SM01 Mini2 Flashcards
Mitochondria
long, ovoid membrane bound organelle found in the cytoplasm
has own DNA
responsible for converstion of food to usable ATP energy
avg. 1000/cell, erythrocytes= 0, more energy demand→ more mitochondria
& distribution vary according to cell type
under basal bodies in ciliated cells to provide ATP for dynein activity in ciliary beating
new ones are made via division, not linked to cell cycle & they do not all divide at the same time
sperm mitochondira are tagged with ubiquitin
Outer Mitochondrial Membrane
contains porin, large channel forming protein, that are ALWAYS open (only pore in the human body)
thus permeable to molecule 5000Da or less
Inner Mitochondrial Membrane
surrounds matrix, infoldings create cristae→ increased surface area
contains proteins that carry out oxidative rxns of electron transport chain & ATP synthase
Intermembrane Space
contains enzymes that use ATP passing out of matrix to phosphorylate other nucleotides
resembles cytosol
pH=7
Mitochondrial Matrix
inside inner membrane
highly concentrated mix of enzymes for oxidation of pyruvate & fatty acids & those for citric acid cycle
pH=7.5
Cristae
finger-like projections that cross the mitochondrion formed by the infoldings of the inner membrane
contain ATP transporters that pump new ATP from matrix to intermembrane space
mDNA
mitochondrial DNA
2-10 circular copies/mitochondrion
<1% of cellular DNA
only 13 out of 615 proteins of mitochondria are coded for on this DNA
oxidative phosphorylation
Lysosome
body where lysis occurs
membrane-bound organelle containing digestive enzymes, typically most active at acidic pH (4.8- proton pumps to acidify lumen)
only in cytoplasm, NOT nucleoplasm
degrades proteins, lipids, carbohydrates, DNA, RNA,
size, #/cell, & appearance vary greatly per need
ALWAYS smaller than nucleus in normal cell
material to be degraded is brought in by vesicles that fuse w/lysosome
M6P Signal
mannose 6 phosphate
signal on proteins to be packaged together to form a primary lysosome
phosphotransferase adds M6P to proteins with lysosomal amino acid sequence with N-linked sugar
primary lysosome
new lysosome that has just budded from the trans Golgi
contains newly synthesized enzymes
before it receives any material to be digested
“virgin lysosome”
sometimes exicytosed to degrade subtances in the ECF
secondary lysosome
primary lysosome after it has fused with vesicles containing material to be degraded
lysosomal storage diseases
>30
most linked to mutation in specifc acid hydrolase
leads to accumulation of partially degraded insoluble metabolite in lysosome
ex. I cell disease & Tay Sachs disease
Tay Sachs Disease
absence of hexosaminidase A→ cannot breakdown glycolipids (highly prevalent in neurons)→ neurons ballooned w/cytoplasmic vacuoles
destruction of neurons
symptoms: 6 months relentless motor & mental deterioration, and early childhood death (2-3 yrs)
more common in Ashkenazi jews
exception of one time lysosomes are bigger than nucleus
autophagy
catabolic process involving degradation of cell’s own components via lysosomal machinary
purpose: provide raw materials to sustain life, seen in starvation
endocytosis
uptake of material into cell by invagination of plasma membrane & internalization of membrane-bound vesicle
function: bring molecules from ECF inside cell &/or retrieve plasma membrane proteins
phagocytosis
endocytosis in which vesicle contains large food particle
ONLY macrophages & neutrofils
proteosome
degrades unneeded or damaged proteins by proteolysis that have been tagged with ubiquitin
