test 3

Question 1

Reduction-Oxidation Reactions

Answer 1

-one substrate reduced, on oxidized; involve addition or removal of ELECTRONS

-oil rig (oxidation is LOSING)

PRO-OXIDANT/ OXIDANT: oxidizing agent; “the oxidizing agent is reduced”, it causes something else to be oxidized so it becomes reduced

ANTIOXIDANT: reducing agent… it becomes oxidized, so it is LOSING electrons

Question 2

Free Radical Chemistry

Answer 2

-atoms have nucleus (pro and neu)
-when molecule is formed by bonds of atoms, orbitals are rearranged
STABLE MOLECULE: each orbital has 2 electrons, with opposite spin
FREE RADICAL: when there is an unpaired electron in an orbital, VERY REACTIVE becaues they are seeking to find another electron
THEREFORE… A* + B -> A + B*
where the * is the radical sign
new radical is LABILE (easy to change) and can rxt w another molecule SOOOO these reactions can keep taking place until a terminal reaction takes place (when radicals react with eachother to form a NON radical species ORRRR if quenched (it donates e to neutralize) by an antioxidant to give a non-radical species).

Question 3

What are the two ways in which a terminal reaction can occur to stop radical reactions

Answer 3

1) Two radicals react with eachother to form a no -radical speices
2) quenched by an antioxidant (antioxidance donates e, it does not become a radical and the radical becomes unradical )

Question 4

Reactive Oxygen Species:

Answer 4

ROS: compounds derived from PARTIAL chemical reduction of o2 (molecular oxygen)… therefore they are reactive bbut not necessarily free radicals because they do not always contain unpaire electrons

SUPEROXIDE RADICAL: o2: formed by one electron reduction of oxygen
o2 +4e- -> 2 h20
o2 + 1e - > o2-

HYDROGEN PEROXIDE:
superoxide is very reactive but shortlived, can spontaneously convert to h202 when reacted with hydroge
o2- + 2H+ -> h202 (NOT RADICAL but ROS)
-very long lived, (unlike superoxide), more stable than o2-, can travel long distances because it is long lived
-can pass through membranes

Nitric Oxide:
NO* (nitrogen based free radical)
-RNS (reactive nitrogen species)
-involved in many imp biological processes as SIGNALLING molecule IE VASOMOTOR factor of blood vessels (promotes vasodilation of smooth muscle)

PEROXYNITRITE:
NO* (Nitric oxide) + o2*- (superoxide) -> ONOO- (peroxynitrite)
-VERY LONG LIVED AND VERY REACTIVE

Question 5

ROS DEFENSE: ANTIOXIDANTS

Answer 5

Why we need defense: free radicals, ROS and RNS can damage: PROTEINS, DNA, LIPIDS… overall cell function
-to overt the oxidative damage we have antioxidant defense and Oxidative damage repair

ANTIOXIDANT DEFENSE:
-mech is foun in the form of antioxidant enzymes and biological (chemical ) antioxidants

-MAIN ENZYMES:-can be MADE IN BODY, and can induced by oxidant stress

1) Superoxide dismutase (SOD)
-catalyzes reaction of Superoxide and H to form H202
O2*- + 2H+ -> H202 + 02

-several forms
a) Copper-zinc containng SOD (CuZnSOD), FOUND IN CYTOSOL

b)Manganese containing SOD (MnSOD), FOUND IN MITOCHONDRIA, made in cytosol then transported; makes up 15-20% of TOTAL SOD

2) catalase
-mitochondria and other organelles
-changes hydrogen peroxide into water and o2
Catalase dismutase: 2H202 -> h20 and o2

3) glutathione peroxidase (GPx)
-mitochondria and cytosol
-h202 into water with glutathione being the electron DONOR
2GSH + h202 -> GSSG (Glutathione disulfide) + 2H20

Biological (chemical) antioxidants:
a) Glutathione- made in body
-water soluble thiol-containing peptide
-found high conc in virtually all cells
-mito, xytosol, extracell fluid
-SERVES AS SUBSTRATE FOR GLUTATHIOD PEROXIDASE (GPX) to convert h202 into h20
-ALSO IMP FOR KEEPING VITA C and E (limited) in reduced state so that they are able to donate electrons to ros or free radical to neutralize it)

b) Antioxidant vitamins (cant be made, so from diet)
-cant be induced from oxidative stress
ex: VitaminE and C

     VITAMIN E: TOCOPHEROL -most important FAT SOLUBLE ANTIOXIDANT IN BODY Found in: cell membranes , including mito inner mito membrane -donates e- to LIPID RADICALS

    Vitamin C: ASCORBIC ACID -main water soluble antioxidant found in the body -CYTOSOL as well as extracell fluid -directly acts as antioxidant by donating e- to o2*- and *OH

Question 6

look at slide 16

Question 7

ROS AND RNS AS SIGNALLING MOLECULES

Answer 7

-although mostlly talked about for etrimentral affects, also important bioactive molecules in CELL SIGNALLING
-at physiological concentrations (normal levels found in the body): regulate
1) CELL GROWTH
2) DIFFERENTATION
3) PROLIFERATION
4) APOPTOSIS

-low levels of oxidants increase antioxidant levels and activity throgh adaptive response
-so low levels are important for gene expression of antioxidant enzymes

ALSO STIMULATE GENE EXPRESSION OF:
Cytokines, transcription factors (synthesis and breakdown), and proteins; as well as regulate kinase and phosphatase activity, triggering a signalling cascade through protein PHOSPHYRLATION AND DEPHOS.

TWO MAIN SIGNALLING PATHWAYS:
1) Transcription Factors- Nuclear Faccotr kB, activated by ros and rns, once it is, goes to nucleus and turns on genes for inflammation, stress, etc

2) Protein Kinases: Mitogen Activated Protein Kinases (MAP KINASES). these are the ones that activate cell growth, differenation, apoptosis, prolif. Switch these mitogen activated protein kinases on and off

Question 8

slide 19

Question 9

Oxidative Stress

Answer 9

-Free rad, ROS, RNS are produced whereas antiox are always present. in cels.body
THEREFORE: oxidative stress is when there is an imbalance of oxidant GENERATION ad ANTIOXI DEFENSE
SOOO any factor that causes an increase in oxi gen and a decrease in anti oxi defense would lead to this STRESS

-when there is oxi stress, biomolecules of tissues and organs are OXIDIZED (PROTEINS, LIPIDS, NUC ACIDS)
CONSEQUENCE BECAUSE:
Proteins make up many active molecules and structures
Lipids make up organelles and cell mems
Nuc acid: DNA, damage to this (nuc or mito) can impact genes for cell functions

Question 10

Names of the stresses that ROS put to the different components

Answer 10

LIPIDS: Peroxidation
-ros attack fatty acids
-creates lipid radicals and lipid hyperoxides
-damages membrane, leaky and dysfunctional

