Sept 26 - Mitochondria Lecture

Question 1

Can yeast live without mtDNA?
Can yeast live without mitochondria?
Can humans live without mtDNA or mitochondria?

Answer 1

Yes - glycolysis
No - biochemical processes like heme synthesis necessary
No

Question 2

AGG codon in humans versus mitochondria

Answer 2

Nucleus - arg

Mitochondria - STOP

Question 3

UGA codon in nucleus versus mitochondria

Answer 3

Nucleus - STOP

Mitochondria - Trp

Question 4

Mitochondrial purification

Answer 4

1. Rupture cells in hyperosmolar solution
2. Differential centrifugation - Low speed to remove unbroken cells and nuclei. High speed to pellet mitochondira.
3. Gradient - sucrose, percoll, or nycodenz

Question 5

What percent of protein functions of mitochondria are unknown?

Answer 5

25%

Question 6

Respiratory complex III

Answer 6

Bc1 complex

Question 7

What do mitochondrial carrier proteins carry? Across inner membrane

Answer 7

Nucleotides, pyrimidines, AAs, ketoacids, other things (phosphate, flavin, CoA

Question 8

Dnm1

Answer 8

aka Drp1. Helps with mitochondiral fisshion

Question 9

Fzo1

Answer 9

aka hFis1. Helps with mitochondrial fusion

Question 10

Fzo1-1 mutant

Answer 10

Mitochondria do not fuse in this mutant; there is a fusion block. When you put the gene back in (pRS416-FZO1 rho) you get a rescue phenotype
Does not grow on glycerol. You can look for fission genes by seeing what can restore the growing-on-glycerol function. (dnm1, fis2/mdv1 were identified this way)
.

Question 11

Fis1

Answer 11

If you block fission the result is a mesh-like yeast mitochondria.

Question 12

Fzo1 and Fis1

Answer 12

Blocking fission and fusion - causes a restoration of function. Normal growth and morphology.

Question 13

Mgm1

Answer 13

No fusion

Question 14

Charcot-Marie-Tooth

Answer 14

Affects the longest nerves in the body - problems with fission and fusion.

Question 15

MFN2

Answer 15

FZO1 homolog. Defects in the MFN2 GTPase domain lead to CMT2A and neuronal degeneration in long neurons of legs, muscle weakness, sensory loss.

Question 16

PINK1

Answer 16

Serine/threonine kinase targeted to mitochondria. Accumulates with loss of psi…

Question 17

PARKIN

Answer 17

Ubiquitin ligase recruited by PINK1. Defects in PARKIN cause inherited parkinson’s.
Mitochondrial fusion is blocked by ubiquitinating Mfn, promoting clearance of damaged organelles/ fission

Question 18

Proteolytically processed, resistant to external protease, requires delta psi. Location? Further studies?

Answer 18

Matrix, inner membrane, or inter-membrane space.
Sonication, carbonate extraction, swelling+protease

Cox4, ABCD7, Cytochrome B2

Question 19

Not proteolytically processed, resistant to external protease, requires psi

Answer 19

Matrix, inner membrane
Sonication (to see if membrane or not), carbonate extraction (membrane or not), swelling+protease (how much of it is in membrane)

ISd11, Aac

Question 20

Not proteolytically processed, resistant to external protease, does not require psi

Answer 20

IMS, OM (buried or facing IMS), protease stable aggregate
Carbonate extraction
Swelling+protease
Detergent+protease

Porin, Mic14, Dre2

Question 21

Not proteolytically processed, resistant to external protease, does not require psi

Answer 21

Outer face of OM, Unspecific binding
Carbonate extraction

Fzo1, Tom7, Bcl2

Question 22

What does MPTP target?

Answer 22

mitochondria