found in cytoplasm & nucleoplasm
Peroxisome
small membrane-bound organelle that uses molecular oxygen to oxidze organic molecules
contains enzymes that produce organic molecules, produce hydrgen peroxide & degrade hydrogen peroxide
NOT found in every cell in body
important for liver & kidney function to detoxify bloodstream
rapid responce to change (proliferation when needed)
EM: dark due to stain rxn with catalase enzyme, but otherwise can’t be distinguished
fucntions of peroxisome
- rid body of toxic substances: hydrogen peroxide, phenols, formic acid, formaldehyde, alcohol
- 1/4-1/2 of ingested alcohol is broken down in perioxisomes
RH2 + O2 → R + H2O2
then catalase used H2O2 to oxidize other hydrocarbons: H2O2 + R’H2 → R’ + 2H2O
- breakdown of long chain fatty acids (>22C) via beta-oxidation→ acetyl-CoA
- NOT coupled with ATP production, but creates H2O2 instead
- synthesis of bile acids in liver
- synthesis of plasmalogens to make myelin→ thus contribution to neurologic symptoms
peroxisome formation
- de novo: from ER & proteins are imported (-ser-lys-leu-COO-)
- fission: an existing one divids into two
aerobic respiration
uses oxygen
oxidative phosphorylation takes place in mitochondria
approx. 30 ATP produced
anaerobic respiration
doesn’t use oxygen
takes place in cytoplasm by glycolysis
makes 4 ATP
Mitochondrial fission
one mitochondrion splits into two
don’t understand why yet
Mitochondrial fusion
two mitochondria fuse into one
don’t understand why yet
Functions of Mitochondria
- acetyl-CoA in oxidative phosphorylation in ATP production
- breakdown of fatty acid molecules to acetyl-CoA
- must be 22C or less
ATP synthase
makes ATP in mitochondrial matrix coupled with protons moving down their electrochemical gradient (intermembrane space to matrix of mitochondria)
found as transmembrane protein in inner membrane of mitochondria
mitochondrial targeting sequence
most mitochondrial proteins are still encoded by nuclear genome
always at amino terminus
binds to mitochondrial chaperones before binding to keep it exposed & target it to the mitochondria after ribosome relase
every fourth aa has a positive charge (this is the recognition site for the chaperones
mitochondrial disease
- defective gene in nuclear genome
- defective gene in mDNA (thus maternally inherited - 15% of total)
How Mitochondria Make ATP
- oxidation of fatty acids & pyruvate to acetyl-CoA, coupled w/reduction of NAD+ & FAD→ NADH & FADH2
- electrons from reduced coenzymes are transferred via 3 electron transport complexes to O2 & H+ are transported frommatrix to intermembrane space
- matrix becomes basic (low [proton]) & negatively charged (electric potential)
- ATP synthase makes ATP coupled with protons moving down their electrochemical gradient (intermembrane space to matrix of mitochondria)
proton-motive force
low proton concentration in matrix + negative charge of matrix relative to intermembrane space
cardiolipin
double phospholipid (four fatty acid tails)
found mainly in mitochondrial inner membrane
catalyzed by mitochondria themselves
packs tighter than regular phospholipids, better to withstand stress
Ubiquination
attaching of ubiquitin to a protein to signal proteosome for degradation
signaled by exposure of string of hydrophobic aa that are normally buried
Special Properties of Lysosomes
- ATP driven proton pump for acidification
- glycoprotein coat on inner surface to protect it against hydrolysis by its own enzymes
- transport channels to transport products out of lysosome: amino acids, nucleotides, glucose, etc.
