高等生物化學及分子生物學期末報告How are proteins secreted out of

animal cells?

6th Team

0050184 李道一 0157030 陳盈孝0157036 陳柔樺 9828518 劉冠佑

Presentation on Jan. 10th 2013

Outline

• Protein Secretory Pathways In Animal Cells

– Post protein synthesis (Ribosomes): Co-translational

Translocation

– Rough Endoplasmic Reticulum

– Golgi Apparatus: Cisternal maturation

– Vesicles and Exocytosis

• Unconventional Secretory Pathways

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Secretory Pathway• A series of steps a cell uses to move proteins out of the cell; a process

known as secretion.

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Early stages of SPFrom ER to cis-Golgi

Cisternal maturationFrom cis-Golgi to trans-Golgi(a kind of nonvesicular process)

Later stages of SPVesicle bud from trans-Golgi network

Copyright © McGraw-Hill Education, LLC. All rights reserved

EARLY STAGES OF THE SECRETORY

PATHWAY

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Co-translational Translocation

• 60s ribosomal subunit is aligned with the pore of the translocon in such a way that the growing chain is never exposed to the cytoplasm and is prevented from folding until it reaches the ER lumen. eukaryotic system

Nature 450, 663-669 (29 November 2007)

Signalpeptidase

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Post-translational Translocation

• Spase: signal peptidase

• ERj1: allow communication between ER lumenal BiP and translating ribosomes; aid polypeptide translocation or folding

nucleotide exchange factors (NEF)2014/4/18 How are proteins secreted out of animal cells? 6

Cargo from ER to Golgi

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Anterograde (Forward)Transport is mediated by COPII vesicles.

Retrograde (Reverse)Transport is mediatedby COPI vesicles.

CISTERNAL MATURATION

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The Golgi Apparatus

• The Golgi complex is organized into three or four sub-compartments, which are often arranged in a stacked set of flattened sacs, or cisterna.

• The sub-compartments of the Golgi differ from one another according to the enzymes that they contain. Many of the enzymes are glycosidases and glycosyltransferases that are involved in modifying N-linked or O-linked carbohydrates attached to secretory proteins as they transit the Golgi stack.

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Cisternal Maturation Model

• The cisternae of the Golgi apparatus move by being built at the cis face and destroyed at the trans face

• Against: Vesicular transport model (cisternalmaturation model lost favour in the 1980s, recently it has made a comeback, because additional evidence comes from the fact that COPI vesicles move in the retrograde direction, transporting endoplasmic reticulum proteins back to where they belong by recognizing a signal peptide)

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LATER STAGES OF THE SECRETORY

PATHWAY

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Coat-protein: Clathrin

• Triskelion shape:– Three heavy chains

– Three light chains

• When the triskelia interact they form a polyhedral lattice that surrounds the vesicle.

• Coat-proteins, like Clathrin, are used to build small vesicles in order to safely transport molecules within and between cells.

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Vesicle Pinch Off Is An Energy Consuming

/ Dynamin-Mediated Process • Dynamin

– Essential for release of complete vesicle.

– When bud formatting, dynaminpolymerizes around the neck portion and then hydrolysis gtp. The energy derived from GTP hydrolysis is thought to drive a conformational change in dynamin that stretches the vesicle neck until the vesicle pinches off.

• AP(adapter protein) complex: – Assemble between the clathrin

lattice and the membrane

H. Lodish, B. Anthony, C. A. Kaiser, A. Amon, A. Berk, M. Krieger, H. Ploegh and M. P. Scott, "Moving Proteins into Memberances and Organelles," in Molecular Cell Biology, 7th. ed., Macmillan, 2013, pp. pp 577-

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What’s the specificity of cargo transport?• Sorting signals: vesicle coat selects cargo molecules is by directly binding to specific

sequences in the cytosolic portion of membrane cargo protein.