PROTEIN: denaturation
-loses 3d shape, h-bonds and disulfid bonds are disrupted
-cant funxtion

Enzymes: Inactivation
-no longer catalyze reaction because structure changes, AA messed up, active sites messed up

Nucelic acid: Modification
-base changes
-strands break
-inducing mutations
-sugar backbone altered

ALL THESE DAMAGES LEAS TO diseases, poisonings, aging

Ways to fix
Protein Turnover
Dna Repair
Lipid Repair

Question 11

CELLULAR SOURCES OF ROS AND RNS

Answer 11

1) MITOCHONDRIA: ELECTRON TRANSPORT CHAIN
remember: e move through chain and are accepted by o2 to make h20 at complex IV (final step)
-normally o2 undergoes 4e reduction, but sometimes e- leak out of ETC and incompletely reduce o2
-2-3% of total o2 used by mito is incompletely reduced, lthoguh this is only a SMALL fraction of total o2 utilized, this can be a significant source of ROS in METABOLICALLY ACTIVE TISSUES
-MAJOR SOURCES OF SUPEROXIDE FORMATION HERE IS complex 1 and Complex 3, greater e flux (movememnt) meanign greater superoxide formation; this is because e can only move when the one infront of it moved, so if there is too much e- it will remain where it is meaning that it leaks out and forms ROS
-more notes for this on next card

2) NAD(P)H OXIDASE
-nicotinamide adenine dinucleotide phosphate oxidase
-6 subunit complex in MEMBRANES
-makes O2- by catalyzingtransfer of electron from reduced NADH / NADPH to OXYGEN
-found in IMMUNE CELLLS (neutrophil, macrophage; funx in host defense by killing microbes using free radical damage), smooth muscle and Skeletal
-COMPLEX IS EMBEDDED IN MEMBRANE AND IT TAKES E FROM NADH OR NADPH AND IT DONATES IT TO o2 to form O2-
-NOT IN ETC, SOLE JOB TO MAKE SUPEROXIDE
-IN IMMUNE CELLS: lysoszyme holds the NADPH oxidase, phagocytosis the bac and then this occurs

3) XANTHINE OXIDASE
-cytosolic enzymes that OXIDIZE XANTHINE AND HYPOXANTHINE to make O2*- and H202

-also participates in cell signalling, oxidizes proteins to lead to changes in proteins

4) NITRIC OXIDE SYNTHASE
-CATALYZES oxidation of L-arginine to L-citrulline to release NO*
a) Neuronal NOS (nNOS)
FOUND IN skeletal muscle, localized to sarcolemma and associated with DYSTROPHIN

b) Inducible NOS (iNOS)
-macrophages and aid in host defense

c) endothelial NOS (eNOS)
-endothelial cells, and aids in vasodilation

Question 12

What are sources of Free Radicals and Reactive Oxygen Species

Answer 12

UV LIGHT
RADIATION
SMOKING
INFLAMMATIN
METABOLISM
AIRPOLLUTION

Question 13

Mitochondrial Content and Function Influence ROS levels

Answer 13

Cytochrome C: in ETC, it moves electrons from complex 3 to 4…
when it is removed: more h202 is produced (Superoxide is produced)
When it is added back: reduced production, but still more than when it was originally depleted

2) Increase mito content:
-in a cell we produce a spcific amount of atp and that is associated with a sepcific number of H that have been pump and e- that is moved.
-if we have low mitochondria, we are making less atp because less ETC
-more mito means more ETC
-regardless of if we have low or high mito ina. cell, the ATP NEEDED BY CELL IS SAME, so the same number of e is needed ot be moved, HOWEVER< when there is more mito in a cell, each individual chain moves less electrons, so they ae distributing electron movement across many chains so it is more efficient and LESS LEAK
-so more etc means LESS ROSb because electrons are moving efficiently
-think factor assembly line, if we need to make alot of boxes but only one line of workers, more mistakes because we have to move faster, but if we have multiple lines with multiple things, each line makes less so they can work more better and efficiently

THEREFORE: for a given rate of oxi phos, decreased mito CONTENT and FUNCTIOn increases ROS generation, while increased content and function of mito decreases ROS

Question 14

how do immune cells like neutrophils and macrophages kill using free radical damage

Answer 14

NADPH OXIDASE
-makes o2*- from NADH AND NADPH

Neutrophil: defense of cell by killing bacteria , neutralizing microbes
PHAGOCYTOSIS: it engulfs microbes and inside the neutrophil is a lysosome which holds NADPH oxidase, and it makes superoxide and this will kill the bacteria by oxidzing the bacteria

Question 15

Main difference between NADPH OXIDASE AND XANTHINE OXIDASE

Answer 15

XANTHINE:
cytosol
uses xanthine and hypoxanthine (oxidizes this)
makes superoxide AND h202

NADPH:
membrane
uses NADH or NADPH

Question 16

ANTIOXIDANT ENZYME ACTIVITY IN SKELETAL MUSCLE

Answer 16

ANTIOXIDANT ARE HIGH IN SLOW , MIXED and FAST MUSCLES respectively

regardless of the antioxidant, it is higher in slow (type 1)

WHY?:
-used more, so more antioxidants
processes happen all day long in these muscles, so assocaited with more cell signalling so more ROS so more Antioxi to
-essentially more signalling means more ROS because it is a bioactive molecule that also helps with signalling and this means that it will induce the gene expression of ANTIOXIDANT ENZYMES so more of them

Question 17

when we need to target something

Answer 17

we need to target it from the source it is coming from

ie if it is in mito, we nee to target the superoixed there

Question 18

ROS REDUCES CELL REPLICATION and INDUCED DNA DAMAGE

Answer 18

SOD deficient animals (antiox enzume defiecient) have increased oxidative damage to DNA

-KNOCKING IT OUT INCREASES DNA DAMAGE IN YOUNG AND OLDER ANIMALS (age therefore does not matter), older animals actually have more dna damage
-uprefulaed ENZYMES so higher enzymes levels

Satellite cells:
-sensitive to ROS
-satellite cells in a dish with H202 shows reduced DIVISION in plate
-number of divisions decreases in the presence of h202

Question 19

Oxidative Stress in Aging Skeletal Muscle

Answer 19

-INCREASED ANTIOXIDANT ENZYME ACTIVITY (SOD, CATALSE, GPX, GST (neutralized by conjugate with GSH, GR (regenerates GSH))
-because increased ROS with age
-mito of aged produces more O2*- an H202
-even though there is an increase in the enzymes, not sufficient enough to deal with increase in ROS production
-net increase in oxidative stress to proteins, lipids (membranes) and DNA

