Mitochondria Flashcards
1
Q
Who called mitochondria the “powerhouse of the cell” and when?
A
Philip Siekevitz in 1957.
2
Q
Who discovered mitochondria and when?
A
Albert von Kölliker in 1857 in insect muscles.
3
Q
Who coined the term mitochondrion and when?
A
Carl Benda in 1898.
4
Q
What are mitochondria primarily known for?
A
ATP production, biosynthetic functions, and signaling hubs.
5
Q
What is the evolutionary origin of mitochondria?
A
Endosymbiosis of a prokaryote by a larger cell over 1.45 billion years ago.
6
Q
What are the two main theories regarding mitochondrial origin?
A
Engulfment by a eukaryote vs. direct endosymbiosis by a prokaryote.
7
Q
who created the endosymbiotic theory
A
Lynn Margulis in 1970
8
Q
What are the four compartments of mitochondria?
A
Outer membrane, intermembrane space, inner membrane, and matrix.
9
Q
What are the functions of the outer membrane?
A
Contains porins, enzymes for lipid synthesis, and fusion/fission proteins.
10
Q
What is the intermembrane space involved in?
A
H+ pumping, cytochrome c storage, apoptosis regulation.
11
Q
What are key functions of the inner membrane?
A
Electron transport chain, ATP synthesis, transport proteins.
12
Q
What happens in the mitochondrial matrix?
A
Krebs cycle, mtDNA storage, gene expression, mtDNA repair.
13
Q
What is mitochondrial fusion?
A
Merging of mitochondria to mix contents and enhance function.
14
Q
What is mitochondrial fission?
A
Splitting of mitochondria for division, quality control, and apoptosis.
15
Q
What proteins regulate fusion?
A
Mitofusin-1 (MFN1), Mitofusin-2 (MFN2), and OPA1.
16
Q
What proteins regulate fission?
A
Drp1 (dynamin-related GTPase)and Fis1 (mitochondrial fission protein 1).
17
Q
How does oxidative stress affect mitochondrial dynamics?
A
Can lead to increased fission and mitochondrial fragmentation.
18
Q
What is the primary function of mitochondria?
A
ATP synthesis through oxidative phosphorylation.
19
Q
What substrates do mitochondria use for ATP production?
A
Pyruvate (from carbohydrates) and fatty acids (from fats).
20
Q
Which cycle generates high-energy electrons for ATP synthesis?
A
The Citric Acid (Krebs) cycle.
21
Q
What is oxidative phosphorylation?
A
Process of ATP production using electron transport chain and proton gradient.
22
Q
What are the five complexes of the electron transport chain?
A
Complex I (NADH-dehydrogenase), II (succinate dehydrogenase), III (cytochrome b-c1), IV (cytochrome c oxidase), V (ATP synthase).
23
Q
What is the role of coenzyme Q (ubiquinone)?
A
Transfers electrons between Complex I and III.
24
Q
What is the role of cytochrome c?
A
Transfers electrons between Complex III and IV.
25
What happens when mitochondria uncouple oxidative phosphorylation?
Proton leak allows energy dissipation as heat instead of ATP production.
26
What is the structure of mitochondrial DNA (mtDNA)?
Circular, double-stranded DNA stored in the mitochondrial matrix.
27
What does mtDNA encode?
13 proteins, 22 tRNAs, and 2 rRNAs essential for mitochondrial function.
28
Why is mtDNA more prone to mutations than nuclear DNA?
Higher exposure to reactive oxygen species (ROS) and lack of histones.
29
What is the average size of mitochondrial nucleoids?
~300 nm in human fibroblasts (confocal), ~99 nm (STED microscopy).
30
How does oxidative stress impact mtDNA?
Causes mutations and fragmentation, leading to mitochondrial dysfunction.
31
What is the function of TFAM?
Mitochondrial transcription factor that helps package mtDNA.
32
What are reactive oxygen species (ROS)?
Highly reactive molecules that can damage DNA, proteins, and lipids.
33
What are examples of oxidatively damaged DNA bases?
8-oxoguanine, thymine glycol, 5-hydroxycytosine.
34
How does mitochondrial DNA repair work?
Through Base Excision Repair (BER) PROVEN and Single-Strand Break Repair (SSBR) NOT PROVEN.
35
What enzymes are involved in mtDNA repair?
DNA glycosylases (OGG1, UNG), APE1, DNA polymerase gamma (Polγ), Lig3.
36
What happens when mtDNA damage is not repaired?
Mutations accumulate, leading to mitochondrial diseases and aging effects.
37
How is mitochondrial function measured experimentally?
Extracellular Flux Analyzer to assess oxygen consumption rates.
38
What is the effect of aging on mitochondrial function?
Reduced repair efficiency, increased ROS, and accumulation of mtDNA mutations.
39
How does oxidative stress affect mitochondrial DNA integrity?
Increased fragmentation, leading to impaired ATP production and apoptosis.