• Recognize soluble proteins in lumen: acts as receptors

• Recognize transmembrane proteins: bind to cytosolic domain

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Animalcellbiology, Chapter 4: Intracellular Compartments & Transport- Via Vesicles. August 11, 2011

Added by Anonymous , last edited by gxt5@psu.edu on Aug 21, 2009 16:25

UNCONVENTIONAL SECRETORY

PATHWAYS

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Unconventional Secretory Pathways

• Discovery of golgi independent trafficking from

brefeldin-a resistance.

• Unconventional secretory pathway utilizes two possible

different routes to secrete proteins out of cells.

– Non-vesicular

– Vesicular

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Golgi Independent Trafficking

• During passage through the Golgi, N-linked glycan

chains are modified so that glycoproteins at the cell

surface are endoglycosidase H (endo H) resistant.

However, an endo H–sensitive form of the receptor

protein-tyrosine phosphatase CD45 rapidly

accumulates at the cell surface in

• T lymphoma cells, so as in the trafficking of the cell

adhesion molecules.

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Brefeldin A (BFA) Resistance

• BFA is a fungal metabolite that inhibits the activation

of ADP-ribosylation factor 1 (ARF1) on Golgi

membranes and, therefore, blocks vesicular

transport along the secretory pathway. Golgi-

independent trafficking occurs only in specific cell

types, indicating that it is a regulated process

limited to specific physiological conditions that can

operate in addition to trafficking along the

conventional secretory pathway.

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Unconventional Secretory Pathways

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Unconventional Secretory Pathways

• Vesicular mechanism

– Lysosomal secretion:• E.g.: Inflammatory cytokines such as interleukin-1b(IL-1b)

– Exosomes / Multivesicular bodies

– Exovesicles / Membrane blebbing• E.g.: Member of the galectin family proteins (galectin-1)

• Non-vesicular mechanism

– Plasma membrane-resident transporters:• E.g.: the FGF-2 transportation

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Conclusions

• The unconventional secretory mechanisms still have

many phenomenon not to be actually explained by now.

• The unconventional secretory pathway is a golgi-

independent, BFA-resistant system. Mainly processing

by vesicular and non-vesicular routes.

• The unconventional secretory proteins mostly have

biomedical properties in higher eukaryotes such as

FGFs and cytokines, except for the ACBP in yeast cells.

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References

• H. Lodish, B. Anthony, C. A. Kaiser, A. Amon, A. Berk, M. Krieger, H. Ploeghand M. P. Scott, "Moving Proteins into Memberances and Organelles," in Molecular Cell Biology, 7th. ed., Macmillan, 2013, pp. pp 577-.

• R. H. Garrett and C. M. Grisham, in Biochemistry, Brooks/Cole CengageLearning, pp. 1095-.

• M. Lee, E. Miller, J. Goldberg, L. Orci and R. Shekman, "Bi-directional protein transport between ER and and Golgi," Annu Rev Cell Dev Biol, vol. 20, p. 87–123, 2004.

• Ken Matsuoka, Lelio Orci, Mylene Amherdt, Sebastian Y. Bednarek, Susan Hamamoto, Randy Schekman and Thomas Yeung, "COPII-Coated Vesicle Formation Reconstituted with Purified Coat Proteins and Chemically Defined Liposomes," Cell, vol. 93, p. 263–275, 1988.

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References (cont’d)

• Keller P and Simons K, "Post-Golgi biosynthetic trafficking," J Cell Sci, vol. 110, p. 3001–3009, 1997.

• W. Nickel, "The mystery of non-classical protein secretion. A current view on cargo proteins and potential export routes.," Eur J Biochem, vol. 270, p. 2109–2119., 2003.

• Prudovsky I, Tarantini F, Landriscina M, D. Neivandt and R. Soldi, "Secterion without Golgi," J Cell Biochem, vol. 103, p. 1327–1343, 2008.

• F. Trombetta and A. Parodi, "Quality control and protein folding in the secretory pathway," Annu Rev Cell Dev Biol, vol. 19, p. 649–676., 2003.

• W. Nickel, "Unconventional secretory routes: direct protein export across the plasma membrane of mammalian cells.," Traffic, vol. 6, p. 607–614, 2003.

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Thanks For Your Attentions

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