scholarly journals AAV-mediated ERdj5 overexpression protects against P23H rhodopsin toxicity

2020 ◽  
Vol 29 (8) ◽  
pp. 1310-1318 ◽  
Author(s):  
Monica Aguilà ◽  
James Bellingham ◽  
Dimitra Athanasiou ◽  
Dalila Bevilacqua ◽  
Yanai Duran ◽  
...  
Keyword(s):  
Quality Control ◽  
Outer Nuclear Layer ◽  
Photoreceptor Cells ◽  
Er Quality Control ◽  
Full Field ◽  
Er Retention ◽  
Function Loss

Abstract Rhodopsin misfolding caused by the P23H mutation is a major cause of autosomal dominant retinitis pigmentosa (adRP). To date, there are no effective treatments for adRP. The BiP co-chaperone and reductase ERdj5 (DNAJC10) is part of the endoplasmic reticulum (ER) quality control machinery, and previous studies have shown that overexpression of ERdj5 in vitro enhanced the degradation of P23H rhodopsin, whereas knockdown of ERdj5 increased P23H rhodopsin ER retention and aggregation. Here, we investigated the role of ERdj5 in photoreceptor homeostasis in vivo by using an Erdj5 knockout mouse crossed with the P23H knock-in mouse and by adeno-associated viral (AAV) vector-mediated gene augmentation of ERdj5 in P23H-3 rats. Electroretinogram (ERG) and optical coherence tomography of Erdj5−/− and P23H+/−:Erdj5−/− mice showed no effect of ERdj5 ablation on retinal function or photoreceptor survival. Rhodopsin levels and localization were similar to those of control animals at a range of time points. By contrast, when AAV2/8-ERdj5-HA was subretinally injected into P23H-3 rats, analysis of the full-field ERG suggested that overexpression of ERdj5 reduced visual function loss 10 weeks post-injection (PI). This correlated with a significant preservation of photoreceptor cells at 4 and 10 weeks PI. Assessment of the outer nuclear layer (ONL) morphology showed preserved ONL thickness and reduced rhodopsin retention in the ONL in the injected superior retina. Overall, these data suggest that manipulation of the ER quality control and ER-associated degradation factors to promote mutant protein degradation could be beneficial for the treatment of adRP caused by mutant rhodopsin.

Anchoring of FLT3 in the endoplasmic reticulum alters signaling quality

Blood ◽  
2009 ◽  
Vol 113 (15) ◽  
pp. 3568-3576 ◽  
Author(s):  
Dirk Schmidt-Arras ◽  
Sylvia-Annette Böhmer ◽  
Sina Koch ◽  
Jörg P. Müller ◽  
Lutz Blei ◽  
...  
Keyword(s):  
Endoplasmic Reticulum ◽  
Cell Transformation ◽  
Intracellular Location ◽  
Er Retention ◽  
Transforming Activity ◽  
Signaling Quality

Abstract The mechanism of cell transformation by Fms-like tyrosine kinase 3 (FLT3) in acute myeloid leukemia (AML) is incompletely understood. The most prevalent activated mutant FLT3 ITD exhibits an altered signaling quality, including strong activation of the STAT5 transcription factor. FLT3 ITD has also been found partially retained as a high-mannose precursor in an intracellular compartment. To analyze the role of intracellular retention of FLT3 for transformation, we have generated FLT3 versions that are anchored in the perinuclear endoplasmic reticulum (ER) by appending an ER retention sequence containing a RRR (R3) motif. ER retention of R3, but not of corresponding A3 FLT3 versions, is shown by biochemical, fluorescence-activated cell sorting, and immunocytochemical analyses. ER anchoring reduced global autophosphorylation and diminished constitutive activation of ERK1/2 and AKT of the constitutively active FLT3 versions. ER anchoring was, however, associated with elevated signaling to STAT3. Transforming activity of the FLT3 D835Y mutant was suppressed by ER anchoring. In contrast, ER-anchored FLT3 ITD retained STAT5-activating capacity and was transforming in vitro and in vivo. The findings highlight another aspect of the different signaling quality of FLT3 ITD: It can transform cells from an intracellular location.