Opsinized
“seasoned”
addition of eat me ligand signals attached something to be phagocytosed
ex. antibody coating of bacterium, Fc is eat me signal
Autophagosomes
organelle wrapped in ER membrane
destined to fused with lysosome
Autolysosome
autosome that has fused with lysosome
residual body
remains of secondary lysosome
can be exocytosed or turn into lipofuscin granules (if pigmented)
M6P receptor
affinity for M6P at pH=6.5, but not low pH of lysosomes
drops cargo off in lysosomes & recycles
I cell disease
defective phosphotransferase→ cannot make M6P targeting signal→ all lysosomal hydrolases are secreted
waste buildup creates “I” (inclusion) cells
symptoms: 6 months failure to thrive & developmental delay, and early childhood death
PTS
peroxisomal targeting signal
ser-lys-leu at COOH terminus of protein
Zellweger’s Syndrome
defect importing proteins to peroxisomes
die soon after birth
homozygous recessive of PTS receptor mutation
sER Structure
continuous with rough ER
distal to nucleus
held in place by mictrotubules
more if cell needs to make a lot of lipid or hormones
sER Functions
- lipid biosynthesis
- all except for cardolipin (made in mitochondria)
- detoxification rxns
- more complex molecules than those in peroxisomes, ex. phenobarbital
- both do ingested alcohol
- cytochrome p450s (CYP)
- more complex molecules than those in peroxisomes, ex. phenobarbital
- regulation of Ca2+
- especially in muscle contraction (sarcoplasmic reticulum)
- cellular signaling pathways
rER Structure
grows out of nuclear envelop
continuous with sER
contains bound ribosomes (they are signaled to attach, but are the same as free ribosomes) on outside of membrane
more of it if cell secretes a lot of protein
held in place by microtubules
rER Functions
- protein synthesis
- secreted proteins
- transmembrane proteins (except mitochondrial)
- lysosomal proteins
- protein modification
- sequestration of Ca2+
- released in signaling pathways
Flippases
moves phospholipids from extracellular leaflet to cytoplasmic leaflet
uses ATP
in Golgi
flips phosphatidylserine (negative charge attracts peripheral membrane proteins, ex. PKC) & phosphatidylethanolanine from lumenal (extracellular) to cytosolic face
Scramblases
does NOT use ATP
moves phospholipids form one leaflet of a membrane to the other in both directions
movement of two lipid in opposite direction
only found in sER
exist because all new lipids are added to cytosolic leaflet
non-specific to polar head group
makes sER membrane homogenous mixture
Lipid transport mechanisms
- lateral diffusion within a bilayer
- scramblase assisted translocation between leaflets
- lipid-transfer protein assisted movement thru cytosol
- non-specific, bump around from membrane to membrane
- incorporation in membrane-bound vesicles
Secretory Pathway
ER signal sequence
usually at amino terminus of protein
usually cleaved in rER lumen by signal peptidase
8 or more non0polar aa at the center
once translated, will be bound by SRP
SRP
signal recognition particle
binds to ER signal sequence→ stops mRNA translation→ binds to SRP receptor on rER→ ribosome binds ribosomal receptor on rER & translocator complex→ mRNA translation resumes→ protein pushed through translocon into rER lumen
Lipid biosynthesis in sER
newly synthesized lipids inserted into outer leaflet of sER bilayer
made on cytosolic side of sER
Floppases
moves phospholipids from cytoplasmic leaflet to extracellular leaflet
uses ATP
Chaperones
2 types: hsp70 (heat shock protein 70 in cytoplasm & BiP in ER lumen) & GroEL family
hsp family binds to hydrophobic domains in unfolded proteins
prevent aggregation of unfolded proteins & aid in proper folding
Protein membrane insertion
- some use ER signal as transmembrane domain
- translocon springs open to laterally release hydrophobic sequence into membrane
- positively charged aa at either end are flipped so charge faces cytoplasm
*
protein modifications in rER
- N-terminal singal peptide cleavage by signal pepidtase
- N-linked glycosylation (on asparagine)
- core= 2x N-acetylglucosamine & v branch of 3 mannose
- other mannose & glucose added
- only core survives trimming in Golgi for many
- formation of disulfide bonds via oxidatioin of cysteine sulfhydral groups
- stabilizes protein conformation
- cannot occur in cytoplasm due to reducting atmosphere of glutathione
- ex. light & heavy chains of antibodies & alpha & beta subunits of insulin receptor
- transmembrane domain/+GPI anchor (lipid)
1. covalent bond- still integral protein
- transmembrane domain/+GPI anchor (lipid)
- protein folding by chaperones
1.
