miller, Elizabeth A.

doi:10.1101/2022.01.31.478454

Cited by 3 publications

(2 citation statements)

References 40 publications

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“…The increased hepatocyte LDLR levels in Surf4 fl/fl Alb-Cre + mice are not accompanied by upregulation of Ldlr mRNA as measured by RNA-seq analysis, consistent with the increase in LDLR abundance being mediated by PCSK9 activity rather than an increase in gene expression. Furthermore, a recent report by Gomez- Navarro et al independently demonstrated that PCSK9 relies on both SEC24A and SURF4 for secretion and that chemical disruption of SEC24A-SURF4 interaction is sufficient to reduce PCSK9 secretion (Gomez-Navarro et al, 2022). Taken together, these data are consistent with the proposed function of SURF4 as a cargo receptor linking PCSK9 in the ER lumen to the SEC24A component of the COPII coat on the cytoplasmic face of the ER (Emmer et al, 2018).…”

Section: Discussionmentioning

confidence: 99%

Hepatic inactivation of murine Surf4 results in marked reduction in plasma cholesterol

Tang

McCormick

et al. 2022

Preprint

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PCSK9 negatively regulates low-density lipoprotein receptor (LDLR) abundance on the cell surface, leading to decreased hepatic clearance of LDL particles and increased levels of plasma cholesterol. We previously identified SURF4 as a cargo receptor that facilitates PCSK9 secretion in HEK293T cells (Emmer et al., 2018). Here, we generated hepatic SURF4-deficient mice (Surf4(fl/fl) Alb-Cre(+)) to investigate the physiologic role of SURF4 in vivo. Surf4(fl/fl) Alb-Cre(+) mice exhibited normal viability, gross development, and fertility. Plasma PCSK9 levels were reduced by ~60% in Surf4(fl/fl) Alb-Cre(+) mice, with a corresponding ~50% increase in steady state LDLR protein abundance in the liver, consistent with SURF4 functioning as a cargo receptor for PCSK9. Surprisingly, these mice exhibited a marked reduction in plasma cholesterol and triglyceride levels out of proportion to the partial increase in hepatic LDLR abundance. Detailed characterization of lipoprotein metabolism in these mice instead revealed a severe defect in hepatic lipoprotein secretion, consistent with prior reports of SURF4 also promoting the secretion of apolipoprotein B. Despite a small increase in liver mass and lipid content, histologic evaluation revealed no evidence of steatohepatitis or fibrosis in Surf4(fl/fl) Alb-Cre(+) mice. Acute depletion of hepatic SURF4 by CRISPR/Cas9 or liver-targeted siRNA in adult mice confirms these findings. Together, these data support the physiologic significance of SURF4 in the hepatic secretion of PCSK9 and APOB-containing lipoproteins and its potential as a therapeutic target in atherosclerotic cardiovascular diseases.

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Section: Discussionmentioning

confidence: 99%

Hepatic inactivation of murine Surf4 results in marked reduction in plasma cholesterol

Tang

McCormick

et al. 2022

Preprint

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“…However, to our knowledge, it has not been directly observed before in kinetic measurements of ER-to-Golgi transport. A recent study with several cargos demonstrated that 4-PBA selectively inhibited secretion to the medium of client cargos without effects on bulk-flow cargos (39).…”

Section: Alg-2 Activation Increases Copii Sorting Stringency Through ...mentioning

confidence: 99%

Steady-State Regulation of COPII-Dependent Secretory Cargo Sorting by Inositol Trisphosphate Receptors, Calcium, and Penta EF Hand Proteins

Held

Hojanazarova

Sargeant

et al. 2020

Preprint

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ER Ca2+ regulates ER-to-Golgi transport machinery. Sustained Ca2+ signaling by inositol trisphosphate receptors (IP3Rs) leads to depression of cargo export through activation of penta EF hand protein (PEF) ALG-2 which reduces outer COPII coat at ER exit sites (ERES). However, we do not know whether tonic Ca2+ signals during steady-state conditions affect ER export rates and if so by what mechanisms. Here we report that partial depletion of IP3 receptors from NRK epithelial cells causes a marked increase of basal ER export of the transmembrane glycoprotein cargo VSV-G. The increased ER-to-Golgi transport required ALG-2 and was actuated by decreased peflin and increased ALG-2 at ER exit sites (ERES) – a condition previously demonstrated to stimulate COPII-dependent ER export. Upon IP3R depletion the amount of outer coat at ERES increased, the opposite to what occurs during ALG-2-dependent inhibition of secretion during agonist-driven Ca2+ signaling. The increased ER export correlated with reduced spontaneous cytosolic Ca2+ oscillations caused by the reduced number of Ca2+ release channels. IP3R depletion also unexpectedly resulted in partial depletion of ER luminal Ca2+ stores. The low Ca2+ conditions appeared to decrease both ALG-2 and peflin expression levels somewhat, but these were the only detectable expression changes in COPII trafficking machinery and the Ca2+ decrease had no detectable impact on ER stress. We conclude that at steady state, IP3Rs produce tonic Ca2+ signals that suppress the basal rate of ER export by maintaining lower outer coat targeting to ERES.

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Quantification of the Chemical Chaperone 4-Phenylbutyric Acid (4-PBA) in Cell Culture Media via LC-HRMS: Applications in Fields of Neurodegeneration and Cancer

et al. 2023

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In recent years, 4-phenylbutyric acid (4-PBA), an FDA-approved drug, has increasingly been used as a nonspecific chemical chaperone in vitro and in vitro, but its pharmacodynamics is still not clear. In this context, we developed and validated a Liquid Chromatography–High Resolution Mass Spectrometry (LC-HRMS) method to quantify 4-PBA in NeuroBasal-A and Dulbecco’s Modified Eagle widely used cell culture media. Samples were injected on a Luna® 3 µm PFP(2) 100 Å (100 × 2.0 mm) column maintained at 40 °C. Water and methanol both with 0.1% formic acid served as mobile phases in a step gradient mode. The mass acquisition was performed by selected ion monitoring (SIM) in negative mode for a total run time of 10.5 min at a flow rate of 0.300 mL/min. The analogue 4-(4-Nitrophenyl)-Butyric Acid served as internal standard. Validation parameters were verified according to FDA and EMA guidelines. The quantification ranges from 0.38–24 µM. Inter and intraday RSDs (Relative Standard Deviations) were within 15%. The developed LC-HRMS method allowed the estimation of 4-PBA absorption and adsorption kinetics in vitro in two experimental systems: (i) 4-PBA improvement of protein synthesis in an Alzheimer’s disease astrocytic cell model; and (ii) 4-PBA reduction of endoplasmic reticulum stress in thapsigargin-treated melanoma cell lines.

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Selective inhibition of protein secretion by abrogating receptor-coat interactions during ER export

Cited by 3 publications

References 40 publications

Hepatic inactivation of murine Surf4 results in marked reduction in plasma cholesterol

Hepatic inactivation of murine Surf4 results in marked reduction in plasma cholesterol

Steady-State Regulation of COPII-Dependent Secretory Cargo Sorting by Inositol Trisphosphate Receptors, Calcium, and Penta EF Hand Proteins

Quantification of the Chemical Chaperone 4-Phenylbutyric Acid (4-PBA) in Cell Culture Media via LC-HRMS: Applications in Fields of Neurodegeneration and Cancer

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