Question 20

Oxidative Stress in satelitte Cells

Answer 20

more lipid peroxidation in aged versus young samples

-Malondialdehyde (MDA is a by production of lipid peroxidation); marker of oxidative damage to lipids is increasedin aged samples)
-more malondialdehyde in aged sames mwans more lipid peroxidation in aged samples

Question 21

Mitochondrial ROS in Mature Muscle

Answer 21

-basal mitochondrial H202 release is elevated in mito from aged skeletal muscle

-ie more h202 is made in aged muscle
-measure of superoxide production from ETC

-more dysfunctiona;

Question 22

Small amount of ros is good because

Answer 22

allows for cell signalling

but when the balance becomes off, this is bad becaue it means that we will have more oxidative stress and DAMAGE to these components

Question 23

REDOX STATUS AND OXIDATIVE STRESS IN OLDER ADULTS

Answer 23

Older adults were taken and seperated into 2 groups
1)More than 40% type 2 fibers
-normal muscle, not experiencing age related changes

2) less than 40% type 2 fibers
-muscle phenotype changes based on ages
-more lipid peroxidation
-so more likle yhtat these are associated with changes in muscle, so it might be influencing these changes
LPO levels are measured (lipid peroxidation)

-for individuals with less type 2 fibers , they have more lipid peroxidation (higher levels)

Question 24

ETC COMPLEX DEFICIENCYoccur in muscle during aging

Answer 24

Aged mito are : fewer and dysfunctional (make less atp)

THEREFORE: deficiency leads to increased ROS at mito an increased muscle oxi damage

ETC proteins decrease in AGED mito:
-so we essentially have fewer chains so we need to use this to make the same ATP that we usually do, so we have more ROS, because high traffic chain
-DNA damage in age muscle is MORE

SO DEFICIENCY IN ETC and PROTEINS lead to less proteins, more damage, more oxi stress that leads to tpye change

PCG-1 GENE: induce more biogenesis, so in aged mice with high PGC-1 gene, there is more ETC proteins

Question 25

upregulating

Answer 25

PCG1 helps with mito dysfunction

Question 26

ANTIOXIDANTS and ROS GEN in DISUSED MUSCLES

Answer 26

Lower levels in antioxidant enzymes for DISUSED muscles -level of activityh help maintain antioxidants at a specific level -as we disuse, we have less signalling, so adaptations that have happened due to signalling is dropped -INCREASED h202 in disused, because fewer antioxidants -INCREASED OXIDATIVE STRESS (high DCFH) SOO if there is a dsease that targets ros generation, and we dont use the muscle, it will make it even worse for you

Question 27

MUSCLE CONTAINS REDOX SENSITIVE SITES IMPORTANT FOR CONTRACTION

Answer 27

-abilityn to generate ros is CRITICAL for signalling and if we take it away, cell does not function Skeletal muscle has proteins that need to be oxidized for them to work properly: so it has NOS, NADPH oxidase and XANTHINE OXIDASE which source ROS -there are enzymes that are impacted by ROS generation ie NO inhibits SERCA ROS inhibits actin and myosin heavy chain etc What are the ways in which ROS is needed: 1) ROS/RNS and OPTIMAL FORCE GENERATION -2 conditions: control and antioxidant supplemented condition -individuals perform exercise ONE LEGGED In ControL: exercised leg gets increase in lipid oxidation IN antioxidant one: lowered oxidation in (no oxidation, ros is being made but it is being quenched before it can oxidize lipids ) -FORCE GENERATION lowered in antioxidant condition because we need some of that oxidarion to stimulate enzymes for optimal muscle contraction FORCE GOES DOWN . this means that when we conrtact normally, we make ROS that stimulates enzymes that make muscle contract optimally. Maximal force that we produce reduces when we use ANTIOXIDANTS, slower relaxation SMALL LEVELS OF ROS: increase musclw force, ros is needed!!!!!!! Maintaining OPTIMAL cellular redox Status may influence FORCE: -over oxidize: performance drops -less oxidize:Nreduced performance -so narrow margine for the optimal function that we must keep

Question 28

Regular exercise positively infleuncies muscle redox status

Answer 28

-aerobic training makes system more effiecient at making atp ANDDD stimulates antioxidant defense systems (positive adaptation), makes ros gen dealing with it better -Following training: basal ROS generation is also reduced (GOOD) -everytime we run we produce ROS, the more we do the better our systems are at adapting tothis and deealing with it ANTIOXIDANT PROCESS: balances it, we want to keep these levels goo

Question 29

Oral Vitamin Intake Prevents Exercise Induced Antioxidant Adaptations

Answer 29

-usually when we train muscle, we get an increase in the experssion of antioxidant enymes HOWEVER: when we supplement with antioxidants such as VITAMIN C and E, we get less increase in the expression of antioxidant ezymes, so the adaptations we usually see (positive adaptation) of exercsie increasing this is not seen). This means that our body is not learning to balacne out the oxidant levels on its own. MnSOD and GPX double with training, but do not ncrease at all when exercise is done with vitamin C intake

Question 30

ROS SINALLING: critical in exercise induced mitochondrial biogenesis

Answer 30

PGC1 increases mitobiogenesis When training, PGC1 increases, and this leads to increased mito when training with VitaminC or E: PGC1 levels do not increase, or barely increase... meaning MITOBIOGENESIS IS NOT HAPPENING . Same thing happens with NRF (helps makes proteins for etc) and MTFA which is neede for mito dna and ensuring it can make its own)

Question 31

What factors influence these positive ros induced adaptaions

Answer 31

REMEMBER pos ros adapations are 1) Increased expression of antioxidant enztnes 2( decreased ROS IxB: inhibits NFkB p-IxB: allows NFkb to go NF-kB: activates antioxidant genes

Question 32

ROS and Exercise indicued signalling

Answer 32

Allopurinol: inhibits Xanthine oxidase which makes ROS so when exercise but have this we hav lower levels of NF-kb and P-erf which are factors that upregulate the anxiodiant enzyme genes

Question 33

What factors influence positive ROS induced adapations relative to disease and aging

Answer 33

1) transient increase in ROS generation and signalling -short term invreases in ros will genereate good signallings for adaptations 2) Lower absolute levels of ROS -lower levels prevents damage to things -in aging, baseline ros is higer so already leads to damage 3) ROS/RNS source, Site of genereation, type of ROS/RNS -dysreg in wrong sources leads to more problems. IE mito and NADPH ros may increase gene expression of antiox (NADPH) and mito biogenesis (mito) XAnthine ROS: can cause damage

Question 34

Oxidative stress and Muscular DYSTROPHY

Answer 34

LIPID (MDA) and Protein (carbonyl) oxidation (oxidative stress) are increased in uscles of Muscular dystrophy mice -exercise provides some protection for this

Question 34

SOOO ROS: good bad or fine in moderation?

