Week 5 Random Flashcards
Two types of on/off swithces
Protein kinease / Protein phosphatase (signalling by phosphorylation) = covalent
GTP binding (GEF) / GTP hydrolysis (GAP) (signaling by GTP-binding) = noncovalent
Why molecular swiches are necessary?
Allow integradation of signal at signal processor
Cdk Kinase requirements for activation
Has three conditions in order for Cdk kinease to take signal downstream (2 phsophates and cyclin)
Where can the intracellular signalling complex assemble?
Type and precisioon of response to signalling molecules
Steroid are usually gradual response
Cooperativity might stimulate quicker response (all or nothing like)
Who positive feedback can affect response?
Can accelearate response
How desintization can occur?
Recepotor sequestration
Receptor down-regulation
Receptor inactivation
Inactivation of signalling protein
Productionof inhibitory proteins
How penile erection occur?
Intracellular receptor
Neuron releases ACh -> Activates NO Synthase ->
arginine converted to NO (endothelial) ->
NO diffusion goes to smooth muscle cell activates to gyanylyl cyclase ->
GTP is converted to cGMP and relaxes smooth muscle
Activation of PKG (phosphorylation) to vascular smooth muscle relaxationand blood vessel dilation
PDE inibitors type V
Levitra
Cialis
Viagra
PDE type 5 action
Prevent cGMP conversionto GMP
Some examples of molecules that bind intracellular recepotrs
Cortisol
Estradiol
Testosterone
Thyroxine
Vitamin D3
Retinoic Acid
Machanism of hormone receptor activating transcription
Two types of cellular response to signals
Altered protein function by intracellular singalling pathway (fast sec-min)
Altered gene expression (mins to ours)
Cell membrane receptors
GPCR’s = Ga, Gi, Golf, Gt, Gq
Ras = MAP kinease
Enzyme-linked receptors = PI3 kinease, PLC-g, IP3, Ca2+, SRc, Jak-STATs, NF-kB
(-mab) in the name of the drug
Humainzied monoclone antibody
Drugs inactivating NFkB signalling
Remicaid, Humira, Cimzia, Enbrel, Simponi
Communication in cells is necessary for:
- Regulate development and organization of tissues
- Control their growth and division
- Coordinationof their functionwith each other
Order the signalling pathways from shortest distance to longest
Synaptic
Paracrine
Autocrine
Endocrine
How is the effector different in endocrine vs. synaptic signalling?
In endocrine signalling, the receptor sees mix of signals while in synaptic signalling, the receptor only sees the specific or limited signals.
Why autocrine signalling evolved?
The strength of the signal might be beneficial
Autocrine signalling is important for development and during immune system development
Importance of eicosanoids
Inhibitors of eicosanoids synthesis
Names of enzymes that are involved oxidation
What prostaglanding mediate
Eicosanoids (signaling molecules made by oxidation of 20-carbon fatty acids)
Inhibitors of eicosanoids include cortisone and Nonsteroidal anti-inflammatory drugs (NSAIDs) like ibuprofen
COX1 and COX2 are enzymes incyclooxygenase dependent pathway
Prostaglandin regulates inflamatory response
Problems with COX-2 inibitors?
Multiple sides efects: CELEBREX, and Vioxx (withdrawn)
Endocrine vs. Synaptic signalling affinity / length
Endocrine: Low ligand conc
Synaptic: High lingad conc >10^-4 M; low affinity; quick termination
Prozac
Inhibits seritonin uptake that allows maintenance of seritonin concentration in synapse
Anti-depressant
Selective serotonin reuptake inhibitors (SSRIs)
They act within the brain to increase the amount of serotonin
For anxiety disorders, obsessive-compulsive disorder, and eating disorders.
Effective in treating premature ejaculation in up to 60% of men.
How cells can respond to signals
Survive
Grow + Divide
Differentiate
Die
How same cell can respond to different signals?
Ach signal
1) Skeletal muscle = contraction
2) Heart muscle = relaxation (different receptors)
3) Salivary gland = secretion (same receptor as heart, but different protome)
Extracellular matrix
Components and types
Sugars and Proteins
Interstitial (surrounds in tissues and abundant in connective tissues)
Basement membrane (sheet of fibers that underlie the epithelium)
Functions:
Collagen
Fibronectin
Laminin
Elastin
Glycosaminoglycans
Proteoglycans
Collagen – ropelike fibers that give tissue tensile strength.
Fibronectin and Laminin – glycoproteins that link the extracellular fibers to the cells.
Elastin – thin fibers that give tissue elasticity.
Glycosaminoglycans – extracellular polysaccharides of defined sequence.
Proteoglycans – gel like slimy mucus that hydrates the space between cells.
