3021-2017-15 slides - Lecture notes 15 PDF

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BCH3021 Lectures 15 - 20 The secretory and endocytic pathways: organelles and molecular mechanisms of protein trafficking

Prof Phil Bird

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Organelles and protein transport routes in the secretory and endocytic systems Protein Transport Pathways: Proteins synthesized here

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(1) default export to surface no specific signal required (2) regulated export from storage vesicle - no specific signal identified

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(3) targeting to lysosome requires specific signal (4) import from surface (endocytosis). May require specific signal

In these lectures, we will be discussing dynamic systems in live cells………….

What we will cover ……….

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1. Functions, relationships and formation (biogenesis) of organelles Alberts 12-5

2. Protein synthesis & trafficking routes

3. Moving proteins around the secretory and endocytic pathways: machinery and mechanisms

“Signal sequences” used to direct specific proteins to organelles Alberts 12-6

Transfer of proteins between organelles involving budding and fusion of transport vesicles

So what? > 60 human diseases associated with defects in these pathways

Global market for protein drugs > US $234 billion in 2014 70% are synthesized / secreted from eukaryotic cells

Nobel Laureates ? Golgi trafficking 1906 Camillo Golgi (shared with Santiago Ramón y Cajal) “in recognition of their work on the structure of the nervous system"

Cell fractionation mitochondria

1974 George Palade, Christian de Duve & Albert Claude "for their discoveries concerning the structural and functional organization of the cell"

Endocytosis 1985 Michael Brown and Joseph Goldstein “for their discoveries concerning the regulation of cholesterol metabolism" http://www.nobelprize.org

lysosomes

Signal sequences 1999 Günter Blobel, for the discovery that "proteins have intrinsic signals that govern their transport and localization in the cell."

Vesicular transport 2013 Randy Schekman, James Rothman and Thomas Sudhof, “for their discoveries of machinery regulating vesicle traffic, a major transport system in our cells"

BCH3021 Lecture 15 Structure and biogenesis of the Endoplasmic Reticulum and Golgi complex CORE REVIEW: Nature Reviews Molecular Cell Biology 8: 429-439 (2007)

Concepts •  The ER is a large network of membrane-bound tubes involved in protein and lipid synthesis, calcium storage and detoxification •  The Golgi is a dynamic organelle that accepts cargo from the ER and passes it further along the secretory pathway. It contains three distinct membrane-bound compartments that further modify proteins as they pass through. •  when a cell divides it has to pass intact organelles or a template on to the daughter cells (organelles can’t be constructed de novo from information in DNA) •  The ER remains intact and functional during mitosis and is split among daughter cells •  The Golgi breaks down during mitosis and re-forms from the ER

The Endoplasmic Reticulum (ER) The first organelle in the secretory pathway

•  network of interconnected membrane tubules enclosing single internal space, the lumen or cisternal space • ER lumen is topologically equivalent to the exterior of the cell •  ER occupies ~ 10 % of the cell’s volume •  The ER membrane comprises over half the membrane in cell •  ER membrane is continuous with the outer nuclear membrane •  ER has a central role in lipid and protein biosynthesis ( ~1 / 3 of a cell’s protein is made in the ER) Alberts 12-7

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Functions of the ER PROTEIN SYNTHESIS, MODIFICATION, TRAFFICKING (discussed in later lectures) •  synthesis and glycosylation of proteins destined for the cell surface •  synthesis of resident proteins of the secretory / endocytic organelles (ER, Golgi, lysosomes, secretory vesicles) •  proteins can be soluble i.e. totally released into the lumen OR membraneassociated MEMBRANE SYNTHESIS •  site of synthesis of lipids incorporated into the membranes of most of the organelles in the cell CALCIUM STORAGE •  Ca++ is stored for release in response to extracellular signals DETOXIFICATION •  ER resident enzymes detoxify lipid-soluble drugs and toxic metabolites

The ER has two parts: the rough ER and smooth ER

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Rough ER and smooth ER structures can be separated by cell fractionation procedures

•  if cells are disrupted by homogenization the ER fragments and reseals into small vesicles called microsomes

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Rough ER is studded with ribosomes involved in secretory protein synthesis •  synthesis of cytosolic, nuclear, mitochondrial and peroxisomal proteins occurs in the cytoplasm on free ribosomes •  synthesis of secretory proteins occurs on ribosomes bound to the membrane of the ER (discussed in next lecture)

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Smooth ER •  smooth ER is a region of ER that lacks ribosomes •  smooth ER is the site of lipid synthesis, Ca++ storage and detoxifying enzymes •  site of assembly and departure of vesicles travelling to Golgi (ER exit sites) – discussed in later lecture •  most cells have much more rough ER than smooth ER

INTRACELLULAR COMPARTMENT Plasma membrane Rough ER membrane Smooth ER membrane Golgi membrane Mitochondrial membrane Nucleus (inner membrane) Peroxisome membrane Lysosome membrane Endosome membrane

PERCENTAGE OF TOTAL CELL MEMBRANE Hepatocyte Acinar cell 2 5 35 60 16...