Answer 34

Some levels needed for signalling, muscle function optimization, force production -exercised induced adapations are dependent on ros (ie increased antioxidant enzymes and decreased ros are dependent on there being a suffieicient amount of ros present); think the experiements with vitamin C and E, it reduced this so no adaptations were formed -therefore; altering basal and exercise induced ros may be detrimental to muscle function and adaptations -need more work to understant the influene of higb antioxidant diets on long-term cellular adaptisons and signalig, as well as suceptibility to stress and disease

Question 35

Slide 52

Question 36

section 7

Answer 36

Apoptosis and Cell death in skseletal muscle

Question 37

Cell Death: Necrosis:

Answer 37

2 main types of cell death: 1) necrosis -accidental -occurs during disease, or acute injury only. not part of NORMAL DEVELOPMENT -results in: rapid cellular swelling and rupture of cell membrane -leads to: major inflammatory response due to the release of intraceullar materials into surrounding environment 2) apoptosis -highly conserved cell ddeath -important process of normal tissue development and homeostasis -increased or decreased in disease states -regulated by intracellular signalling -RESULTS IN: chromatin condensation, cell shrinking, membrane BLEBBING (plasma membrane does this) -cell membrane remains in tact, little to no inflammation therefore What does CELL BLEBBING DO -packs the cell for removal sooo 1) dna damage, withdrawl of growth hormones, extrisinc cell death signals (TNF), cytotoxic lumphocytes) all issue pathway 2)excectuioner caspases activated 3) endouclease action and breakdown of cytoskeleton 4) bud forms 5) phacogytic cell receptors on outside of bleb bring it to phagocyte which eats it

Question 38

essentially

Answer 38

Necrosis: compromised membrane, cell swelling, release of intracellular components, inflammation response Apoptosis: cell shrinkage chromtin condensation cell blebbing blebs get into phagocytosed by phagocyte an other immune cells -controlled

Question 39

BASICALLY for blebbing to occur

Answer 39

1) executioner caspases activated by various things: dna damage, extriscinc signals, mitro pathway, cytotoxic lymphocytes 2) breakdown cytoskeleton (actin and other things) 3)membrane detaches 4) found by phagocytes

Question 40

Cell morphology during Apoptotic and Necrotic cell death

Answer 40

Apoptotic: lots of blebs outside necrotic: lookes like stringy, damages from inside out seperarting very stringly

Question 41

Common features of Apoptosis

Answer 41

DEFINING FEATURE: "HALLMARK" : fragmented DNA ALSO breakdwon of structural, regulatory and repair proteins

Question 42

look at slide 8

Question 43

Apoptosis Research

Answer 43

-one of the most popular research areas in science and medicine 149241

Question 44

Apoptosis during development and disease

Answer 44

A) Lumen formation (Development): cells in the center go through apoptosis to create a hollow cavity -needed for proper tissue and organ creation B)Sculpting of digits: DEVELOPMENT -apoptosis removes webbing that leads to fingers and toes -creates form c)Tail of Tadpole when frog -tail goes thorugh apoptosis to removeit once it becomes a frog SO FOR DEVELOPMENT: apoptosis is precise, allows for formation of structures, scult organs, remove unnessecary items Disease: Stress causes it to shrink and be removed (cell) Neurodegenerative diseases (too much apoptosis → neuron loss) Heart failure (cardiomyocyte apoptosis) Diabetes complications Aging-related tissue atrophy -may be excessive, leads to damge

Question 45

Apoptosis : NOT ENOUGH

Answer 45

-inhibition of apoptosis leads to cell ACCUMULATION -this leads to UNDESIRALE TISSUE GROWTH; and in this case an accumulation of immune cells and immune disorders Bax and Bak: pro-apoptotic genes that execute mitochondrial mediated apoptosis by permebealizing the mitochondria membrane --sit on outer mito membrane and when they recieve signal, they make holes in it which causes cytochrome C to come out and start caspase event SO when bax and bak are inhibited, apoptotic signalling is inhibitted, so cell accumulation happens so extra large things are formed

Question 45

Rate of Apoptosis on Tissue homeostasis

Answer 45

If rate of cell prolif is high but rate of cell death is low: DISORDERS OF CELL ACCUMULATION If rate is equal: Homeostasis If rate of cell prolif is LOWer than cell death: Disorders of cell LOSS

Question 46

Apoptosis: TOO MUCH

Answer 46

-promotion of apototis leads to cell loss -if toomuch leads to undesirable tissue atrophy, retarded development, decreased immune function and premature death BCL-2: member of the bcl2 family that PREVENTS bax and bak from permeating the outer mito membrane by binding to them -when this is knockout, it results in too much apoptosos

Question 47

Diseases assosciated with decreased apoptosis

Answer 47

-cancer -cardiovasc disease -autoimmune disease -Crohn's diease, ulcerative collitis

Question 48

Diseases associated with Increased APOPTOSIS

Answer 48

-diabetes -neutrodegenerative diesease -hiv -viral andbac infec =osteoporosis -neuromuscular disorder -sarcopenia, atrophy -heart disease, stroke, hypertension

Question 49

BCL-2 Family Proteins: regulators of apoptosis

Answer 49

-first identified as a gene that alters growth and cell death in B-cell lymphoma (BCL-2) -25 BCL-2 family proteins identified -all have similar structures and functions to inhibit or promote cell eath in tissues -regulate organelle and cell function by acting on various membranes (mito, ER/SR, nuclear) 2 main classes: 1) Anti-apoptotic: BCl2, BCL-x1 2) apoptotic: Bac, Bid, Bak ratio of anti to pro proteins is used as a measure of cells susceptibility to apoptosis

Question 50

CASPASES

Answer 50

-executers of apoptosis -CYSTEINE ASPARTIC ACID PROTEASES -14 identified -found in inactive (pro form) and active form 2 main classses: 1) INITIATOR: (caspase 1, 8 , 9 ,12) 2) effector: 3, 6, 7 -cleave numberous cellular substrates that results in apoptosis (400+ protein targets) including a) ACTIN b) Poly ADP ribose polymerase (dna repair enzyme) c) lamins (nuclear structure proteins)

Question 51

Other important Apoptosis Regulatory Proteins or Factors

Answer 51

INDUCERS: 1) Increased Free radicals, ROS, RNS 2)Chemotherapeutic drugs 3)High stress hormones (glucocorticoids, catecholamines) 4) Increased cytosolic Ca2+ levels 5)UV, Radiation, ethanol, growth factor withdrawl, nutrient deprivation INHIBITIORS: 1) antioxidants (SOD, gluthione, catalsae, vitamin C n E 2) Growth factors (IGF-1, GH)

