Mitochondria

Question 1

Mitochondrial structures and their Functions

Answer 1

1. Outer mitochondrial membrane
   
   1. Permeable to SMALL molecules
   2. Contains apoptosis signal receptors
2. Intermembrane space
   
   1. Contains apoptotic proteins
3. Inner mitochondrial membrane
   
   1. Electron transport chain
   2. ATP synthesizing complex
   3. Impermeable to small and large molecules
4. Matrix (involved in breaking down food)
   
   1. TCA cycle
   2. Lipid oxidation
   3. houses mtDNA

Question 2

Food converted to ATP

Answer 2

1. Polymeric molecules are broken down
2. Displaced into mitochondrial matrix in the form of small carbon chains
3. NADH is produced from oxidation of food stuff
4. NADH releases high energy e-
5. E- passed along e- transport chain in inner membrane
6. Energy released by transfer used to drive process of ATP synthesis
   
   1. Consumes O2

Question 3

Production of mitochondrial proteins by 2 separate genetic systems

Answer 3

1. Most of the proteins in mitochondria are encoded by nuclear DNA, transcribed into mRNA, and made into proteins via cytosolic ribosomes which then are transported into the mitochondria.
2. Other mitochondrial proteins are encoded by mitochondrial DNA, synthesized by ribosomes within the organelle using mitochondrial produced RNA (all of these processes take place in matrix)
3. Protein traffic tends to be unidirectional cytoplasm à mitochondria with exception of apoptosis

Question 4

Human mtDNA replication

Answer 4

1. Replication of mitochondrial genome (mtDNA) occurs out of phase with cell cycle
2. Under constant conditions, process is regulated to ensure total number of mtDNA doubles every cell cycle to ensure the maintenance of a constant amount of mtDNA
3. In Euk, mtDNA is replicated in replisome
   
   1. Twinkle, a mtDNA helicase/primase unwinds ds circular mtDNA and adds primers
   2. mtDNA encoded SSB proteins bind to DNA and keep it single stranded.
   3. The replisome is made entirely with nuclear DNA and it replicates the mtDNA. It consists of:
      
      1. DNA Pol Gamma
         
         1. DNA polymerase activity
         2. Exo/proofreading activity
         3. Lyase activity for MMR
      2. 2 accessory subunits
      3. Catalytic subunit
   4. After replication, mtDNA is packaged with proteins into nucleoids each of which contains 1 or 2 mtDNA.

Question 5

Mitchondrial Transcription

Answer 5

1. Requires
   
   1. Mitochondrial Transcription factors B1 or B2.
   2. TFAM: transcription activator
   3. Mitochondrial RNA polymerase
2. Mitochondrial transcription generates polycistronic transcripts (one transcript contains many genes to code for many proteins i.e. lac operon)
   
   1. Can be processed to make mRNA, tRNA, and rRNA

Question 6

Mitochondrial Genome Translation

Answer 6

1. mtrRNA makes mt ribosomes
2. mtDNA encodes IF2 and IF3 translation factors
3. mtDNA encodes mttRNA
4. mtmRNA is used
   
   1. Very small or no 5’UTR
   2. No 5’ cap
   3. Poly A tail added
5. Codon/anticodon matching is lax in mitochondria
   
   1. Allows translation to occur with fewer tRNA since 1 tRNA can recognize 4 different types of codons

Question 7

Very high mutation rates of mitochondria

Answer 7

1. 10-20 times greater mutations in nuclear genes
2. mtDNA (matrix) is close to source of ROS generated during oxphos in inner membrane
3. mtDNA repair systems are not effective against oxidative damage
4. DNA pol gamma with proofreading susceptible to mutations and defects
5. No introns in mtDNA so mutations are mostly within the coding region
6. Accumulation of mutations in mtDNA decreases mitochondria efficiency in adulthood

Question 8

Maternal inheritence

Answer 8

1. During mammalian zygote formation, sperm mtDNA is removed by Ub
2. Therefore, there is only maternal inheritance of mtDNA
3. Characteristics of mtDNA inheritance
4. All offspring of an affected or carrier female are AT RISK of being affected
5. All carrier or affected daughters are AT RISK of transmitting the condition
6. Affected males cannot pass the condition on

Question 9

Bottleneck effect

Answer 9

1. Number of mitochondrial molecules within developing oocyte is reduced before subsequently being amplified to amount seen in mature oocyte
2. Reduction and amplification results in random shift in mtDNA mutational load between cells.

