Ims: Histo Block 1 Lecture 2

Front 1

What is the endoplasmic reticulum comprised of? What are its functions?

Back 1

Network of membrane tubules and sheets which are continuous through the cell.

The ER membrane produces many proteins and lipids for organelles, membranes, and export.

Front 2

How does the ER membrane expand itself?

Back 2

Enzymes on the cytosolic surface of the ER use fatty acids to make phospholipids which are added to the cytosolic side of the ER membrane and then transferred to the cisternal surface by flippase.

Front 3

What are the functions of the smooth ER? Where may SER be found?

Back 3

Cholesterol synthesis, detoxification, calcium release

Found in cells producing steroid hormones (glands), which use cholesterol to make hormones

Found in cells that detoxify molecules, such as in the liver

Sarcoplasmic reticulum and oocytes require Ca++ from SER

Front 4

What are the key features of Rough ER?

Back 4

Polyribosomes on surface that translate proteins of ER, and other organelles

Front 5

What path do soluble proteins follow once they've eneter the ER cisterna? What about membrane proteins?

Back 5

They may stay there, move to another compartment, or, in the case of the membrane proteins, enter the ER membrane

Front 6

What are free polysomes?

Back 6

Cluster of free polyribosomes bound to mRNA

Front 7

What is the role of free polysomes?

Back 7

Code for proteins of the cytosol, mitochondria, chloroplasts, and nuclei; also some peroxisomal proteins

Front 8

What occurs after a free polysome has completed translation?

Back 8

mRNA, nascent protein, large and small subunits separate and join a common pool; they can then be used again as either free polysomes or polysomes on RER

Front 9

Describe the steps that occur when a protein is being translated by a RER polysome (before enters lumen).

Back 9

In the cystosol, small ribosome subunit binds mRNA
tRNA and large subunit bind small subunit to begin translation
ER signal sequence of 16-30 aa's formed at amino end of protein
Signal sequence binds signal recognition particle in ER membrane; halting translation
SRP released, translation is resumed and growing protein chain inserted into translocation channel

Front 10

Describe the steps that occur when a protein is being translated and has entered the ER lumen.

Back 10

Signal peptidase removes signal sequence
Soluble protein enters ER cisterna and assumes 3D configuration

Front 11

Contrast co-translational with posttranslational in terms of protein synthesis. Gives examples.

Back 11

Ex: entry of protein into ER lumen during translation in proteins synthesized by RER polysomes VS entry of protein into organelle post-translationally in protein synthesized by free polysomes

Front 12

What happens if a protein is misfolded in the ER cisternae?

Back 12

It's exported to the cytosol and broken down in proteasomes

Front 13

Describe how transmembrane proteins are inserted into the ER.

Back 13

Transmembrane proteins have a stop-transfer signal that is recognized by the translocation channel.
This causes chanel to discharge protein laterally into membrane.
Stop-transfer signal is an alpha helix which anchors the protein into the membrane.
The sequence is enzymatically removed from N-terminus which extends into ER lumen.

Front 14

What's the most common pathway for a vesicle? Start at the ER.

Back 14

ER -> Golgi -> Outside

Front 15

Describe the enzymatic pathway for vesicle formation.

Back 15

Clathrin is joined to adaptin, which binds specific carbo molecule

Clathrin molecs assemble on cytosolic surface and shape membrane into a coated pit

Dynamin causes coated pit to pinch off to form coated vesicle

Uncoating enzyme later removes clathrin

Front 16

Where can clathrin-coated molecules move things to?

Back 16

Can move material from Golgi to other destinations or even inwards from cell surface

Front 17

Describe how vesicles are directed to their respective targets.

Back 17

Transmembrane SNARE proteins:
v-SNARES on vesicles and t-SNAREs on target membrnaes ensure vesicles reach appropriate compartment.
The two SNAREs wrap around each other to fuse the vesicle and target membranes together

Front 18

How and where does glycosylation occur?