Question 52

WAYS WE ACTIVATE CELL DEATH

Answer 52

A) Death Receptor Pathway Relies on receptors on cell membrane -Cytokines FasL or TNF-alpha activate caspases 1) TNF binds to TNFR on membrane and this activates death receptor pathway (extrinsic) 2) signalling 3) results in activation of caspase 8 (initiator caspase) 4) caspase 8 activates caspase 3 (effectir caspase) 5) caspase 3 cleaves substrates (Cytoskeletal proteins, dna repair enzymes (polyADPribose polymerase), nuclear proteins (laminns) B) Mitochondrial Pathway (CASPASE DEPENDENT, CYTO C) -cytochrome C activates Caspases -remember mito houses 1500 proteins, it is not all involved in metabolism , some for apoptosis 1) in response to various stresses, pore formation by bax and bak occurs, at mito outer membrane, read flashcard57 2)proteins housed here leave, ie CYTOCHROME C, not anchored at etc 3)now in cytosol, it acrivates apoptotic pathway, formes activae complex which is caspase 9 4) Caspase 9 activates caspase 3 5) caspase 3 cleaves substrates Another proteins that leave when mito membrane is pored is AIF and ENDO G, these dont activate caspase, but transclocate to nucleus and begin breakding down DNA (REMEMBER HALLMARK OF APOPTOSIS IS FRAGEMENTED DNA) C) ER/SR STRESS PATHWAY: -CA is stored in ER/SR. -Calcium release is regulated, however in response to stressors, there is stress at ER/SR, so there is sustained ca at cytosol, whicb activate m-CALPAINS. -CALPAINS ACTIVATE CASPASE 12, qhich activates caspase 3, and then we get cleavage of substrates

Question 53

SO WHAT ARE THE DUAL ROLES OF MITOCHONDRIAL FACTORS

Answer 53

1) Cytochrome C : MITO: shuttles e- from comple 3 to 4 APOPTOTIC: Apotosome formation and caspase activation PATHWAY: Caspase-dependent 2) ENDONUCLEASE G: mito: mito biogen apop: direct dna fragmentation pathway: caspase independent 3) AIF (Apoptosis inducing factor): mito: proper function of COMPLEX 1 Apoptotic: direct DNA fragmentation Pathway: caspase independent ROLEin complex1 activity: 1) aids in assembly of complex 1 (46 NADH DEHYDROGENASE COMPONENTS) 2)may direct aid in electron transport to complex 1 by acting as NADH OXIDASE (DONATES ELECTRONS TO COMPLEX) therefore altering these locations is based

Question 54

Consequences of CYTOCHROM C and AIF release from mito

Answer 54

1) Formation of Apoptosome (caspase 9 complex), For AIF release: translocates to nucleus and degrades DNA 2) Less ATP because less cytochrome C moving e from 3 to 4 an AIF is not shuttling it to complex 1 or complex 1 thing is not properly formed. SO less e means less H pumping, so less atp through atp synthase 3) Increase ROS generation because cytochrome C is not shuttling proteins. More electrons will leak due to this. AIF will cause leake at complex 1, cytochome causes leak at 3 4) OVERALL CELL DYSFUNCTION AND DEATH.

Question 55

MECHANISMS OF MITOCHONDRIAL PROTEN RELEASE

Answer 55

ONE WAY: BAX PORE BAX: found in cytosol NORMAL. IN response to apoptotic signalling it will move from cytosol to outer mitochondrial membrane, and forms a complex with BAK. THIS FORMS PORE. NOW CYTOCHROME C AND AIF AND ENDO G CAN LEAVE BCL2: inhibits BAX from inserting itself on to OUTER MITO MEMRBANE. BALANCE IS IMPORTANT because it means homeostasis, soon as one is more, then something occurs SECOND WAY: PERMEABILITY TRANSITION PORE VDAC (voltage ependant anion channel) ON OUTER MEM and ANT (adenonine nucleotide translocator) ON INNER MITO MEM -usually has good control on what leaves and enters, however in response to stress, BAX moves from cytosol to VDAC and embeds itself and this changes the conformation fo VDAC an d this lines up the two channels (VAC AND ANT) that makes the mito more permeable which allows more ions and water to enter mito more rapidly and this is a problem BECAUSE INNER MEMBRANE HAS LARGER SURFACE AREA, SO as inner membrane is filled, and this will burst holes and rupture outer membrane, so then things move out through this hole including cytochomr C and AIF. BCL TRIES TO STOP THIS TOO.

Question 56

ER/SR Stress PATHWAY

Answer 56

C) ER/SR STRESS PATHWAY: -CA is stored in ER/SR. -Calcium release is regulated, however in response to stressors, there is stress at ER/SR, so there is sustained ca at cytosol, whicb activate m-CALPAINS. -CALPAINS ACTIVATE CASPASE 12, qhich activates caspase 3, and then we get cleavage of substrates

Question 57

Apoptotic signal and apotosis in skeletal msucle

Answer 57

A)different mito content in each muscle fiber type, some have more , some less B)muscle specific apoptotic proetins and expression patterns depend on fiber type too -ie ARC. (antiapoptosis protein in death receptor and mito pathway) -HIGH IN SKELETAL MUSCLE but absent in other healthy tissue -also expressed differently across muscles (more in type 1) C)multinucleated, so fibers will undergo MYONUCLEAR apoptosis rather than complete Cell apoptosis -remmeebr in normal apoptosis AIF and endo g go to nucleus and fragment dna and this causes cell to die, however for muscle there are so manynnucleus so this is going to beb a. bit different -small number will therefore get fragmented, not all of them so entire cell does not die. so MYONUCLEAR APOPTOSIS -basically, there will be cell death signals in the cell , and this will go and target loss of SPCIFIC nuclei and when this is targeted we will lose that NUCLEI and the CYTOPLASM THAT THAT NUCLEI IS ASSOCIATED WITH ( CALLED NUCLEAR DOMAIN) -leads to muscle atrophy and dysfunctional muscle

Question 58

What is the NUCLEAR DOMAIN

Answer 58

area that that nuclei is responsible for when cell death happens in muscles, this area dies too

Question 59

SOOO essentially myonuclei apoptosis leads to myscle atrophy because

Answer 59

THINK: healthy muscle has a certain number of myonuclei that support a portion of cytoplasm (nuclear domain) -this nuclei number determines how many proteins are made, such as myosin and actin and this results in muscle size -so if a muscle nuclei dies, due to apop, we have reduced myo number, and therefore reduced muscle proteins and the muscle now needs to adapt because cant be bigger cuz we dont have enough proteins so it becomes smaller , so remodelling helps maintiain Nucleus: cytoplasm RATIO SO ATROPHY]]]] so eventually keeps loosing until we lost so much nuclei that it can no longer support the fiber and it will die then only, so myonuclear apoptosis does not always lead to cell death.