BIP
binding protein
homologous to HSP70
in rER
correct folding required to leave rER or will tag for degradation
binds to hydrophobic patches as they come through the translocon
N-glycanase
enzyme that removes N-linked glycosylation for protein ubiquination & degradation
Cystic Fibrosis
most common fatal genetic disease in US
death caused by repeated chest infection
mutation in cystic fibrosis transmembrane conductance regulator (CFTR) gene
chloirde ion transporter of epithelial cells misfolds→ transporter absent from plasma membrane
chloride imbalance causes cells to secrete less water, cells swell, secretions are very thick
secretory pathway
ER (rough or smooth)→Golgi→ secretory vesicle→ plasma membrane
regulatory signal required if bound for elsewhere (lysosome or regulated secretion)
SNAREs
(Soluble NSF Attachment protein Receptor)
v-snares on vesicles; t-snares on target membranes
used for targeting & drive membrane fusion reaction
35 different SNARES, each associated with particular membrane enclosed organelle
snap membranes together so strongly it drives water out→ stalk formation→ hemifusion cytosolic leaflet fused but not lumenal leaflet)→ fusion
Rabs
small GTP binding proteins
found on vesicle membranes (different types for direction)
contribute to specificity of docking
binds to tetherin protein on target membrane→ brings SNARES in closer proximity
NSF
(N-ethylmaleimide-sensitiven factor)
solubel protein responsible for breaking apart v- & t-SNAREs for recycling with help of SNAPs, uses ATP
SNAPs
acessory proteins that aide NSF in recycling of v- &t-SNAREs
Botox
uses various forms of botulinum toxin to paralyze muscle activity
cleaves SNAREs for exocytosis of regulated secretory vesicles at neuromuscular junction→ no neurotransmitter release→ no muscle contraction
Golgi
membraneous complex of vesicles, vacuoles, & flattened sacs in the cytoplasm
involved in portien modification, intracellular secretion & transport
located on one side of nucleus on top of centrosome (mictrotubule organizing center)
cis (closest to ER), medial (mulitple sacs), trans (faces plasma membrane-exit)
modifications in Golgi
- trimming of N-linked carbohydrates
- addition of sialic acid to glycoproteins & glyolipids
- aka NANA
- turned black w/ Golgi stain
- onlly carbohydrate group w/ - charge
- give extracellular leaflet of plasma membrane negative charge
- addition of O-linked sugars to serine & threonines
- glycosylation of some lipids, ex. ceramide
glycocalyx
cell coat created by thick rim of carbohydrates fanning out from plasma membrane
functions: protection, cellular recognition, slows rate of degradation of secreted & membrane proteins
regulated secretion
signal mediated secretion
directed to lysosome or secretory vesicles
ex. insulin, neurotransmitters (acetylcholine, glutamine)
rise of intracellular Ca2+ often triggers release
constitutive secretion
secretion without signal mediation
operates continuously
M6P signal mechanism
phagocytosis mechanism
types of molecules: 0.1-10micrometers in size
pintocytosis mechanism
“cell drinking”
each budding vesicle traps a drop of extracellular fluid as it pinches off
types of molecules: indiscriminate
receptor-mediated endocytosis
100-500nm
mediated by clathrin coat proteins
receptors for specific proteins cluster in clathrin pits, can have many different receptors in same pit
viruses like to exploit (ex. flu)
types of molecules: insulin, EGF (epidermal growth factor), transferrin, LDL (low density lipoprotein), & polymeric IgA
possible fates: recycling, transcytosis, degradation
LDL endocytosis pathway
receptor recycles, ligand degrades
receptor needed for multiple round of endocytosis
ligand is degraded for the cell to use the cholesterol
LDL= low-density lipoprotein, carries cholesterol made in liver through the blood to the body (75% from liver/25% from food)
10-15 min process
proton pump of endosome acidifies vesicle after pinching off→ 6.5 receptor unbinds LDL & pinches off vesicle→ 4.5 merge with lysosome for LDL degradation
