L5: Cell Biology IV

Question 1

What is atlastin? What does increased amounts of atlastin lead to? Atlas deficiency?

Answer 1

• GTPase involved in constructing and shaping the ER
• Increased ER membrane fusion and normal Golgi absent
• Atlastin deficient = ER that is fragmented. Deficient atlastin linked to hereditary spastic paraplegia - - symptoms = leg stiffness, gait disturbances, mental retardation, extrapyramidal symptoms, dysmorphic features

Question 2

Proteasome. Location, function?

Answer 2

• ATP-dependent protease that is located in the cytoplasm and nucleus
• Function: proteolysis of regulatory proteins, malformed/denatured/damaged proteins and antigenic proteins

Question 3

Effect of prions on proteasomes?

Answer 3

• Abnormal prions inhibit proteasomes

Question 4

Clinical relevance of proteasome

Answer 4

• Defective ubiquitination of proteins implicated in Parkinson’s disease
• Form of CF involves proteasomal degradation of a CFTR ABC transport variant that is slow to fold, but is competent

Question 5

What inhibits proteasomes?

Answer 5

• Abnormal prions
• Bortezomib (used in treating multiple myeloma) causes partial inhibition of proteasomes. Proteasome inhibition decreases degradation of pro-apoptotic factors that then promotes apoptosis of cancer cells.

Question 6

Structure of Golgi apparatus? Location?

Answer 6

• Organelle that is typically located in proximity to nucleus (in polarized cells, located between nucleus and apical face of cell).
• Comprise of 3-10 slightly curved flattened membranous sacs (cisternae)
• Cis (convex) face presents towards transitional ER
• Trans (concave) face is opposite to cis face

Question 7

Golgi apparatus function?

Answer 7

Major sorting and distribution center

• Proteins sorted based on chemical moieties such as sequences of AA or attached CHOs
• Proteins are modified in Golgi (glycosylation, sulfation, phosphorylation, oligosaccharides previously added are trimmed, proteolytic cleavage of proteins)
• Proteins packaged into vesicles

Question 8

Histology of Golgi – describe

Answer 8

• Neither acidophilic or basophilic. Pale-staining area of cytoplasm may be seen in H&E.

Question 9

Differentiate between what is occurring at cis vs trans Golgi

Answer 9

• Cis: transport vesicles carrying newly synthesized proteins from ER to Golgi are being formed for movement through Golgi
• Trans: secretory granules, lysosomes and membrane protein-transporting vesicles are being formed

Question 10

What is Wilson’s disease?

Answer 10

• AR disorder caused by mutation in protein pump for copper in trans-Golgi network of liver cells.
• Result= impaired biliary excretion of copper. Kayser-Fleicher rings surrounding iris can sometimes be seen, which is copper deposition in Descemet’s membrane

Question 11

What would happen to ceruloplasmin serum level in Wilson’s disease? Explain

Answer 11

• ATP7B is a copper transporter in the Golgi membrane, takes copper from cytoplasm into Golgi. Apoceruloplasmin in Golgi is protein not bound with Cu. When Cu is present, apoceruloplasmin binds it and forms ceruloplasmin, which then can be secreted into plasma.
• In Wilson’s, copper transporter is defective, not moving Cu into Golgi, don’t form ceruloplasmin, therefore it decreases.

Question 12

Discuss mutations in dysferlin gene, clinical relevance and organelle implicated

Answer 12

• Membrane protein-transporting vesicles containing dysferlin and important in repair of microperforations formed in skeletal muscle PM during contraction / relaxation.
• In repair process, dysferlin-containing vesicles are shipped from Golgi to cytosolic leaflet of sarcolemma where they accumulate and reseal microperforations in a calcium-dependent manner
• In mutated gene, damage membrane is not resealed and muscle degenerates
• Results in three types of muscular dystrophy (miyoshi myopathy, limb-girdle muscular dystrophy type 2b and distal myopathy tibialis anterior muscle onset). All forms are characterized by muscle weakness and atrophy of distal (principally) leg or proximal limb muscles

Question 13

Describe proinsulinemia in context of Golgi

Answer 13

• Mutated proinsulin is missorted and placed into constitutive pathway
• Prohormone converting enzymes are sorted correctly into regulated pathway
• Consequently, there is absence of prohormone coverting enzyme activity in upregulated pathway

Question 14

Function of endosomes?

Answer 14

• Organelles involved in sorting and identifying molecules for recycling or degradation in lysosomes

Question 15

4 endosomal compartments

Answer 15

1.) Early endosomes
2.) Recycling endosomes
3.) Multivesicular bodies
4.) Late Endosomes
• Differ by cytoplasmic location, luminal pH, possess unique molecular markers

Question 16

Describe early and recycling endosomes.