40
What is LA-qPCR used for?
To assess mitochondrial DNA integrity and damage levels.
41
How does the Total Repair Assay work?
Uses enzymatic steps to measure base excision repair efficiency.
42
What is an incision assay used for?
Detects the activity of APE1 and other endonucleases in BER pathways.
43
What is the significance of mitochondrial fusion assays?
Used to visualize mitochondrial mixing in live cells.
44
What microscopy techniques are used for mitochondria?
Confocal microscopy, STED super-resolution microscopy, electron microscopy.
45
Why is mitochondrial fission important in apoptosis?
It facilitates the release of cytochrome c, triggering programmed cell death.
46
How does glycolytic rate relate to mitochondrial function?
Compensatory mechanism when oxidative phosphorylation is impaired.
47
what diseases are mitochondria involved in
Alzheimers
Parkinsons
nonalcoholic fatty liver
cardiac ischemia-reperfusion injury
cancer
48
mitochondria catalyze a major conversion of energy by
oxidative phosphorylation
49
mitochondrial uncoupling allows
controlled leak of protons into the
mitochondrial matrix down the
concentration gradient without
entering the ATP synthase and
phosphorylating ADP to ATP.
50
What is the purpose of an Extracellular Flux Analyzer?
It measures cellular bioenergetics by assessing oxygen consumption rate (OCR) and extracellular acidification rate (ECAR).
51
What does OCR (oxygen consumption rate) indicate?
It reflects mitochondrial respiration and ATP production efficiency.
52
What does ECAR (extracellular acidification rate) indicate?
It measures glycolytic activity by detecting acidification due to lactate production.
53
What key inhibitors are used in extracellular flux analysis?
Oligomycin (ATP synthase inhibitor), FCCP (uncoupler), and Rotenone/Antimycin A (electron transport chain inhibitors).
54
oligomycin
ATP synthase inhibitor
55
FCCP
uncoupler
56
Rotenone and Antimycin A
ETC inhibitors
57
How does oligomycin affect OCR?
It inhibits ATP synthase (Complex V), reducing ATP-linked respiration and revealing proton leak-driven respiration.
58
What is the role of FCCP in mitochondrial stress tests?
It uncouples oxidative phosphorylation by dissipating the proton gradient, maximizing mitochondrial respiration capacity.
59
Why are rotenone and antimycin A used together in extracellular flux analysis?
They inhibit Complex I and Complex III, completely shutting down mitochondrial respiration to assess non-mitochondrial oxygen consumption.
60
What does a mitochondrial stress test measure?
Basal respiration, ATP production, maximal respiration, and spare respiratory capacity.
61
How does the extracellular flux analyzer help in studying oxidative stress?
It compares OCR changes under normal and oxidative stress conditions to assess mitochondrial dysfunction.
62
What does a reduced spare respiratory capacity indicate?
A cell’s inability to respond to increased energy demands, which is often linked to mitochondrial dysfunction and disease.
63
what two enzymes are found in mitochondria but not nucleus
EXOG
Poly
64
how are BER activities measured
Incision cleavage assay
total repair assay
65
What is mitochondrial import?
Process of transporting nuclear-encoded proteins into mitochondria.
66
What happens when mitochondrial import is defective?
Proteins may mislocalize, leading to oxidative stress and disease.
67
What disease is linked to defective mitochondrial import of MnSOD?
Severe alcoholic liver disease.
68
What does Mn superoxide dismutase (MnSOD) do?
Converts superoxide to hydrogen peroxide to reduce oxidative stress.
69
What happens when MnSOD is mislocalized to the intermembrane space?
Increases oxidative stress within mitochondria.
70
What is EXOG, and what happens when it is depleted?
A mitochondrial 5'-exonuclease; its depletion leads to mitochondrial dysfunction and programmed cell death.
71
What happens to mtDNA-deficient cells when EXOG is depleted?
They become resistant to cell death.
72
How does mitochondrial dysfunction affect apoptosis?
Triggers cell death pathways by increasing oxidative stress and DNA damage.
73
Why is mitochondrial membrane depolarization important in mitophagy?
It signals the destruction of damaged mitochondria through mitophagy.
74
How does fission and fusion help maintain mitochondrial function?
Fusion mixes mtDNA to repair damage; fission segregates damaged mitochondria for removal.
75
How does mitochondrial metabolism influence cancer?
Affects bioenergetics, oxidative stress, apoptosis, and signaling in tumors.
76
What happens when tumor cells lose mtDNA?
They show delayed tumor growth.
77
How can tumor cells regain mtDNA function?
Through horizontal transfer of mtDNA from the host tumor microenvironment.
78
What is the role of mtDNA in tumor progression?
Essential for bioenergetics and tumor-initiating capacity.
79
Why do tumors from rho0 cells have altered bioenergetics?
They lack mtDNA and rely on glycolysis instead of oxidative phosphorylation.