scholarly journals Circular RNA CREBBP Suppresses Hepatic Fibrosis Via Targeting the hsa-miR-1291/LEFTY2 Axis

2021 ◽  
Vol 12 ◽  
Author(s):  
Ya-Ru Yang ◽  
Shuang Hu ◽  
Fang-Tian Bu ◽  
Hao Li ◽  
Cheng Huang ◽  
...  
Keyword(s):  
Hepatic Fibrosis ◽  
Circular Rna ◽  
Vehicle Group ◽  
Potential Biomarker ◽  
Function Loss ◽  
And Function ◽  
Pro Inflammatory Cytokines

CircRNAs (circRNAs) are commonly dysregulated in a variety of human diseases and are involved in the development and progression of cancer. However, the role of circRNAs in hepatic fibrosis (HF) is still unclear. Our previous high throughput screen revealed changes in many circRNAs in mice with carbon tetrachloride (CCl4)-induced HF. For example, circCREBBP was significantly down-regulated in primary hepatic stellate cells (HSCs) and liver tissue of HF mice induced by CCl4 compared to those in the vehicle group. Overexpression of circCREBBP with AAV8-circCREBBP in vivo prevented CCl4-induced HF worsening by reducing serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) contents, liver hydroxyproline levels, collagen deposition, and levels of pro-fibrosis genes and pro-inflammatory cytokines. Furthermore, in vitro function loss and function gain analysis showed that circCREBBP inhibited HSCs activation and proliferation. Mechanically, circCREBBP acts as a sponge for hsa-miR-1291 and subsequently promotes LEFTY2 expression. In conclusion, our current results reveal a novel mechanism by which circCREBBP alleviates liver fibrosis by targeting the hsa-miR-1291/LEFTY2 axis, and also suggest that circCREBBP may be a potential biomarker for heart failure.

scholarly journals Fer exacerbates renal fibrosis and can be targeted by miR-29c-3p

Open Medicine ◽  
2021 ◽  
Vol 16 (1) ◽  
pp. 1378-1385
Author(s):  
Chen-Min Sun ◽  
Wen-Yi Zhang ◽  
Shu-Yan Wang ◽  
Gang Qian ◽  
Dong-Liang Pei ◽  
...  
Keyword(s):  
Renal Fibrosis ◽  
Target Gene ◽  
Rat Kidney ◽  
Inflammatory Factors ◽  
Strongly Correlated ◽  
Tnf Α ◽  
Function Loss

Abstract Aim Renal fibrosis (RF) is a common clinical condition leading to irreversible renal function loss. Tyrosine kinase proteins and microRNAs (miRs) are associated with pathogenesis and we aim to investigate the role of Fer and its partner miR(s) in RF. Method In silico reproduction of Mouse Kidney FibrOmics browser was performed to identify potential miR(s) and target gene(s). In vivo validation was performed in C57BL/6 mice with unilateral ureteral obstruction (UUO). In vitro validation was performed in rat kidney fibroblast NRK-49F cells. Mimics and inhibitors of miR-29c-3p were constructed. The target gene Fer was monitored by RT-PCR and western blotting. The levels of interleukin (IL)-6, IL-1β, and tumor necrosis factor (TNF)-α in serum and media were measured by ELISA. Results The Fer expression and protein level were gradually increased during 14 days of UUO modeling. miR-29c-3p expression was strongly correlated with that of Fer. In vivo validation showed increased expressions of fibrosis-associated genes and increased phospoho-Smad3 level in the UUO model. Fer-knockdown (KD) significantly decreased expressions of fibrosis-associated genes. Pharmaceutical inhibition of Fer showed similar effects to miR-29c-3p, and miR inhibition showed a significant decrease of excretion of inflammatory factors. Conclusion Dysregulation of miR-29c-3p and Fer plays a role in RF. Pharmaceutical or genetic inhibition of Fer may serve as the potential treatment for RF.