Question 60

Myonuclear Number and Fiber Cross sectional area

Answer 60

- alothough entire cell (fiber)may not be lost due to apoptosis in muscle, loss of nuclei is an important factor to muscle atrophy -number of nuclei dictates fiber XROSS SECTIONAL AREA -more nuc, more area -same for each fiber type

Question 61

Skeletal muscle Apoptosis and Oxidative Stress

Answer 61

ROS can induce skeltal muscle apoptoisis therefore APOPTOSIS xan be inhibited by Vitamin C (ascoribc acid) more ros for longer time, more aposotisis CHRONIC (long time exposure, bad) VS ACUTE (samll transient oses of ros its fine)

Question 62

MTDNA mutations lead to increased skele muscle apop

Answer 62

Aged muscle: higher levels of CASPASE 3 compared to young Young mice with accumulated MtDNA mutations have high Caspase 3 too SOOOOO mutations in mitodna leads to more caspase 3 , and aged muscles with no disease aslo leads to caspase 3 mitochondria dysfunction leads to etc dysfunc leads to ros leads to apotosis leads to aging phenotype

Question 63

apop in skel muscle satellite cells during AGING

Answer 63

Satellite cells (MYOD cells) in old animals have higher BAX than young and lower BCL2 Older animal satellite cells are also more suceptible to TNF-alpha induced apoptosis (death pathway, caspase 8 to 3 pathway) this means that they are dying easier which also contributes to why aginign muscle is less nuceli because satelitte cells which are the machinery to get back nuclei have more bax less bcl2 and more suceptibility to death so the machinery to make up for nuclei dies faster too so this is why we get more muscle wasting and atrophy because we are losing it through apoptosis of normal nuclei and paotosis of the satelitle cells in general, the satellite also dont fuse or proliferate as well with age

Question 64

Skel Muscle Apop in DISUSE

Answer 64

apoptosis is high in aging and even more higher in inactivity -increases apoptosis means decreased CSA and weight

Question 65

Apoptosis Regylates muscle mass in muscular dystrophy

Answer 65

-mice lacking LAMININ have CONGENITAL MUSCLE DYSTROPHY -inhibition of BAx (apoptosis) reverses muscle loss, functional decline and early death essentially cogenital muscular dystrophy means laminin is lost, when the mixe has low laminin and high BAX (meaning apotosis) we see alot of muscle loss but when there is no laminin and NO bax (apop inhibited) we see a decreased in muscle loss, function decline and early death

Question 66

Skeletal muscle apoptosis ELEVATED IN CHF

Answer 66

Congenital Heart Failure: altered MHC expression whicb means lowered BCL2 so increased caspase 3 and myonuclear Apoptosis *TUNEL, dna fragmentation is tunel this is higher) -these people with apopotoic pos skeletal muscle has 34% reduction in VO2max even though heart function not different -muscle undergoin much more dysfunction so this is just lower, -more apoptotic nuclei means less cross sectional area for CHF pateint

Question 67

Exercise protection against muscle disease and dysfunction by decreasing apoptosis

Answer 67

exercise: increases levels of antiapoptotic proteins such as BCL2, ARC, and XIAP in healthy, aged, hypertensive rats -it decreases: myonuclei apoptosis, and leads to positive morpholigcal adaption in old and hpyer tensive rats IN HUMANS: moderate regular exercise leads to less TNF and FAsL in CHF patients. (propapototic signalling things) -if we detrain, it will go back to whart it was pretraining EXERCISE ALSO: increases mitochondria biogenesis so we push it to be better even in how etc works Mitophagy:exercise activates this, finds and degrades dysfunctional mito, so SO WHY IS EXERCISE SO GOOD: 1) makes more mito, 2) removes dysfunctional mito (mitophagy) so not only does exrecise lower apoptotic signalling and incrrease non apoptoci signallonglike BCL2, ARC and XIAP, it also makes more mito and gets rid of bad mito

Question 68

Mitochondrial Pathway snd how exercise helps this

Answer 68

When Protein stress or damage occurs: 1) ca is reesase at high levels, it is relased around mito (not to contractile aparatus for this) 2) causes swelling of mitochondria because it enters it 3) permeability tranisiton pore if formed (Vdac and ANT pore) 4) release of factors 5) caspase activation and dna fragmentation OR M-calpains get activated and this activates caspase 3 and this cleaves substrates on sacromere (reiterated ways to engage cell death) but with exercise: induce mito biogenesis -so more mito so more ca can be taken up tha is releaae due to stress, so now we distribute it so the mito doesnt swell so pore is less likely to be formed. load is distributed. less in cytosol so less m-calpain is activated + anti apop proteins are reduced, so this akes an environment of cell that is less susceptible to cell death signalling

Question 69

Summary

Answer 69

Aging dsease all of this lead ti change in potein patterns that is much more pro apoptotic so this increases pore formation and ros gen, release of proteins that activate cell pathways -downregulat factors b=promoting mito bio, ros generaton due to etc non function LOOK at slide 43

Question 70

lecture 8

Question 71

Protein Homeosstasis and Protein Turnover

Answer 71

-for cell function need proteins -made and degraded for proper function however rate of degredation is important , dysregulation can increase or decrease this rate Abnormal proteins that are rapidly degraded in the cell: A) INcomplete: nonsense mutation B) missense: mutations C)Free subunits of multimeric complex: excess subunits d) Postsynthetic damage: ROS e) Genetic engineering f) Misfoled:

Question 72

Ubiquitin Proteasome System (UPS)

Answer 72

eals with the proteins -responsible: turnover of short lived, damaged or misfolde proteins -recognizes target protein by the presence of poly-ubiquitin chain PROCESS INVOLVED 3 main enzymes: 1) E1: Activation of pthway 2) E2: conjugation 3) E3: ligation SYSTEM PATHWAY: DAMAGED DENATUED OR MISFOLEDED PROTEINS -we dont want these proteins to accumulate because they will go to ER to try and be dealt with here, they will acccumulate and lead to ER stress. This leads to Ca release, we dont want ER stress so we want to make sure these proteins dont accumulate so we degrade them 1) Chaperone proteins (HSP40 and HSP70, heat shock protein) find denatured and damaged proteins 2) ubiquitin chain is made by E1 binding to ubiquitin with ATP 3) Ubiquitin conjugation occurs and the chain is added on 4) Proteasome has protelytic enzymes on inside, it can recognize ubiquitin chain on protein and take it in, this will degrade the proteins into peptides and amino acids which can be used to make other proteins SPECIFIC PATHWAY: 1) UB is activated by E1 (ubiquitin activating enzymes) 2) UBis then transfered to ubiquitin carrier protein (E2: ubiquitin conjugating enzyme) 3) Proteins to be degraded are found by e3 m, these proteins have chaperone proteins (hsp40 and hsp70), and e2 is akso brought to e3 (ubiquitin ligation enzyme) and both bind 4) Activated structure transfers ubiqutuin to protein, as it forms a scaffolding with e2nubiqutiin and the protein 5) now it acts as a signal oto target the protein for degreation by the 26S proteasome