Collagen Synthesis
Prepocollagen made in ER
(1) Signal sequence is removed -> polyproline helix
(2) Intra chain dilsufide bonds form with a-chains
(3) ER: Various prolines and lysines are hydroxylated by prolyl or lysyl hydroxylases (Vit C)
(4) ER: Collagen is O-glycosylated on hydroxylysines and N-glycosylated on asparagines with galactose or galactosyl-glucose.
(5) ER: The 3 α-chains assemble into a soluble right handed super helix.
(6) Procollagen is secreted
(7) Extracellular proteases cleave the ends generating collagen molecules (tropocollagen).
(8) Fibrils self-assemble into insoluble fiber complexes in a quarter staggered array.
(9) Fibrils are covalently cross-linked
Lysyl oxidase – oxidative deamination of lysine and/or hydroxylysine (Cu2+ dependent ). Followed by an aldol condensation.
More crosslinking = more rigid
Collagen
where is it found?
of types?
examples?
Bones, tendons, and skin
Most common protein in mammals (25-30%)
28 types
examples:
Collagen I, II, and III (90%)
Collagen IV - basal lamina
Non-collagen collagens - C1q, pulmonary surfactant proteins (SPD, SPA)
Pyrimidine synthesis
Source of atoms in pyrimidine synthesis
Name enzymes
ATP + AMP -> 2ADP
ATP + NMP -> ADP + NDP
ATP + NDP -> ADP + NTP
UTP -> CTP
Adenylate kinease
Nucleoside monophosphate kineases
Nucleoside duphosphate kineases
Cytidylate Synthetase
Salvage pathway
Names of enzymes in salvage pathway that form XMP
Defect in one of them
Pyrimidines: orotic acid transferase (existing)
Adenine: adenine phospho ribosyl transferase (A-PRT)
Guanine/Hypoxanthine: G/HX phospho ribosyl transferase (G/HX-PRT)
Lesch-Nyhan syndrome: defect in G/HX-PRT
Source of atoms in purine synthesis
How AMP and GMP are formed from IMP?
Regulation by ATP / GTP levels
How deoxyribonucleotides are synthesized from ribonucleotides?
How the reaction is driven?
By NADH
GMP and AMP syntehsis regulation
General regulation of nucleotide synthesis
Thymidine Synthesis
Cystonie and Uracil degradation
Thymidine degradation
GMP and AMP degradation
Gout
Cause?
Treatments?
The average uric acid concentrations in humans is near the solubility limit. This has a selective advantage because uric acid is a highly effective scavenger of reactive oxygen species. However, if at low pH uric acid crystals form, they can irritate joints and cause gout.
Treatments:
Colchicine: anti-inflammatory
Probenecid: Increases uric acid excretion
Allopurinol: Xanthine oxidase inhibitor
Cobalamine absorption
Reactions that B12 catalyzes
Lack of B12?
B12 (required by methylmalonyl CoA mutase and methionine synthetase)
Megaloblastic anemia. Neurological dysfunction. Deficiency of folate.
Pernicious anemia: autoimmune disease destroys parietal cells.
Treat with B12 supplements or monthly injections.
Folate Importance
B9 vs. B12 deficiency
Folate deficiency
Deficiency? (1) Folate deficiency decreases purine and dTMP synthesis, arresting cell cycle in the S-phase and resulting in megaloblastic anemia. (2) Hyperhomocysteinemia with increased risk for cardiovascular disease. (3) Deficiency in pregnancy can lead to neural tube defects (spina bifida) in baby.
Why needed? THF; one carbon carrier involved in amino acid metabolism and nucleotide synthesis
Source: Yeast, liver, fruits, green vegetables
ECM functions
Connecting cells together
Guides cell migration (e.g. would healing)
Relay of environmental signals
Collagen I
Charactersitics
Left handed tripple helix of three
Repetitive AA sequence: (Gly-X-Y)n
33% Glycine and about 20.5% Proline/Hydroxyproline at X and Y respectively.
3 α-chains assemble into a right handed super helix.
Mechanism of collagen quaternary structure assembly
Lysyl oxidase – oxidative deamination of lysine and/or hydroxylysine (Cu2+ dependent ).
Followed by an aldol condensation
Diseases associated with collagen mutations
Basal Membrane vs. Basal Lamina
The epithelial ECM the term “basement membrane” is used with light microscopic observation and “basal lamina” is used with electron microscopy.
Basement Membrane = Basal Lamina + Retircular Lamina
Basal lamina characteristics
(about 40–120 nm thick) consists of fine protein filaments embedded in an amorphous matrix.
Membrane proteins of the epithelial cells are anchored in the basal lamina, which is also produced by the epithelial cells.
major component of the basal lamina are two glycoproteins - laminin and (usually type IV) collagen
Reticular lamina charactersitics
consists of reticular fibers embedded in ground substance.
fibers of the reticular lamina connect the basal lamina with the underlying connective tissue.
components of the reticular lamina are synthesized by cells of the connective tissue underlying the epithelium.
