Transcript No Slide Title - Docenti.unina

How proteins get out of the Golgi apparatus?
TGN
Proteins that exit the Golgi apparatus are
directed to three possible destinations
Lysosomes are ’bags’ filled with hydrolytic enzymes
Many different enzymatic activities are present in lysosomes
1.
2.
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10.
Acid phosphatase
Deoxiribonuclease
Ribonuclease
Galattosidase
Mannosidase
Esosaminidase
Carboxipeptidase
Amminopeptidase
Elastase
D-cathepsin
Acid phosphatase can be demonstrated by hystochemistry
ACID PHOSPHATASE
glycerol +
-glycerophosphate
lead nitrate +
P
P
lead phosphate
Enzymes can be visualized by hystochemical reactions
Acid
phosphatase
DIFFERENT
INTRACELLULAR
COMPARTMENTS
HAVE DIFFERENT pH
pH CAN BE
MEASURED BY THE
USE OF pHSENSITIVE
FLUORESCENT
PROBES
LYSOSOMES
HAVE A LOW
pH OF ≈5-red
ENDOSOMES
pH5.5-6.5
blue-green
Lysosome are heterogeneous
Mannose 6-phosphate is a signal that directs a lysosomal
hydrolase to lysosomes
Lysosome targeting signal forms in 2 steps
PROTEIN
MANNOSE
UDP-GlcNAc
phosphotransferase
UMP
PROTEIN
MAN-6-P-GlcNAc
phosphoglycosidase
PROTEIN
MAN-6-P
GlcNAc
Mannose-6-phosphate signal formation
A phosphotransferase recognizes lysosomal
hydrolases in the Golgi apparatus
A phosphotransferase recognizes lysosomal
hydrolases in the Golgi apparatus
LYSOSOMAL
HYDROLASE
UDP-GlcNAc
UMP
PHOSPHOTRANSFERASE
catalytic
site
recognition
site
Newly synthesized lysosomal hydrolases are
delivered from Golgi apparatus to endosomes
Lysosomal enzymes are transported to endosomes
Clathrin
E
+
M6P
ENDOSOME
GOLGI
APPARATUS
M6P receptor
What happens if the phosphotransferase is lacking?
Lysosomal diseases are rare and severe, genetic diseases
Lysosomal diseases are caused by a defect in the enzymes that degrade
a variety of molecules within the cell. Non-degraded compounds
accumulate in the lysosomes and alter their function.
CLASSIFICATION
MUCOPOLYSACCHARIDOSIS
OLIGOSACCHARIDOSIS
SPHINGOLIPIDOSES
They are progressive diseases that lead to severe functional alterations.
Some of them are lethal. Autosomal recessive. Frequency ≈ 1/5000.
Multiple pathways deliver materials to lysosomes
Autophagy degrades proteins and organelles, when needed
Secretory pathways can be constitutive or regulated
Secretory pathways can be constitutive or regulated
lipide di
membrana
proteina
solubile
SECREZIONE
COSTITUTIVA
proteina di
membrana
TGN
segnale
SECREZIONE
REGOLATA
Secretion can be either constitutive or regulated
CONSTITUTIVE
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all cells
continuous
vescicles do not accumulate
short-lived vesicles
signal not necessary
no concentration
REGULATED
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•
specialized cells
non-continuous
vesicles accumulate
long-lived vesicles
signal necessary
concentration
We can envisage 4 steps in protein secretion
1.
Vesicle formation from the TGN (BUDDING)
2. Vesicle maturation
3. Vesicle transport to the plasma membrane
4. Vesicle fusion to the plasma membrane
1
VESICLE FORMATION (BUDDING)
2
Secretory granules undergo maturation
Some proteins undergo processing during secretory
granule maturation
3
Secretory granules move along microtubules
3
Secretory granules move along microtubules
DIC MICROSCOPY
3
Secretory granules move along microtubules
Two families of molecular motors move along microtubules
Kinesin motors ‘walk’ on microtubules
4
Exocytic vesicles and granules fuse to the plasma
membrane
4
When a vesicle fuse to the plasma membrane:
1. Its content is discharged in the extracellular space
2. The vesicle membrane fuse with the plasmamembrane
3. Proteins of the vesicle membrane are delivered to the
plasmamembrane
Ca++, Na+
Ca++ IONS
PLAY A KEY
ROLE IN THE
EXOCYTIC
PROCESS
Ca++ pump
Exocytosis of a mast cell is very rapid after the signal
histamine secretion
Exocytosis can be polarized
Figure 13-69 Molecular Biology of the Cell (© Garland Science 2008)
Synaptic vesicle fusion occurs very rapidly
For fusion to occur membranes should be closer than 1.5 nm
SNARE proteins play a crucial role in the fusion process
In some cases the fusion step can be very short
KISS AND RUN
Rapid formation of synaptic vesicles occurs in a nerve cell
Exocytosis is needed for plamamembrane recycling
Figure 13-70 Molecular Biology of the Cell (© Garland Science 2008)
Protein sorting in the secretory pathway requires signals
CONSTITUTIVE
REGULATED
Aggregation: pH e Ca++
No signal needed
Disulfide bond: CgB, POMC
Protease cleavage site:
Prorenin
Carboxypeptidase E: Proinsulin
and Proenkephalin
Carboxi terminal: PC2
Many cell types, like epithelial cells, are polarized
Tight
junction
Apical
membrane
Basolateral
membrane
Epithelia cells can be cultured on filters in bicameral systems
They form polarized monolayers
TER
BIOTIN
Protein secretion in polarized cells requires sorting signals
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Ap •
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Bl
cytosolic sequence
transmembrane domain
GPI anchor?
(N or O) sugars
cytosolic sequences
similar to endocytic
signals
In a polarized epitelial there are two ways to sort
proteins to the plasma membrane
Nerve cells are also well polarized
Figure 13-71 Molecular Biology of the Cell (© Garland Science 2008)
There are similarities in the polarity of nerve cells
and epithelial cells
THE END