Question 73

Ubiquitin-proteasome system and pathophysiology

Answer 73

Pathological states (diseased states) arise if UPS system does not function properly, number of them arise from dyfunctional ups activity INcrased UPS ctivity: accelerated protein degredation Decreased UPS: stabilizaion of certain proteins, or inhibition of 26Sproteasome resulting in reduced degredation

Question 74

Accelerated removal of spceific proteins in cancer

Answer 74

-too much degredation p53. is a tumour supressor gene, it supresses tumors Control Condition: p53 levels low in cancer tissues (means cancer is growing faster than this) Result: little to no ubiquinated p53 because proteasome is working V Condition: cells treated with something irrelevant (carrier of drug) MG132 (drug): proteasome inhibitor. binds to it and doesnt let it do its work, this is preventing protein degredation. RESULT: HIGHER P53 levels, this stops p53 from being degraded so allows ubiquinated p53 to build up SOOO: this cancer lives by increasing the rate of p53 degredation, therefore if we block proteasome activitym ubiquinated p53 increases, and since this is not being dealt with, normal p53 is still there too

Question 75

Inhibition of Proteasome Acitivty promotes Cell Cycle Arrest and Decreases Tumor Size

Answer 75

-too much degredation MG132: does not cell cycle impactor but it makes the cells in G1 (they are arrested, theya re not making to g2), so they are not proliferating -mg132 didnt do this directly, it just stoped p53 from being degraded so it is able to do its job better because it is now able to reduce the amount of cells that proliferate so more stay in g1 SO WE ARENT TARGETING HE GROWTH we are targeting what is stopping the growth from stopping -tumour size reduces because proteasome inhibition allows drug to do its job We dont have to target the disease or wha is going wrong, we can target proteins in the process thatcan help us have an effect... anotehr way we could have done this then is by increases p53 transcription for esample , or introduce something that increases p53, but this way we just stopped its degredation which normally occurs in cancer which allows for cell

Question 76

ACUMMULATION of protein aggregates contributes to neudegeneraive disease

Answer 76

-too little protein degredation goin on HALLMARK FEATURE OF HUNTINGTONS: accummulation of proteons (specifically, increase in glutamine on HTT protein (huntintons protein) -leads to formation of iniclusion bodies which are toxic and kill cells -large globs in cells that kills neurons REASON: proteasome system is not goo at dealing with the accumulation of HTT proteins -essentially proteasome enzymes activity is lower which makes these proteins accumulate and inclusion bodies form -when we inhibit proteasome more, it makes more inclusion bodies so the rate is really impoetnat -if we activate proteosome, it clears out the aggregates at much faster rate

Question 77

UPS system in skeletal muscle specifcallhy

Answer 77

2 main E3 ligases in skeletal muscle 1) MuRF1: muscle ring finger protein 2) MAFbx: atrogin 1 (muscle atrophy F box) These target the polyubquit of proteins that target them for proteolysis by proteosome - MURF1 and MAFxb : regulated by myostatin/TGFB 1) Signals like insulin and IGF activate pathways that drive up protein syntehsis through MTOR pathway. -level of e3 ligases is low, so these transcription of it is relatively low. So protein degredation is relatively low -therefore synhtesis or muscle is more than degredation, so HYPERTROPHY In response to diseaase states, and pathological onditions: Protein synthesis still happens but the rate of it is reduced. Caspaes ad stuff are also activated, or transcription of e3 ligases are increased so rate of degredation goes up. -imbalance so ATROPHY balance between protein degredation and synthesis depends on if musclew will remain same size, increase or decrease.

Question 78

UPS and Muscle Atrophy: denervation and Immobilization

Answer 78

Inactivity in muscle: muscle weight decreased through hind limb suspension, immobilization and denervation -also see more murF and maFBX too , more e3 ligases SO FOR MUSCLE ATTROPHY: denervation TO OCCUR: must have an increase in MurF1 and MAFBX. -Wasting inresponse to stimulus if these genes are gone are SIGNIFICANTLY LESS

Question 79

So what genes are important for muscle atrophy via denervation

Answer 79

MAFBx and MurF1: e3 ligases

Question 80

Longer Periods of UPS inhibition can preserve MASS but result in muscle damage

Answer 80

WILDTTPE: loses mass in response to denervation if u knockout murf or mafbx: muscle mass is preserved much better (makes sense because protein degredation pasthway is inhibitied) HOWEVER: muscle that is preserved looks weird: fiber sized are bigger than wildtype (less protein degredation so more protein in muscle) however some of the muscle does not look good, spaces and vacuoles present -muscle damage is seen: beacuse accumulation of damaged proteins in muscle, some are ones that should be turnover

Question 81

So what does knocking out MUrf1 do specifically in aging

Answer 81

1) preserved muscle mass (fibers and stuff because these bad proteins not being taken out) BUT does not preseve function or caspase activity during aging -fiber area is higher (so mass is preserved) -Caspase activity : increased -force generation: decreased . so even though it has more muscle mass, it has decreased force because the mass is stuff that we really should be getting rid of -levels of oxized proteins are also higher in knockout mice because damaged proteins are not being gettin rid of so they are accumulating in ER and this leads to ER stress which activates caspase 12 through m-calpain and calcium release

