Cell Bio 1: Membrane and Golgi

What three types of biomolecule are cell membranes composed of?

Lipids (primarily phospholipids), proteins and carbohydrates

Features of cell membranes

flexible, resealable, control import and export, receive information- therefore they are required to be fluid

Thickness of phospholipid bilayer

5nm

How do phospholipids differ in archaea compared to in eukaryotes and bacteria?

• In eukaryotes and bacteria, phospholipids have a hydrophilic head, D-glycerol, an ester linkage and 2x unbranched fatty acid chains

• In archaea instead of fatty acid chains they have branched isoprene chains and L-glycerol instead of D-glycerol

Give four examples of membrane protein functions

Transport, linkage, receptors and enzymes

Give an example of a membrane protein with more than one function

• Integrins are linkers as well as receptors

• RTKs (receptor tyrosine kinases) eg. insulin receptor catalyse phosphorylation of other proteins ie. they are receptors and enzymes

What is the old model of cell membranes that was replaced by the fluid mosaic model in 1972?

Lipoprotein Sandwich model

How do somatic cell fusion experiments support the fluid mosaic model of cell membranes?

• Fusion of human and mouse cells induced using a virus

• Red and green antibodies are added

• At first the red and green are separate but after 40 mins incubation at 37 degrees C, ‘mosaics‘ are formed showing that proteins diffuse

How does FRAP (fluorescence recovery after photobleaching) support the fluid mosaic model of cell membranes?

• Labelled molecules within a defined area of the membrane are permanently bleached using a laser

• If labelled components are free to diffuse in the membrane, then gradually the bleached area will be repopulated with neighbouring fluorescent molecules

• Rapid recovery of fluorescence is observed within minutes

How do freeze-fracture electron microscopy experiments support the fluid mosaic model of cell membranes?

Membrane proteins (nuclear pore complexes) will move across the membrane in the direction of an electric field

How much of the membrane content of a cell does the plasma membrane constitute?

2-5%

Which two organelles have very big membranes and why?

• The ER needs a large SA for protein secretion

• The mitochondrial inner membrane must be large for efficient ATP production

What is meant by nascent polypeptide?

A newly-synthesised polypeptide

What are three potential fates of the nascent polypeptide?

1. Co-translational import into ER lumen, then transport to destination via vesicles (happens if ribosome associates with ER membrane)

2. Protein remains free in cytosol (if ribosome remains free in cytosol)

3. Post-translational import into other organelles such as mitochondria

All secretory proteins have a signal protein recognised by a signal peptide on which organelle?

The endoplasmic reticulum

What is the signal hypothesis?

The theory that co-translational protein targeting to the ER is triggered by. a signal peptide encoded at the start of the protein

What is meant by membrane traffic?

the movement of membrane (and associated cargo or membrane proteins) throughout the cell via vesicles

Membrane traffic: anterograde pathway

Transport of new proteins towards the plasma membrane = SECRETION or the secretory pathway

Membrane traffic: retrograde pathway

Transport of existing or external proteins away from the plasma membrane (towards interior of cell) = ENDOCYTOSIS or endocytic pathway

Give some early evidence for membrane traffic

• Pulse-chase experiments with pancreatic cells from guinea pigs

• Cells are incubated with radioactive amino acid (leucine) for a brief period (pulse)

• Radioactive aa is removed and cells are observed after different time periods to see where labelled proteins have travelled to (chase)

What is the order of the secretory pathway in the guinea pig cell experiment?

ER to vesicles to golgi to secretory granules in cell surface. After 20 mins produced proteins were in golgi apparatus then later in secretory granules

How can cell fractionation be used in conjunction with radioactivity to investigate the secretory pathway?

• Cell fractionation: cells can be homogenised by gently disrupting plasma membrane, then organelles can be isolated by centrifuging

• The nucleus (largest, most dense) can be separated out first and then the supernatant centrifuged again to obtain fractions with different organelles

• Radioactivity can be followed through different fractions

What does the golgi apparatus do?

Receives the entire output of newly-synthesised proteins and lipids from the ER, processes them in the stacked cisternae and then distributes them to their correct destination either inside or outside the cell

What do scaffolds in the golgi do?

Maintain the polarised stack of cisternae

Cis face of golgi

Recruits from the rough ER

Trans face of golgi

Exit side of organelle

What is meant by the cisternal maturation model of the golgi?

• The cisternal maturation model suggests that (rather than material just moving through stacks) the stacks themselves mature and move throughout the Golgi

• Material emerging from ER forms a stack at the cis face but that stack will mature, move through Golgi and eventually become the trans face

• Identity of stacks is maintained by backwards flow of material