Answer 16

• At cell’s periphery
• pH 6.2-6.5 (aided by membrane-bound proton-ATPases)
• Glucose transport in response to insulin??

Question 17

Describe multivesicular bodies

Answer 17

• Located bw early and late endosomal compartments
• pH 5.0-6.2 (aided by membrane-bound proton-ATPases)
• Contain large amounts of membrane and vesicles
• Eventually fuse with late endosomes, also have secretory role via exosomes
• Migrate along microtubules

Question 18

Describe late endosomes

Answer 18

• Located near Golgi apparatus and nucleus
• pH ~ 5.0 (aided by membrane-bound proton-ATPases)
• These fuse with or mature into lysosomes. Aka prelysosomes

Question 19

Describe 4 fates of receptor-ligand complexes. Provide examples.

Answer 19

1. ) Receptor is recycled and ligand is degraded or released for cellular use. Example = LDL and its receptor
2. ) Receptor and ligand are recycled. Example = iron and it transferring receptor
3. ) Receptor and ligand are degraded. Example = FGFR3 with ligand FGF
4. ) Receptor and ligand are transported across cell and secreted (transcytosis): how glandular cells secrete antibodies

Question 20

Achondroplasia at the cellular level

Answer 20

• FGF binds FGFR3 (fibroblast growth factor) and is endocytosed
• Both ligand and receptor are usually targeted for lysosomal degradation as a means to terminate signaling
• In achondroplasia, lysosomal targeting is degraded as there is a mutation in FGFR3. Instead, FGFR3 (still active) is recycled to plasmalemma and amplifies FGF signaling. Result = abnormal bone growth leading to most common cause of dwarfism.

Question 21

Lysosomes. Function, location? Example of lysosomal hydrolase transport

Answer 21

Function?
- Transport of hydrolases (with enzymatic activity greater at acidic pH) and other chemicals that are able to digest most biologic molecules.
Location?
- Found in most cells, but especially abundant in professional phagocytic cells such as neutrophils and macrophages
Example?
- Lysosomal hydrolase precursor is tagged when it arrives from ER in Golgi with mannose
- As it moves through cisternae of Golgi, it reaches trans Golgi. Packaged into vesicle.
- M6P (mannose-6-phosphate) receptor recognizes it in vesicle and transported to lysosome
- Defect in M6P synthesis leads to lysosomal storage disease

Question 22

Three autophagic pathways

Answer 22

1. ) Macroautophagy
2. ) Microautophagy
3. ) Chaperone-mediated direct transport

Question 23

What happens to the material after the lysosome has completed its action?

Answer 23

• Digested material diffused out of lysosomes into cytoplasm OR
• Residual bodies (undigested wastes) form. Waste product can be eliminated from cell via exocytosis or waste accumulates as cell ages. Eg. Lipofuscin in cardiac and nerve cells.

Question 24

Are lysosomes acidophilic or basophilic?

Answer 24

• Lysosomes are acidophilic

Question 25

Name lysosomal storage disease. What enzyme is deficient, what metabolite accumulates?

Answer 25

1. ) Type II Pompe’s dz: alpha-1,4-glucosidase defiency causing accumulation of glycogen
2. ) GM1 gangliosidosis: GM1 ganglioside beta galactosidase deficiency causing accumulation of GM1 ganglioside, galactose-containing oligosaccharides
3. ) GM2 gangliosidosis (Tay Sachs): hexosaminidase-alpha-subunit deficiency causing accumulation of GM2 ganglioside
4. ) GM2 gangliosidosis (Sandoff): hexosaminidase-beta-subunit deficiency causing accumulation of GM2 ganglioside and globoside
5. ) Metachromic leukodystrophy: arylsulfatase A deficiency causing accumulation of sulfatide
6. ) Gaucher: glucocerebrosidase deficiency causing accumulation of glucocerebroside
7. ) Hurler: alpha-L-iduronidase deficiency causing accumulation of dermatan and heparan sulfate

Question 26

23 yo male admitted to hospital for fulminant hepatorenal failure. Pt was jaundiced, Kayser-Fleischer rings absent and pt was confused, combative and unable to cooperate or communicate during exam. Ratio of alk phos to total bili = 0.05, very low value, ceruloplasmin was normal. Microscopic exam of liver biopsy show steatohepatitis and was negative for copper. Pt ultimately died a few days after admission. On autopsy, greenish-yellow appearance on sectioning of the liver with signs of cirrhosis. Copper was 979 ug/gram (normal range: 10-35). What is the diagnosis?

Answer 26

• Wilson’s disease
• Ceruloplasmin is normal (normally decreased in Wilson’s)
• Kayser-Fleischer rings may or may not be seen in Wilson’s (not in this case)
• Biopsy didn’t show copper while pt was alive. Biopsy sample was area that wasn’t affected. Multiple biopsies should have been performed.