Lecture 12 - Using Yeast to Understand Membrane Trafficking Flashcards
Why do eukaryotic cells need membrane trafficking ?
Compartmentalisation allows more complexity
Enzymes can modify specific subsets of proteins in certain environments e.g glycosylation and proteolytic cleavage.
For sequential modifications, proteins need to be exposed to distinct sets of enzymes.
Membrane trafficking is also important in retrieval of proteins back to their ‘resident’ compartment.
What is the route of the secretory/ exocytic (biosynthetic) ?
ER -> Golgi - > endoscope/llysosome
What is the endocytic pathway (recylcing or degradative) ?
Cell surface -> endosome -> Golgi/ER/lysosome
The endo and exocytic pathways intersect and proteins can be modified as they transit the ER and Golgi
How are proteins modified during endo and exocytosis
Glycosylated by addition of oligosaccharides and proteolytically cleaved.
Both N- and O- linked glcosylation can occur
What is the purpose of glycosylation ?
To assist folding
As a ligand
i. Intracellular for trafficking/ sorting
ii. Outside the cell for interactions with extracellular matrix and with proteins/ sugars on the other cells .
Which genetic organisms are commonly used to identify genes and proteins involved in trafficking pathways ?
Mouse, Drosophila melanogaster (Fruit fly), Zebrafish, Caenorhabditis elegans (nematode), Dictyostelium discoideum, Saccharomyces cerevisiae (budding yeast)
What makes a model suitable for studies on membrane trafficking ?
Simplicity - trafficking occurs on a cellular scale so a single celled organism is likely to provide information.
Analysis of specific types of secretion e.g regulated secretion, would need a model system that is able to perform this function.
Often different systems reveal different information.
What are the advantages of yeast as a model organism ?
Amenable for genetic studies (can grow as haploid and diploid cells)
Entire genome sequence known since 1996 (and is fully annotated)
Cheap and easy to grow in large quantities (good for biochemical studies)
Limited gene diversity (both ±)
Fundamental pathways conserved
What are the disadvantages of yeast as a model organism ?
Limited cell-cell contact so unlikely to be informative about multicellularity
Small (5μm), so high resolution imaging studies of intracellular
compartments is difficult.
Has a cell wall which can preclude some types of studies
What is endocytosis ?
Endocytosis is the process through which the plasma membrane invaginates into the cell resulting in the production of a vesicle that is then able to fuse with endosomes and enter the endo-lysosomal membrane system.
Why is endocytosis important ?
Retrieval of molecules that formed part of the secretory vesicle for recycling
Downregulation of signals
Remodelling cell surface lipid and protein composition
Note: endocytosis is also a means of entry into cells for many pathogens and toxins
What are the stages in the endocytic pathway ?
Plasma membrane to endocytic vesicle
Endocytic vesicle to early endoscope
Early endoscope to late endoscope (MVB) or recycling to the plasma membrane
Late endoscope to Golgi or vacuole
What is the major function of the lysosome (vacuole) ?
The degradation of extracellular material taken up by endocytosis as well as certain intracellular components by a process termed autophagy
Lysosomes contain many degradative/ proteolytic enzymes that must one separate
What occurs during Vacuolar/ Lysosomal sorting ?
The lysosome’s resident enzymes are transported to the lysosome through the secretory pathway. At the late Golgi compartment (Trans Golgi Network), they are sorted into a pathway destined for lysosomes rather than the plasma membrane.
Genetic studies in yeast were again central to the identification of genes/proteins involved in this sorting and trafficking pathway.
How many vps genes have been identified ?
60
