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Related Concept Videos

Export of Misfolded Proteins out of the ER01:32

Export of Misfolded Proteins out of the ER

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After folding, the ER assesses the quality of secretory and membrane proteins. The correctly folded proteins are cleared by the calnexin cycle for transport to their final destination, while misfolded proteins are held back in the ER lumen. The ER chaperones attempt to unfold and refold the misfolded proteins but sometimes fail to achieve the correct native conformation. Such terminally misfolded proteins are then exported to the cytosol by ER-associated degradation or ERAD pathway for...
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Protein Folding Quality Check in the RER01:29

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ER is the primary site for the maturation and folding of soluble and transmembrane secretory proteins. The calnexin cycle is a specific chaperone system that folds and assesses the confirmation of N-glycosylated proteins before they can exit the ER lumen. The primary players of this quality check pipeline are the lectins, ER-resident chaperones, and a glucosyl transferase enzyme. In case the calnexin system in the lumen fails to salvage a misfolded protein, it is transported to the cytoplasm...
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ER Retrieval Pathway01:45

ER Retrieval Pathway

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In the secretory pathway, vesicles transport proteins from one cellular compartment to another in forward transport to deliver the protein to its correct location. Occasionally, misfolded proteins and incorrect proteins escape their original compartments, and a retrieval pathway is used to return the escaped proteins to their original compartment.
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Oligosaccharide Assembly01:24

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Protein glycosylation starts in the ER lumen and continues in the Golgi apparatus. Glycosyltransferases catalyze the addition of sugar molecules or glycosylation of proteins. Usually, these enzymes add sugars to the hydroxyl groups of selected serine or threonine residues to form O-linked glycans or the amino groups of asparagine residues to form N-linked glycans. Different positions on the same polypeptide chain can contain differently linked glycans.
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GPI-anchoring is a post-translational, reversible protein modification that is ubiquitous in eukaryotes. Such proteins are primarily present on the exoplasmic leaflet of the plasma membrane.
GPI-anchor structure
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Regulation of the Unfolded Protein Response01:31

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Inositol-requiring kinase one or IRE1 is the most conserved eukaryotic unfolded protein response (UPR) receptor. It is a type I transmembrane protein kinase receptor with a distinctive site-specific RNase activity. As the binding mechanics of the misfolded proteins with the N-terminal domain of IRE-1 are unclear, three binding models — direct, indirect, and allosteric -- are proposed for receptor activation. Nevertheless, it is known that once a misfolded protein associates with IRE1, it...
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What Happens If a Human Galectin Enters the Endoplasmic Reticulum?

Tanja J Kutzner1, Alonso M Higuero2, Martina Süßmair3

  • 1Faculty of Veterinary Medicine, Institute of Physiological Chemistry, Ludwig-Maximilians-University Munich, Munich, Germany.

Methods in Molecular Biology (Clifton, N.J.)
|March 23, 2022
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Mammalian galectins, lacking a signal peptide, were engineered to enter the secretory pathway. This study investigates the fate of these modified galectins within the endoplasmic reticulum (ER) and Golgi apparatus.

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Area of Science:

  • Cell Biology
  • Molecular Biology
  • Glycobiology

Background:

  • Mammalian galectins are known to lack a signal peptide, a crucial element for protein export.
  • The intracellular fate and processing of galectins directed into the classical export pathway remain largely uncharacterized.
  • Understanding galectin trafficking is key to explaining their leaderless nature and synthesis on free ribosomes.

Purpose of the Study:

  • To investigate the consequences of directing mammalian galectins into the endoplasmic reticulum (ER) via signal peptide engineering.
  • To determine the fate of signal peptide-bearing galectin variants after their entry into the ER.
  • To elucidate the reasons behind the leaderless nature of galectins.

Main Methods:

  • Engineering of galectin-specific cDNA to include a signal peptide.
  • Affinity chromatography and mass-spectrometric analysis to assess N-glycosylation.
  • Cell-based binding and functional assays.
  • Subcellular fractionation via density gradient ultracentrifugation.
  • Immunocytochemical colocalization with ER/Golgi markers.

Main Results:

  • Analysis of N-glycosylation occupancy on potential sites.
  • Assessment of galectin binding and cellular functions in engineered cells.
  • Determination of subcellular localization within ER/Golgi compartments.
  • Identification of post-translational modifications and processing events.

Conclusions:

  • The study provides insights into the intracellular processing of galectins when artificially directed to the secretory pathway.
  • Findings contribute to understanding why endogenous galectins are leaderless and synthesized on free ribosomes.
  • This research clarifies the fate of signal peptide-bearing galectin variants within the ER and Golgi apparatus.