transferrin endocytosis pathway
ligand & receptor recycle
transferrin blinds to Fe in bloodstream
apotransferrin= no bound Fe
diferric-transferrin= bound to Fe (2Fe3+)
7.2 transferrin receptor has low affinity for apotransferrin, but high for diferric-transferrin→ 6.5 of early endosome Fe release from transferrin & leaves endosome→ transferrin & transferrin receptor are recycled
EGF endocytosis pathway
ligand & receptor are degradated by lysosome→ mechanism for down-regulation of signaling pathway
EGFR only cluster in clathrin pits when they are bound to ligand
IgA secretion
ligand & receptor are translocated acorss cell & released on the other side
IgA antibodies coming from bloodstream→ bind to receptor on one side of a polarized cell→ travel with receptor in vesicle to other side of cell→ released to ECF on opposite side
caveolae
“little caves”
small invaginations of plasma membrane, specialized lipid rafts
many cell types, but neurons have none
abundant in endothelial blood vessel cells (most transcytosing vessels in this cell type)
caveolin: protein that causes invagination
proteins found in them: GPI-linked & proteins w/longer than average transmembrane domains (signaling pathways)
functions: signal transduction & caveolar endocytosis
secretory vesicles
regulated ones are often transported intracellularly by dyenin & kinesin on MT to plasma membrane→ actin & myosin take over→ vesicles enmeshed in actin waiting for trigger to release (often rise of intracellular Ca2+)
usually darker on EM due to aggregation of contents
Coat proteins
clathrin, COPI, COPII, (caveolin possible 4th)
vesiculation requires coat proteins
water soluble
assemble on membrane face & induce curvature→ serve to cluster membrane cargo proteins→more added to shape membrane into sphere→vesicle pinches off & coat falls off
Clathrin
protein coat portein for receptor-mediated endocytosis of plasma membrane
triskelion= unassembled state
lattice= assembled on membrane
COPI
protein coat portein for receptor-mediated endocytosis of Golgi
COPII
protein coat portein for receptor-mediated endocytosis of ER
mitosis
eukaryotic cell division
stages: prophase, metaphase, anaphase, telophase
prophase
second stage of mitosis
chromatin condenses to chromosomes, nuclear envelop breaks down, & initiation of mitotic spindle but centrosomes
4X chromosomes, 2n DNA
metaphase
third stage of mitosis
chromosomes line up on equatorial plate, spindles bind to kinetichores, centrosomes are at opposite poles
4X chromosomes, 2n DNA
anaphase
fourth stage of mitosis
sister chromatids are pulled apart to opposite poles & initiation of cleavage furrow
4X chromosomes, 2n DNA
telophase
fifth (last) stage of mitosis
chromosomes unravel to chromatin, nuclear envelop forms, & cytokinesis forming two daughter cells both exactly like the parent cell
2X chromosomes, n DNA
actin function in mitosis
formation of contractile ring during cytokensis
microtubule function in mitosis
form spindle fibers that bind to kinetochore of chromosomes to pull the sister chromatids apart
minus-end directed motor protein is part of kinetochore protein complex
some motor anchor MT to plasma membrane & pull
other motors push overlapping MT to push the poles apart
intermediate filaments function in mitosis
break down nuclear envelope in prophase triggered by phosrylation of nuclear lamins
create two new nuclear envelopes during telophase
cohesins
proteins that cross-link two adjacent sister chromatids, multiple along the length of chromosome
critical for chromosome segregation
degraded at start of anaphase
condensins
proteins that mediate intramolecular cross-linking to coil DNA during chromosome condensation
Taxol
anti-microtubule drug used for cancer treatment
arrest mitotic cells because spindle fibers cannot form so that they perform apoptosis
G0
quiescent phase
inactive
neurons stay in this phase permenantly
G1
phase most variable in length, dependent on tissue type (bone= 25h)