Question 10

Replicative segregation of mitochondria and mtDNA

Answer 10

1. Mitochondrial fission and fusion leads to random distribution of mitochondria and mtDNA
2. Random replicative segregation of mtDNA
   
   1. Can occur in mitosis or meiosis
   2. During cell division, many copies of mtDNA in each mitochondria replicate/sort randomly into daughter mtDNA
   3. These daughter mitochondria then distribute into daughter cells randomly
   4. After gaining a certain number of mutated mitochondria within cell, you can get disease phenotype in which there is a spectrum of different symptoms depending on number of mutants
   5. Mitochondrial diseases are characterized by reduced penetrance, variable expression, and pleitrophy
      
      1. Homoplasmay: when cell contains purely mutated or purely unmutated mtDNA in mitochondria within the cell
      2. Heteroplasmy: mixture of mutant and normal mtDNA containing mitochondria in same cell

Question 11

Threshold Effect

Answer 11

1. Threshold amount of ATP required for cell survival varies from cell type in a given tissue
   
   1. Cardiac muscle, skeletal muscle, and CNS cells have a higher threshold amount of ATP
2. Random partitioning of mutant and wildtype (wt) mitochondria through multiple rounds of mitosis causes collection of daughter cells varying in proportion of mutant and wt mitochondria.
3. When mutant mitochondria increase such that threshold amount of ATP is not produced, you get disease phenotype.
   
   1. Threshold level of ATP varies based on mutation and tissue type.
   2. Differences in levels contribute to characteristic patterns of organ vulnerability for different mitochondria. Diseases and clinical heterogeneity in patients with same mtDNA mutation

Question 12

MtDNA Changes with Age

Answer 12

1. mtDNA mutations increases with age, especially deletions, due to ROS damage and defective DNA pol gamma
2. Oxphos levels decline
   
   1. Result of increasing number of mutations in mtDNA
   2. OR random distribution of mutated mtDNAs in mitochondria in tissue
3. ATP synthesis levels in cell drop below threshold level and once it does, you get pathological symptoms

Question 13

Diseases with no Ragged Red Fibers

Answer 13

1. Leigh syndrome
   
   1. Caused by mutations in mtDNA and nuclear DNA that encode proteins in energy metabolism
   2. Inherited in autosomal recessive (if mutation is on 22 chormosomes), X-linked, (if mutation is on X chromosome) or maternal inheritance pattern (if mutation in in mtDNA)
   3. Symptoms
      
      1. Progressive encephalopathy (brain)
2. Leber hereditary optic neurpathy (LHON)
   
   1. Caused by mutations in mtDNA only
   2. Maternally inherited and shows male prevalence
   3. Symptoms
      
      1. Painless progressive loss of central vision

Question 14

Diseases with Ragged Red Fibers

Answer 14

1. CPEO and Kearns Sayre Syndrome (KSS)
   
   1. Succinate dehydrogenase = nuclear DNA encoded protein
   2. Cytochorme C= partially mtDNA encoded protein
      
      1. Presence of succinate dehydrogenase and absence of Cytochrome C indicates mtDNA mutations
   3. Pathology
      
      1. Mitochondria in diseased individuals
      2. Mitochondria large and structurally abnormal
      3. Abnormal cristae with inclusions
      4. Muscle biopsy
         
         1. Myofibers with clumps of disease mitochondria that appear as red deposits = Red Ragid Fibers (RRFs)
2. MELAS (me loss hearing lol)
   
   1. Deafness
3. MERRF
   
   1. Epilepsy