scholarly journals The Reverse Side of a Coin: “Factor-Free” Ribosomal Protein Synthesis In Vitro is a Consequence of the In Vivo Proofreading Mechanism

Biomolecules ◽  
2019 ◽  
Vol 9 (10) ◽  
pp. 588
Author(s):  
Finkelstein
Keyword(s):  
Quality Control ◽  
Protein Synthesis ◽  
Amino Acid ◽  
Amino Acid Sequence ◽  
Ribosomal Protein ◽  
Elongation Factor ◽  
Ribosomal Protein Synthesis

This paper elucidates a close connection between two well-known facts that until now have seemed independent: (i) the quality control (“proofreading”) of the emerging amino acid sequence, occurring during the normal, elongation-factor-dependent ribosomal biosynthesis, which is performed by removing those Aa-tRNAs (aminoacyl tRNAs) whose anticodons are not complementary to the exhibited mRNA codons, and (ii) the in vitro discovered existence of the factor-free ribosomal synthesis of polypeptides. It is shown that a biological role of proofreading is played by a process that is exactly opposite to the step of factor-free binding of Aa-tRNA to the ribosome-exposed mRNA: a factor-free removal of that Aa-tRNA whose anticodon is not complementary to the ribosome-exhibited mRNA codon.

scholarly journals Endoplasmic Reticulum Quality Control of Unassembled Iron Transporter Depends on Rer1p-mediated Retrieval from the Golgi

2004 ◽  
Vol 15 (3) ◽  
pp. 1417-1424 ◽  
Author(s):  
Miyuki Sato ◽  
Ken Sato ◽  
Akihiko Nakano
Keyword(s):  
Quality Control ◽  
Endoplasmic Reticulum ◽  
Membrane Proteins ◽  
Model System ◽  
Er Quality Control ◽  
Iron Transporter ◽  
Er Retention ◽  
Endoplasmic Reticulum Quality Control ◽  
Sorting Receptor

Endoplasmic reticulum (ER) quality control is a conserved process by which misfolded or unassembled proteins are selectively retained in the endoplasmic reticulum (ER). Failure in oligomerization of multisubunit membrane proteins is one of the events that triggers ER quality control. The transmembrane domains (TMDs) of unassembled subunits are determinants of ER retention in many cases, although the mechanism of the TMD-mediated sorting of unassembled subunits remains elusive. We studied a yeast iron transporter complex on the cell surface as a new model system for ER quality control. When Fet3p, a transmembrane subunit, is not assembled with the other membrane subunit, Ftr1p, unassembled Fet3p is exclusively localized to the ER at steady state. The TMD of Fet3p contains a determinant for this process. However, pulse-chase analysis and in vitro budding assays indicate that unassembled Fet3p rapidly escapes from the ER. Furthermore, Rer1p, a retrieval receptor for ER-resident membrane proteins in the Golgi, is responsible for the TMD-dependent ER retrieval of unassembled Fet3p. These findings provide clear evidence that the ER quality control of unassembled membrane proteins can be achieved by retrieval from the Golgi and that Rer1p serves as a specific sorting receptor in this process.

scholarly journals The role of UDP-Glc:glycoprotein glucosyltransferase 1 in the maturation of an obligate substrate prosaposin

2010 ◽  
Vol 189 (5) ◽  
pp. 829-841 ◽  
Author(s):  
Bradley R. Pearse ◽  
Taku Tamura ◽  
Johan C. Sunryd ◽  
Gregory A. Grabowski ◽  
Randal J. Kaufman ◽  
...  
Keyword(s):  
Quality Control ◽  
Molecular Chaperones ◽  
Control Factor ◽  
Dramatic Decrease ◽  
Er Quality Control ◽  
Basal Expression ◽  
Er Retention ◽  
Chaperone Binding ◽  
Quality Control Test