the differentiated the longer it will stay in this phase
S phase
synthesis phase
DNA is replicated, 2n DNA, 4X chromosomes (but they are not condensed yet)
in bone= 8hrs
G2
growth and preparartion for mitosis
in bone G2 + mitosis= 2.5-3hrs
Preprophase
first stage of mitosis
intranuclear condensation of chromatin & centriole duplication to two centrosomes
organelles during mitosis
ER: vesiculates (breaks down) when the nuclear envelope does, reforms during telophase
Golgi: vesiculates (breaks down) when the nuclear envelope does, reforms during telophase
mitochondria, lysosome, & perxoisomes: nothing, but stay out of spindle region (unexplained as of yet)
necrosis
premature, accidental death
cells swell & break open, releasing their contents
effects on organism: can damage surrounding tissue & possibly damaging inflammtory response
apoptosis
programmed cell death
appearance: round up, appear bigger but are NOT actually bigger
intracellular changes: fragmentation of DNA, shrinkage of cytoplasm, membrane changes→ pieecs bleb off & are phagocytosed by macrophages
effects on organism: no lysis/no inflammation→ no damage to surrounding cells, imperceptible to organism
causes of necrosis
mechanical trauma, eposure to toxic agent, burning, freezing, intense UV radiation, anything that quickly depletes ATP of cell (ie hypoxia→ ischemic stroke & heart attack)
mitochondrial role in apoptosis
pro-apoptotic BCL-2 family member forms channel in outer mitochondrial membrane releasing cytochrome c (part of the apoptosome) & apoptosis inducing factor
anti-apoptotic BCL-2 family member can bind to these to inhibit channel formation
caspases
protein family of proteases that play a role in necrosis, apoptosis & inflammation
cysteine proteases that cleave just C-terminal to asp residues
synthesized as inactive pro-enzymes
cells die in few hours-a day after receiving negative signal or withdrawal of positive signal
BCL2
family of proteins that regulate when apoptosis occurs, some pro & some anti
they can regulate each other by forming heterodimers
caspase cascade
pro-apoptotic signal→ activation o f initiator caspases→ cleaves & activates effector caspases→ break down of cellular targets
1 molecule of activated caspase can amplify & kill the cell
cell survival requirements
produce ATP
be able to maintain barrier to external environment
Effects of increased [Ca2+]cytoplasmic
major cause of cellular injury
denatures protein
poisons mitochondria
inhibits cellular enzymes
inflammation
protective attempt by organism to remove the injurious stimuli & initiate healing process
characterized by redness, pain, heat, & swelling
caused by: increased blood flow & leakiness of capillaries→ bringing white blood cells to affected tissue
hypoxia
oxygen deficiency that causes cell injury & death by reducing oxidative respiration in mitochondria
ex. ischemic stroke & heart attack
causes: ischemia (reduced blood flow), inadequate oxygenation of blood due to cardiorespiratory failure, decrease oxygen carrying capacity (anemia, CO poisoning, severe blood loss)
calpains
calcium-activated neutral proteases
activated in brain by high calcium (which can occur during hypoxia b/c neuron can’t make enough ATP to maintain strong ion gradient)
cause a lot of damage in brain trauma
functions of apoptosis during development
- deleting unwanted structures (tadpole tail)
- sculpting specific tissues by ablating fields of cells (developing digits)
- controlling cell # (50% of neurons eliminated during maturation)
- eliminating cells during development that are abnormal, nonfunctional, or potentially dangerous (T & B lymphocytes that recognize self)
functions of apoptosis during adulthood
- maintaining homeostatis- cell #
- eliminating damaged, mutated, or infected cells
- withdrawal of growth factors
- viral infection (hopefully before virus can infect surrounding cells)
cell loss disorders
AIDS, Alzheimer’s, Parkinson’s, aplastic anemia, myocardial infarction