80
How do tumors compensate for loss of electron transport chain function?
By restoring mitochondrial gene expression from host mtDNA.
81
What is the significance of COX1 and 12S rRNA in cancer?
Their presence indicates functional mitochondrial respiration in tumor cells.
82
How does oxidative stress impact mitochondrial function in cancer?
It influences tumor survival, adaptation, and resistance to therapies.
83
What is P-glycoprotein (Pgp) in cancer cells?
A drug efflux pump that reduces chemotherapy effectiveness.
84
How does mitochondrial-targeted doxorubicin (mtDox) differ from standard doxorubicin (Dox)?
It remains toxic even in drug-resistant cancer cells.
85
How does mtDNA trigger inflammation?
It can activate immune responses similar to bacterial DNA.
86
What is unique about mtDNA transcription?
Almost the entire genome is transcribed in both directions, increasing the chance of dsRNA formation.
87
How does mtDNA structure contribute to inflammation?
It has a three-stranded D-loop structure that can activate immune receptors.
88
How does oxidative stress affect mtDNA integrity?
It induces damage, leading to inflammatory responses in cells.
89
What are key inflammatory cytokines activated by mtDNA damage?
IL-1α, IL-6, IL-8, and TNFα.
90
What signaling pathway is activated by mtDNA damage?
ZBP1/TBK1/IRF3 pathway.
91
What is TLR9, and how is it linked to mtDNA?
A toll-like receptor that detects mtDNA and triggers immune activation.
92
How does mitochondrial damage contribute to chronic inflammation?
Persistent release of mtDNA and ROS sustains immune activation.
93
What diseases are associated with mtDNA-induced inflammation?
Autoimmune disorders, cancer, and neurodegeneration.
94
How can mtDNA integrity be measured experimentally?
Using PCR-based assays and imaging techniques like PLA (Proximity Ligation Assay).
95
How does mtDNA damage contribute to aging?
Mutations accumulate over time, leading to mitochondrial dysfunction.
96
What happens to mice when mtDNA depletion occurs?
They develop premature aging symptoms like skin wrinkles and hair loss.
97
Can aging effects caused by mtDNA depletion be reversed?
Yes, restoring mtDNA replication reverses aging phenotypes in mice.
98
What is the dominant-negative mutation in PolG?
A mutation that impairs mtDNA replication, leading to accelerated aging.
99
How is mtDNA repair linked to longevity?
Efficient repair mechanisms help maintain mitochondrial function and delay aging.
100
What imaging methods are used to study mtDNA damage in aging?
Confocal microscopy and super-resolution imaging techniques.
101
How does oxidative stress accelerate mtDNA mutations?
ROS-induced DNA lesions lead to replication errors and mitochondrial dysfunction.
102
What role does mtDNA play in neurodegeneration?
Mutations contribute to diseases like Alzheimer’s and Parkinson’s.
103
Why is mitochondrial quality control important in aging?
Prevents accumulation of damaged mitochondria through mitophagy.
104
What dietary interventions may support mitochondrial health?
Caloric restriction and antioxidants reduce oxidative damage.
105
What is mitochondrial replacement therapy (MRT)?
A technique to prevent mtDNA-related diseases by replacing defective mitochondria.
106
What diseases can MRT help prevent?
Leber's hereditary optic neuropathy (LHON), MELAS, MERRF.
107
What is Maternal Spindle Transfer (MST)?
A technique where nuclear DNA from the mother is transferred to a donor egg with healthy mitochondria.
108
What is Pronuclear Transfer (PNT)?
A method where nuclei from a fertilized egg are transferred to an enucleated donor zygote.
109
What are ethical concerns regarding MRT?
Embryo manipulation, long-term risks, and accessibility.
110
What are potential risks of MRT?
Mitochondrial DNA carryover, epigenetic modifications, and developmental issues.
111
How does MRT impact future generations?
Mitochondrial changes are passed down maternally.
112
Which MRT method has the least mtDNA carryover risk?
Polar Body Transfer (PBT).
113
What is a Proximity Ligation Assay (PLA)?
A technique to study protein interactions and modifications in situ.
114
What is a cleavage assay used for?
To measure mitochondrial DNA repair activity
115
main complexes involved in mitochondrial protein transfer
TOM, SAM, TIM, MIA, MIM
116
proteins involved in hypoxia and mitochondrial ROS signaling
HIF1a: Hypoxia-inducible factor 1-
alpha
PHD2: prolyl hydroxylase domain-containing protein 2
VHL: Von Hippel–Lindau syndrome
protein
117
4 ways to study mitochondria
1. Imaging modalities: MitoTracker, co-staining, PLA,
2. Biochemical assays: cleavage assay, incorporation assay to study mtDNA damage and repair
3. Extracellular Flux Analyzer: mitochondrial respiration (and glycolysis)
4. PCR approach to examine mtDNA damage and repair
