Intracellular Vesicular Traffic 2 Flashcards
What are lysosomes?
Membrane enclosed compartments filled with hydrolytic enzymes.
What are lysosomes derived from?
Heterogeneous - derived from late endosomes.
What are lysosomes important for?
Intracellular digestion of macromolecules.
How many different types of enzymes are found in lysosomes?
About 40:
- proteases
- mucleases
- glycosidases
- lipases
- phospholipases
- phosphatases
- sulfatases
What is required for optimal activation of lysosomes?
acidic environment and proteolytic cleavage
What happens to the end products of digestion in the lysosome?
Transporters in membrane pump out end products.
How is the acidic pH in lysosomes maintained?
Vacuolar ATPase pumps H+ into lysosomes.
also to drive transport of small metabolites
Approximately what is the pH in the lysosome?
pH of about 5
How can histochemical visualization of lysosomes be achieved?
by staining the acid phosphatases
What is the order of lysosome maturation starting at the plasma membrane?
- early endosome > microtubule-mediated transport - late endosome - endolysosome - lysosome
List the multiple pathways that deliver materials to lysosomes.
> phagocytosis
endocytosis
autophage from mitochondrion
macropinocytosis (“cell drinking”)
Of the multiple pathways for material to get to the lysosome for degradation, which two pathways do not go through the late endosome process?
Autophagy –> turns into autophagosome and goes straight to fuse with lysosome.
Phagocytosis –> forms phagosome and goes straight to fuse with lysosome.
Regarding the model of autophage, list the steps in this mechanism.
> nucleation and extension
closure
fusion with lysosome
digestion
Where are the membrane proteins and hydrolases that are required for lysosome function transported from?
Trans Golgi Network (TGN)
- co-translationally transported into rER and then transported via Golgi complex to TGN.
- TGN buds off into endosomes
- develop into lysosomes
What is the sorting signal attached to lysosomal hydrolases?
mannose-6-phosphate (M6P)
Where is the mannose-6-phosphate (M6P) sorting signal on lysosomal hydrolases attached in?
CGN
What recognizes the sugar (M6P) in the TGN?
M6P receptors in the TGN recognize sugar.
What two things do receptors bind to in the TGN?
Receptors bind to hydrolases and to adaptor proteins in assembling clathrin coats.
What are lysosomal hydrolases packaged into?
Clathrin-coated vesicles that bud from TGN.
What causes the dissociation between the lysosomal hydrolase and the M6P receptor?
acidic pH inside the early endosome
Once the lysosomal hydrolase and M6P receptor are dissociated in the early endosome, what is removed from the lysosomal hydrolase?
phosphate is removed and you now have a lysosomal hydrolase precursor
Once the addition of P-GlcNAc occurs in the cis Golgi network (CGN), what happens to the M6P sorting signal before the protein reaches the TGN?
uncovering of M6P sorting signal
so that it can bind to the M6P receptor
What enzyme transfers P-GlcNAc to the mannose in the catalytic site during the assembly of the M6P sorting signal to the lysosomal hydrolase?
GlcNAc phosphotransferase
Once the lysosomal hydrolase is released from GlcNAc phosphotransferase, what is removed to complete the addition of the M6P sorting signal?
GlcNAc is removed - leaving only M6P bound to the lysosomal hydrolase.
What causes lysosomal storage diseases?
Genetic defects in lysosomal hydorlases.
Why do genetic defects in lysosomal hydrolases lead to disease?
Accumulation of undigested material in lysosomes.
What enzyme has a mutation that leads to Hurler’s disease?
mutation in the enzyme required to break down glycosaminoglycan chains.
What is Inclusion Cell Disease?
ALL of the lysosomal hydrolases are missing in many cell types.
- undigested substrates accumulate as “inclusions”
- defective or missing GlcNAc phosphotransferase, an enzyme that adds M6P to lysosomal hydrolases.
- the enzymes are NOT phosphorylated and hence not sorted into vesicles and NOT delivered into lysosomes
- they are instead carried to cell surface and secreted (found in blood)
What enzyme adds M6P to lysosomal hydrolases?
GlcNAc phosphotransferase
What is endocytosis?
Uptake of macromolecules from exterior across plasma membrane.
During endocytosis, what type of vesicles ingest large particles?
Large particles ingested by vesicles called phagosomes - phaocytosis.