Question 82

Balance between enough and too much degredation is really important

Answer 82

-have to be in optimal zone

Question 83

AUTOPHAGY

Answer 83

Self Eating degredation fo LONG LIVED proteins, portions of cytosol and organelles (oppsotie of UPS: ups was shortlivwd, mifolded or damaged proteins proteosome can only fit proteins because it is a tube) -works with UPS to remove damaged stuff -not limited by size of protein or structures like how the UPS is -plays role in metabolism (breaking down into amino acids, fatty acids, lipids, carbohydrates, can be used as fuel source) cell cycle, differentiation and death -key role is deatg, promoting cel surivval by clearing damaged proteins and organelles -gets rid of cell stress too 3 main types a) Micro-autophagy b) chaperoe-mediated Autophagy c) macro-autophagy (usually this, most common) DEFINING FEATURE: development of double membrane vesicle -CALLED AUTOPHAGOSOME -main machinery, signals in cell produce this double membrane vesicle , inside it are vaarous cellular components that needed to be degraded (so think if these are stress signal, it is now gone, in cell nbut in the autophagosome) -Lysosome has proteolytic enzymes so it fuses with autophagosome to make a autolysosome or autophagolysosome. now things are being broken inside (enzymes only degrade the things inside) -then they are spit out to be used in metabolism or biomaking of other things -cell recycling system PHAGOPORE: start of an autophagosome PATHWAY Important Enzymes: a) ATG7: like E1, critical for autophagosome formation; when it is turned on, it drives this process b)LC3: ubiquitin like protein that forms the autophagosome membrane c) p62: adaptor protein that bings to ubiquitin and LC3 d) LAMP2: aids in autophagosome and lysosome fusion Two kinases that Initiation of Autophagosome formation and activation/induction: two main complexes drive this a) Beclin 1 complex: macromolecular complex that once activated promotes autophagosome membrane generation -negatively regulated by BCl2, BCLx1 -when apoptotic stress is high, beclin 1 will activate autophagosome formation to help survival, they wil look for stress signals ; if it cant solve it aoptotic signals will continue and this will kill it b) mTOR substrate Complex: -highly regulated by mtor and ULK, once acrivated promotes autphag membrane gen -very sensitive to changes in energy status, and substrate status / availability -ampk is activatedin low atp levels. increased ADP levels. this drives formation of autophagosomes (if cell needs atp, it is going to break things down to get it to fuel the cell) Starvation and execise are energy stressors that activate this MTOR pathway Completion and Fusion of Autophagosome: 1) phsopholipid double membrane forms but there is a protein that lines the inside of that membrane called LC3II resides 2) forms around various parts of the cell 3)LAMP2 conjugates the membrane of lysoszome and autoghag membrane so they are fused 4) allows them to be together, outer membrane is intact but inner membrane goes away and things get spit out

Question 84

Non selective and Selective targeting of proteins and organelles to the autophagosomes

Answer 84

Non Selective: as the auto are formed, things are found and they are degraded. Whatever is there gets degraded Selective: Recognizes proteins with an identification called P62 -remember LC3 is on innner membrane, and proteins that are broken can be ubiquinated and p62 can bind to both ubiquitin and lc3, so binds these structures to the autophagosome Damaged mito: p62 accumulates on outer membran, it starts to degrade, and same with bacteria

Question 85

Regulation of Autophagy is linked with Cell stress responses and Cell death

Answer 85

Mitochondria: released proapoptotic factors and ROS-> leads to dna fragmenetation and damage Protein aggregates: leads to ER stress these signals influence cell death and other things autophagy there to mitigate cell death, sometimes too well IE in some cancers it mitigates death signals that are meant to get rid of the cancer, so cells keep living Or autophage low: so it accumulates

Question 85

Mitophagy

Answer 85

Targeting of mitochondria for Autophagy -damage and /or loss of mitochondria membrane potential can initiate the targetting of mitochondria fro degredation of autophagy Change in membrane potential in mito is a signal : hydrogen gradient drives this, high membrane potential is inner membrane compared to matrix, means better at making ATP, pores formed here dropped membrane potential During low membrane potential PINK1: found on outer membrane in accumulation Parkin: e3 ligase, normally found in cytosol but moves to mitochondria whe loss of membrane potential (ie pores there) -uppromotes ubiquitanition of mitochondrial proteins such as VDAC -autophagosome forms around this because P62 binds to ubiquitin and LC3 so this can now form a thing around it FOR DAMAGED MITO: BNIP3/NIX : BNIP3 and NIX are autophagy specific receptors that are expressed in DAmAGED proteins, they can interact with LC3 directly (do not need ubiquitination) -these proteins are part of BCl2 family, accumulate in stress for damaged mito

Question 86

Autophagy is required for muscle maintence

Answer 86

ATG7: if they dont make ATG7 they wont make autophagosomes, need it for autophag formation -should have reduced activation of autophagy -LC32 is needed for autophagosome, 2 is the specific one that is activated by atg7 -p62 is degraded when system works well So no atg7 means no lc32 (accumulation of LC3), and accumulation of p62, so accumulation fo structures that have p62 that are accumulating -centralized nuclei occur when ATG7 KO happens, muscle wastes away: so increased apoptosis instead of autophagy so more smaller so FORCE DECREASES IN ATG7 KO mice We also see increase in MURF and MAFBX (e3 ligase of UPS) because it is trying to do UPS system but these wont fit there so why would it be increasing? because it is trying to compensate there is high ubiqutin tagged things so it is trying upregulate other processes

Question 87

Basal and Starvation Induced Autophagy is dependant on FT

Answer 87

different levels of LC3 and p62 in different muscle types Soleus (Slow) : Fast: no p62 Starvation induced: autophagosomes go up, more dots in fast muscle (more sensitive starvation induced autphacgy) WHY FAST MORE THAN SLOW: we want to keep type 1 fibers around because we use it more often... or maybe because the fast is relying on carbohydrates, but soleus can survive on fat so if it relies on carbs it will see more energy stress. (so type 2 more affected)

Question 88

Autophagy fkux is higher in fast muscle: wasting instarvation state

Answer 88

Fast Glycolytic : have lower mito, auto proteins are less but activation in times of starvation is HIGHER (flux is faster) -mitophagy flux is faster in this which is why their mito content is low becausse they degrade it faster Energy Stress: changes in ATP -AMPK energy sensitive enzyme that senses ADP and AMP, this gets activates and it drives autophagy (MtOR pathway) -and signalls for mitochdonrial biogenesis -does both bruh (breaks down di=ysfunctional, makes new)

Question 89

Autphoagy is required for mito maintence in aging

Answer 89

ATG7 KO: accumulate p62 Reduction in auphoagy leads to accumulation of abnormal mito, swollen lacking cristae , dysmorphic -Aged animals that have reduced autophagy have impaired mitochondria oxygen consumption, without altered ETC complex expression (less o2 consumptio mean atp is reduced) OLD ADULTS: accumulation of damaged and dysfucntional mitochondria, fewer mito, more dysgunctional ones

Question 90

Autophagy and msucular dystrophy

Answer 90

-downregulated autophagy -accumulation of abnormal mito and sr -lower lc3 ( so low autophagy) -INCREASED APOPTOSIS -starvation removed damaged organella nd reduces apoptosis WHY because we are inducing autoophagy ( we are forcuing muscle to find fuel to use as fuel source) *********** Starvation protection in MD: -upregulation of autophagy (increased lc3 and beclin), and increased mitophage (BNIP3) -KO of BECLIN and BNIP3 eliminate protective effects of starvation (Apoptotic nuclei goes back up)... starvation does work (BNIP IS FOR MITOPHAGY, BECLIN FOR AUTOPHAGOSOME INDUCER)

Question 91

Exercise increase AMPK and ULK activation

Answer 91

Exercise activates AMPK leads to ulk increase ULK (part of mTOR pathway), increased with exercise causes acute energy stress that increases autophagy to use as a fuel source so biogenesis and mitophagy is induced mTDNA: exercise lowers mtdna muations, and exercise lowers atrophy of muscle