An endoplasmic reticulum (ER) quality control system assists in efficient folding and disposal of misfolded proteins. N-linked glycans are critical in these events because their composition dictates interactions with molecular chaperones. UDP-glucose:glycoprotein glucosyltransferase 1 (UGT1) is a key quality control factor of the ER. It adds glucoses to N-linked glycans of nonglucosylated substrates that fail a quality control test, supporting additional rounds of chaperone binding and ER retention. How UGT1 functions in its native environment is poorly understood. The role of UGT1 in the maturation of glycoproteins at basal expression levels was analyzed. Prosaposin was identified as a prominent endogenous UGT1 substrate. A dramatic decrease in the secretion of prosaposin was observed in ugt1−/− cells with prosaposin localized to large juxtanuclear aggresome-like inclusions, which is indicative of its misfolding and the essential role that UGT1 plays in its proper maturation. A model is proposed that explains how UGT1 may aid in the folding of sequential domain–containing proteins such as prosaposin.

scholarly journals A cell-based reglucosylation assay demonstrates the role of GT1 in the quality control of a maturing glycoprotein

2008 ◽  
Vol 181 (2) ◽  
pp. 309-320 ◽  
Author(s):  
Bradley R. Pearse ◽  
Luke Gabriel ◽  
Ning Wang ◽  
Daniel N. Hebert
Keyword(s):  
Quality Control ◽  
Control Test ◽  
Er Quality Control ◽  
Influenza Hemagglutinin ◽  
Er Retention ◽  
Nascent Chain ◽  
A Cell ◽  
Polypeptide Backbone ◽  
Quality Control Test

The endoplasmic reticulum (ER) protein GT1 (UDP-glucose: glycoprotein glucosyltransferase) is the central enzyme that modifies N-linked carbohydrates based upon the properties of the polypeptide backbone of the maturing substrate. GT1 adds glucose residues to nonglucosylated proteins that fail the quality control test, supporting ER retention through persistent binding to the lectin chaperones calnexin and calreticulin. How GT1 functions in its native environment on a maturing substrate is poorly understood. We analyzed the reglucosylation of a maturing model glycoprotein, influenza hemagglutinin (HA), in the intact mammalian ER. GT1 reglucosylated N-linked glycans in the slow-folding stem domain of HA once the nascent chain was released from the ribosome. Maturation mutants that disrupted the oxidation or oligomerization of HA also supported region-specific reglucosylation by GT1. Therefore, GT1 acts as an ER quality control sensor by posttranslationally reglucosylating glycans on slow-folding or nonnative domains to recruit chaperones specifically to critical aberrant regions.

The Role of Polyphenols in the Modulation of Plasma Non-Enzymatic Antioxidant Capacity (NEAC)

2012 ◽  
Vol 82 (3) ◽  
pp. 228-232 ◽  
Author(s):  
Mauro Serafini ◽  
Giuseppa Morabito
Keyword(s):  
Antioxidant Capacity ◽  
Dietary Intervention ◽  
Ex Vivo ◽  
The Body ◽  
Dietary Polyphenols ◽  
Enzymatic Antioxidant ◽  
Endogenous Antioxidant

Dietary polyphenols have been shown to scavenge free radicals, modulating cellular redox transcription factors in different in vitro and ex vivo models. Dietary intervention studies have shown that consumption of plant foods modulates plasma Non-Enzymatic Antioxidant Capacity (NEAC), a biomarker of the endogenous antioxidant network, in human subjects. However, the identification of the molecules responsible for this effect are yet to be obtained and evidences of an antioxidant in vivo action of polyphenols are conflicting. There is a clear discrepancy between polyphenols (PP) concentration in body fluids and the extent of increase of plasma NEAC. The low degree of absorption and the extensive metabolism of PP within the body have raised questions about their contribution to the endogenous antioxidant network. This work will discuss the role of polyphenols from galenic preparation, food extracts, and selected dietary sources as modulators of plasma NEAC in humans.

Anti-obesity role of Aster glehni extract: in vivo and in vitro effects

Planta Medica ◽  
2012 ◽  
Vol 78 (11) ◽  
Author(s):  
HM Lee ◽  
TG Ahn ◽  
CW Kim ◽  
HJ An
Keyword(s):  
In Vitro Effects
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