Back 18

Oligosaccharide of 14 sugars is transferred from lipid dolichol in ER membrane to asparagine on nascent protein as it enters the ER (Co-translational modification)

The asparagine that is glycosylated occurs as part of a tripeptide sequences: asp-x-serine or asp-x-threonine, where x is any AA

Front 19

If a protein translated on the ER ribosome is not an ER resident, via what pathway can it get to the Golgi?

Back 19

COP (coat protein) vesicles

Front 20

If an ER resident protein mistakenly enters the Golgi, how is it able to return to the ER?

Back 20

ER resident proteins have an ER retention signal (KDEL) at their carboxyl end. If they should enter the Golgi, they'd bind KDEL receptors and would be returned to the ER by COP (coat protein) vesicles.

Front 21

What are the three compartments of the Golgi app? Describe their locations in relation to the ER.

Back 21

Cis (closest to ER), medical, trans (furthest from ER)

Front 22

What is the function of COPII-coated vesicles?

Back 22

Transport material from ER to cis Golgi

Front 23

What is the function of COPI-coated vesicles?

Back 23

cist to medical to trans Golgi (anterograde, aka fwd, transport) transport

can also go retrograde: trans to medial to cis; ER resident proteins bound to KDEL receptors follow this to get back to ER

Front 24

Describe the general vesicular processes for movement from the ER to the Golgi.

Back 24

Budding of vesicle from donor compartment, fusion of vesicle to target compartment, release of contents in target compartment

Front 25

How do the contents of the three regions of the Golgi differ? What do they have in common?

Back 25

Contain different enzymes; all modify, package, and sort proteins.

Front 26

Where on a protein would the Golgi add/remove sugars?

Back 26

From/to oligosaccharide added in ER

Front 27

Contrast constitutive secretion with regulated secretion.

Back 27

In constitutive secretion, GOlgi products are immediately discharged (such as in production of plasma membrane)

In regulated secretion, Golgi products are stored as secretory vesicles which are released in response to a stimulus (such as insulin release)

Front 28

What is phagocytosis? What structures allow for it?

Back 28

When cells engulf whole cells or parts of cells; actin involved in pseudopod formation for phagocytosis

Front 29

What is pinocytosis?

Back 29

Cells ingest extracellular fluid by pinocytosis using small vesicles; this is non-selective

Front 30

What is receptor-mediated endocytosis? How does it occur?

Back 30

Ligands (STUFF TO BE PLACED IN VESICLE) bind externally to transmembrane receptor proteins which are attached to clathrin molecules inside plasma membrane.

Dynamin causes coated pit to pinch off as coated vesicle

In cytoplasm, uncoating enzyme removes clathrin coat. Clathrin is recycled to plasma membrane, and vesicle is fused with an endosome.

Front 31

Describe the interior of the endosome and how it is achieved. What is the reason for this?

Back 31

Endosome interior has a low pH due to ATP-driven proton pump.

This can cause ligands to dissociate from their receptors. A vesicle rich in receptors then buds off to recycle receptors to plasma membrane.

Front 32

How is a late endosome achieved? What happens in a late endosome?

Back 32

Proton pumps continually lower pH to form a late endosome. This allows for hydrolytic digestion of the endocytosed material.

Front 33

What is the main role of an early endosome?

Back 33

Sorting station

Front 34

What are the various fates of ligands enclosed in endosomes?

Back 34

Can remain bound to receptor within endosome, can leave endosome while bound to receptor, can later be hydrolyzed

Front 35

What role do lysosomes play?

Back 35

Contain enzymes which digest endocytosed material or old cell components (autophagy); resulting small molecules enter cytosol via transport proteins

Front 36

What is the relative pH of lysosomes? How is it maintained?

Back 36

Acidic (pH ~5); maintained by proton pumps (ATP synthase working in reverse: ATP --> ADP + Pi)

Front 37

Why don't lysosomal enzymes digest themselves?

Back 37

Lysosomal enzymes are protected against proteolytic digestion because they're glycosylated.

Front 38

How would lysosomes appear when using cytochemistry? Assume an ultrathin section is floated on a solution containing glycerol phosphate and a soluble lead compound.

