1- Mitochondria Lecture

Question 1

functions of the mitochondria

Answer 1

• atp production
• regulate cell metabolism (oxidation of sugars/fats, synthesis/breakdown/interconversion of amino acids)
• cell death
• calcium storage
• biosynthesis (heme, steroids, iron sulfur clusters)
• cell signaling via ROS (reactive O2 signaling)
• participate in urea cycle

Question 2

what type of major organs are impacted by mitochondrial disease?

Answer 2

most high energy organs (ex: muscle, eye, brain, heart, liver, GI)

Question 3

mitochondria are composed of…

Answer 3

bigenomic proteins (from both nuclear and mitochondrial gene products)

Question 4

why is mitochondrial-nuclear crosstalk so important?

Answer 4

essential for mitochondrial functions including correct synthesis, import/folding/assembly, and functioning of respiratory chain enzymes

Question 5

Respiratory subunits in mitochondria

Answer 5

complexes I, III, IV, V

all encoded by mitochondrial genome except complex II (succinate dehydrogenase)

Question 6

6 must know facts about mitochondrial DNA (re: inheritance and replication)

Answer 6

1. each mitochondrion has 2-10 copies of mtDNA: tons of cytoplasmic inheritance in each cell
2. mtDNA is maternally inherited
3. mtDNA replication is not dependent on cellular division
4. mtDNA undergoes replicative segregation duirng mitosis and meiosis
5. mtDNA has mutation rate 10-20x higher than nuclear DNA (polymerase is not as good)
6. mtDNA mutation has a threshold expression; when the threshold level is crossed, the cell (thus organ) will suffer

Question 7

there are mutations in mitochondrial genome that cause disease…what are the 4 categories

Answer 7

1. giant deletions (tend to be spontaneous and not inherited)
2. tRNA mutation
3. rRNA mutation
4. mRNA mutation

Question 8

How does nuclear DNA interact with mitochondria? (3 ways)

Answer 8

1. cooperates with mtDNA in synthesis and assembly of multi-subunit enzyme complexes of oxidative phosphorylation
2. encodes all components of protein import machinery
3. encodes transcription factors for mtDNA transcription & replication & for proteins that target mitochondria

Question 9

can mutations in different genes cause the same syndrome?

Answer 9

yes

Question 10

Leigh’s syndrome, why is it so interesting?

Answer 10

it is caused by mRNA mutations of complexes I, II, III, and IV. Severity is different depending on the legion.

Question 11

3 possible sources of mutations in mitochondria?

Answer 11

1. nuclear DNA: mendelian inheritance
2. cytoplasmic mtDNA: non-mendelian; maternal inheritance
3. X-linked

Question 12

3 major mtDNA mutations

Answer 12

1. attached to IMM, source of ROS
2. lacks protective histones
3. limited repair system
   - copy number mutations also exist

Question 13

missense mutations

Answer 13

aa substitutions

LHON (Lebers hereditary optic neuropathy)

Question 14

biogenesis mutations: tRNA point mutations affects protein synthesis

Answer 14

• MERRF (myoclonus epilepsy and ragged red fibers)

- MELAS (mitochondrial encephalomyopathy, lactic acidosis, and stroke-like symptoms)

Question 15

insertion-deletion mutations

Answer 15

deletions of 1.3-7.6 kb

• often flanked by direct repeats of nucleotides
• usually no family history
• KSS (Kearns sayre syndrome)
• PEO (Progressive external ophthalmoplegia)

Question 16

mtDNA homo vs. heteroplasmy

Answer 16

homoplasmy - normal

heteroplasmy- diseased/aged

Question 17

explain the mitochondrial genetic bottleneck

Answer 17

primordial germ cell -> primary oocyte -> mature oocyte

primordial has a ton of mitochondria and it splits off into primary oocyte then they mature and you will have different levels of good and mutant mitochondria

low level mutation- offspring unaffected

high level mutation- offspring affected

intermediate level mutation- offspring may or may not be affected

Question 18

Fission and fusion as quality control events

Answer 18

fission- breaks apart the mitochondria and the good parts are distributed, the bad are mitophagy (taken to trash) or if its really bad then apoptosis occurs

the good parts are then fused

after fission there should be a spread out of damaged mtDNA so it can still be below the threshold

Question 19

How are nuclear encoded proteins incorporated?

Answer 19

> 90% of mitochondrial proteins have to be imported

Targeting sequence (signal, leader, or presequence)

• 20-70 aa
• rich in basic and hydroxylated aa at N-terminus
• can form amphipathic structures
• cleaved by specific peptidases in the mitochondria

Cytosoilic factors (Hsp70): unfolding, need ATP

Question 20

are mitochondria the same in every tissue

Answer 20

no! they are sculpted in a tissue-dependent manner

Question 21

major pathway affecting mitochondrial biogenesis (part that was boxed)

Answer 21

AMPK -> PGC-1a -> expression of multiple mitochondrial genes

Question 22

AMP kinase

Answer 22

phosphorylates PGC-1a and its the main sensor in the cell due to adenylate cyclase (high AMP levels stimulate AMP kinase)

• master metabolic regulator
• maintains nucleotide pools over wide range of enzyme demands
• AMP stimulates glycogenolysis and glycolysis
• AMPK inactive until phosphorylated
• AMP binding inhibits dephosphorylation and allosterically activates
• ATP antagonizes these effects
• > 30 downstream targets

Question 23

PGC-1a

Answer 23

induces the expression of multiple mitochondrial genes by driving transcription factors which drive protein synthesis and import into the mitochondria

Question 24

mitophagy

Answer 24

removes damaged good. (cell apoptosis in the substantia nigra for all of these)

Question 25

apoptosis is mediated at the mitochondria

Answer 25

• important during development and in fighting viral infections
• misregulation of apoptosis causes or contributes to a large number of disease
• regulation occurs at 3 steps BUT the committed step is cyt c release
• Bcl-2 proteins control cyt c release
• Bcl-2 proteins come in two flavors:
  1. pro-apoptotic (Bax, Bak) or pro-survival (Bcl-2, Bcl-xL, McI1, A1, Bcl-w)
• Pro-apoptotic thought to form channel that allow cyt c (and others) release

Question 26

3 person IVF

Answer 26

potential treatment for mitochondrial disease

mothers nucleus (with unhealthy mitochondria) goes into a donor egg with healthy mitochondria

Question 27

diseases of aging in which mitochondrial dysfunctions are implicated

Answer 27

you are burning fuel and that causes oxidative damage. as you get older this builds up and you start to reach that threshold. this threshold is different for different people and since some people
eat better/exercise more they have different amounts of mitochondrial damage.

1. Normal aging
   - reactive oxygen species (superoxide, H2O2, OH- radicals): major mtDNA-damaging agents
   - mtDNA deletions accumulate with age
   - oxidative phosphorylation normally declines with age
2. parkinson’s (paralysis agitans)
3. alzheimer’s

Question 28

Mitochondrial dysfunction factor in neurodegenerative diseases

Answer 28

mitochondrial dysfunction and oxidative stress factor in these delayed onset diseases….

• parkinson’s
• alzheimer’s
• huntington’s
• amyotophic lateral scerosis (ALS)

mtDNA mutations arent primary cause of disease
-cause/effect relationship leading to disease currently under investigation but mitochondra do play a central role