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Related Concept Videos

Rab Proteins01:14

Rab Proteins

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Rab proteins constitute the largest family of monomeric GTPases, of which 70 members are present in humans. Rab proteins and their effectors regulate consecutive stages of vesicle transport such as vesicle transport, docking, and fusion to the correct recipient membrane.
Rab proteins switch between a cytosolic, GDP-bound inactive state and a membrane-anchored, GTP-bound active state. By themselves, Rabs show slow rates of GDP/GTP exchange and GTP hydrolysis. Thus, Rab proteins are considered...
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Rab Cascades01:25

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Rab GTPases act in a regulated cascade during membrane fusion, helping the lipid bilayers mix. The Rab family of proteins are active when bound to GTP, and inactive when bound to GDP. Hence, they act as guanine nucleotide-dependent molecular switches. Rab-GTP recognizes and binds to long or short-range tethering proteins to capture the target vesicle. These tethers coordinate with SNAREs on the vesicle and the target membrane to assemble the trans SNARE complex that locks the mixing bilayers.
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Regulation of Nuclear Protein Sorting01:45

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Nuclear protein sorting regulates nucleus composition and gene expression, crucial for determining the fate of a eukaryotic cell. Hence, the entry and exit of molecules across the nuclear envelope is a tightly controlled process. Nuclear protein sorting can be inhibited by one of the following ways: 1) masking cargo signal sequences, 2) modifying the nuclear receptor's affinity for cargo, 3) controlling the nuclear pore size, 4) retaining the cargo during its transit to the cytosol or the...
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Nuclear Export01:42

Nuclear Export

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The nucleus restricts several proteins within and allows others to pass. The restricted proteins possess a nuclear retention sequence or NRS, anchoring them to the nuclear lamins and preventing their transport to the cytosol. The non-restricted proteins, after their synthesis, are transported to their site of action, such as the cytosol or other organelles, with the help of nuclear export signals or NES.
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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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Recycling Endosomes and Transcytosis00:58

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The recycling endosome, also known as the endosomal recycling compartment (ERC), is a part of the slow-recycling process of the endocytic pathway. Molecules internalized through receptor-mediated endocytosis are either degraded in the lysosomes or are recycled to the plasma membrane through the fast- or slow-recycling route.
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The Microscopy-Based Assay to Study and Analyze the Recycling Endosomes using SNARE Trafficking
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A trimeric Rab7 GEF controls NPC1-dependent lysosomal cholesterol export.

Dick J H van den Boomen1, Agata Sienkiewicz2, Ilana Berlin3

  • 1Cambridge Institute of Therapeutic Immunology & Infectious Disease, University of Cambridge, Cambridge, UK. djhv2@cam.ac.uk.

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|November 4, 2020
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Researchers identified over 100 genes regulating cholesterol import. C18orf8 is crucial for Rab7 activation, endosomal LDL transport, and lysosomal cholesterol export, maintaining cellular cholesterol balance.

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Area of Science:

  • Cell Biology
  • Molecular Biology
  • Genetics

Background:

  • Cholesterol import in mammalian cells primarily occurs via the low-density lipoprotein (LDL) receptor pathway.
  • Understanding the intricate mechanisms governing cholesterol trafficking is crucial for cellular homeostasis and disease prevention.

Purpose of the Study:

  • To identify novel genes involved in LDL-cholesterol import using a genome-wide CRISPR screen.
  • To characterize the function of C18orf8 in cellular cholesterol metabolism and trafficking.
  • To elucidate the role of the Mon1-Ccz1-C18orf8 complex in regulating lysosomal cholesterol export.

Main Methods:

  • Genome-wide CRISPR screen utilizing an endogenous cholesterol reporter.
  • Characterization of C18orf8-deficient cells.
  • Analysis of Rab7 activation, late endosome morphology, and LDL trafficking.
  • Investigation of lysosomal cholesterol accumulation and export.

Main Results:

  • Identification of over 100 genes critical for LDL-cholesterol import.
  • C18orf8 identified as a core subunit of the Mon1-Ccz1 guanidine exchange factor (GEF) for Rab7, essential for its stability and function.
  • C18orf8 deficiency leads to impaired Rab7 activation, defective endosomal LDL trafficking, and cellular cholesterol deficiency.
  • Accumulation of free cholesterol in lysosomes indicates a defect in lysosomal cholesterol export.
  • Active Rab7 interacts with NPC1 to license lysosomal cholesterol export, a process dependent on the Mon1-Ccz1-C18orf8 complex.

Conclusions:

  • The trimeric Mon1-Ccz1-C18orf8 (MCC) GEF complex is central to cellular cholesterol homeostasis.
  • The MCC complex coordinates Rab7 activation, endosomal LDL trafficking, and NPC1-dependent lysosomal cholesterol export.
  • Dysregulation of this pathway has significant implications for cellular cholesterol metabolism and potential disease states.