Back 38

Phosphatase in lysosomes would cause the lead to be precipitated on the lysosome. Densely packed with lead.

Front 39

How are endosomes converted into lysosomes? Begin with discussing the manufacture of proteolytic enzymes in the ER.

Back 39

In the ER lumen, oligosaccharides are added to the enzyme.

In cis Golgi, enzymes are recognied by a signal patch of AA, and mannose 6-phosphate (M6P) is added to the oligosaccharide.

At trans Golgi, M6P binds M6P receptors and enzyme is enclosed in clathrin-coated vesicle, which leaves Golgi and loses coat.

Vesicle fuses with endosome where low pH causes M6P receptor to dissociate from enzyme.

Removal of phosphate group from mannose prevents enzyme from re-attaching to receptor.

Front 40

What would happen if a vesicle with lysosomal enzymes fused with the cell membrane?

Back 40

Limted harm; exterior of cell has pH at about 7.4, which is diff from pH optimum of lysosomal enzymes.

M6P receptors on the plasma membrane would bind the enzyme and internalize them via receptor-mediated endocytosis

Front 41

What would be the effects of a lysosomal enzyme lacking M6P?

Back 41

If lacked M6P to direct it to a lysosome, there could be a damaging accumulation of the enzyme's usual substrate, much like in Tay-Sachs Disease.

Front 42

What are peroxisomes and what is their role in the cell?

Back 42

Spherical organelles with single membrane surrounding granular matrix. They produce H2O2 by oxidizing a substrate.

Can also oxidize fatty acids to form acetyl CoA which goes to cytosol.

Front 43

What is catalase? Where is it found?

Back 43

A powerful oxidant which breaks down excess H2O2 into H2O and O2. Also inactivated toxic molecules by oxidizing them with H2O2.

Found in PEROXISOMES.

Front 44

Describe the formation and growth of peroxisomes. Begin with protein translations.

Back 44

Peroxisomal precursor vesicles arise from RER, and contain proteins translated on polyribosomes of RER.

Vesicles then import proteins synthesized on free polysomes in cytosol. Can now divide into daughter peroxisomes.

Front 45

Describe the physical layout of mitochondria.

Back 45

2 membranes, 2 compartments

Front 46

Describe mitochondrial DNA.

Back 46

Circular molecule, no histones. Mito can synthesize proteins.

Front 47

Briefly describe the citric acid cycle.

Back 47

In cytosol: 1 glucose --> 2 pyruvate + 2 ATP + 2 NADH
NADH and pyruvate imported into inner mitochondria
Pyruvate--> acetyl CoA-->enters TCA

TCA:
acetyl CoA-->GTP, NADH, FADH2
Electrons from GTP, NADH, FADH2 pass through electron-transport chain and create proton gradient
Gradient drives ADP-->ATP phosphorylation

Front 48

How is oxidative phosphorylation driven in the IMM?

Back 48

IMM is impermeable to protons, so must be pumped out by electron-transport chain.
H+ want to enter IMM along [ ] gradient, so they pass through ATP synthase
As they go through, rotor rotates and phosphorylation of ADP occurs

Front 49

What would happen if thermogenin were present in the IMM?

Back 49

This is an uncoupling agent. Rather than phosphorylation of ADP, heat would be generated as H+ pass through ATP synthase.

Front 50

What does UGA stand for in most cells? In mitochondrial DNA?

Back 50

universal STOP codon; codes for tryptophan in mitochondria

Front 51

How many circles of DNA present per mitochondrion?

Back 51

2-10

Front 52

During what cell phase is mitochondrial DNA replicated?

Back 52

Interphase

Front 53

Describe how a protein is imported from the cytosol into the IMM.

Back 53

Import sequence on peptide + cyt Hsp70 (holding peptide chain unfolded) bind import receptor on mitochondria (TOM COMPLEX)

Snake it through translocation channel and through TIM complex, mit Hsp70 chaperons attach to keep from backsliding through channel

Signal peptidase cleaves signal

ATP and membrane potential across IMM drive import of proteins

HSP60 may be req'd