Basement membrane compnents
- Collagen IV
- Laminin
- Heparin Sulfate
- Proteoglycans
Laminin (Ln) characteristics
A glycosylated cross-shaped heterotrimer.
A multi-adhesive ECM component enriched at the basal lamina where it binds cells to collagen IV and integrins.
Multiple isoforms.
In general, Ln is associated with cell differentiation.
Laminin vs Lamin A vs Lamina
Laminin 2 – basement membrane protein that links integrin (or dystroglycan) to ECM components.
Lamin A is a nuclear envelope protein that forms filaments.
Lamina is the ECM component of basement membranes seen by electron microscopy (basal lamina).
Elastin + Fibrillin characteristics
Elastic fibers that allow tissue to expand and contract.
Abundant in blood vessels, lung and skin.
Highly cross-linked, insoluble, amorphous structure rich in proline (11%), alanine (22%) and glycine (31%).
Like collagen, lysyl oxidase initiates crosslinking of allysines.
The crosslinks formed are called a desmosine.
Defects in elastin cause Williams-Beuren syndrome and plays a causative role in aortic stenosis.
Elastic Fiber synthesis
Problems with fibrillin?
Marfan Syndrome: defects in fibrillin 1 gene.
Glycosaminoglycans (GAGs) types
Hyaluronan (or hyaluronic acid) at the cell surface
main glycosaminoglycan in connective tissue
high molecular weight (~ MW 1,000,000 )
length of about 2.5 µm hyaluronan
“backbone” for the assembly of other glycosaminoglycans
Hyaluronan is also a major component of the synovial fluid, which fills joint cavities, and the vitreous body of the eye.
Other:
(attach through core and link proteins to hyaluronic acid backbone)
Chondroitin sulphate
Dermatan sulphate
Keratan sulphate
Heparan sulphate (UK sulphate, US sulfate)
What are gags made of?
Unbranched polysaccharides of repeating disaccharide units built from amino sugars and uronic acids.
Formerly: Mucopolysaccharides
How proteoglycans are formed?
GAGs are attached to protein cores.
The proteins can be bound to Hyaluronan
Function of GAGs and PGs
Provide flexible mechanical support to tissues.
Acts as a molecular sieve allowing the diffusion of small molecules but slowing the diffusion of proteins and the movement of cells.
Acts as a lubricant in joints and tissues subject to friction and compression/extension forces.
Binds and sequesters soluble ECM proteins thereby maintaining high local concentrations at the cell surface.
Cell surface receptors that mediate to the ECN and neighbouring cells
Dystroglycan Complex and Integrins
Cohesins
Condensin
Cohesins cross-link two adjacent sister chromatids, gluing them together.
Centrosome
Microtubule Organizing Center
MTOC
centrosomes at their – ends. + ends grow outwards towards the cell periphery.
Before a cell divides it must duplicate its centrosome to provide one for each daughter cell.
Centriole
Perpendicular cylindrical pairs
Microtubulles types
Astral
Kinetochore
Overlap
MAPS
Microtubule associated proteins (allow extension of microtubules)
Catastrophins (Depomylerization)
Separation of the two spindle poles
Kinesin
Overlaping MTs
Checkpoints in cell cycle
Enter mitosis
Exit mitosis
Enter S phase
Two key components of the cell-cycle controlled system
Cyclin-dependent kinease (Cdk)
Cyclin (oscillates)
Regulates of Cdk by ihibitory phosphorylation
Wee1 kinease (mutations cause uncontrolled replications making small cells)
Cdc25 phosphatase
CKI
Regulate Cyclin-cdk complexes
CDK INHIBITOR PROTEINS
p27
Rate limiting step in cyclin destruction
final ubiquitin transfer step by enzymes known as ubiquitin ligases
In G1 and S phase SCF is responsible for the ubiquitylation and destruction of G1/S cyclins and certain CKI proteins that control S phase initiation.
In M Phase, the anaphase–promoting complex (AMP) is responsible for the ubiquitylation of M-cyclins.
Initiation of DNA replication cycle
How is re-replication block insured?
Origin recognitioncomplex recognizes the ORC binding site
Cdc6-MCM forming pre-replicative complex
Cdc degradation
Origin recognitioncomplex becomes phosphorylated
(1) S-Cdk activtity remains high during G2
(2) M-Cdk ensures re-replication by phopshorylating Cdc6 and Mcm
The activation of M-phase
MCdk’s; Cyclin D
M-cyclin destruction
Destruction of M-cyclin is not required for sister-chromatid separation but is required for the subsequent exit from mitosis.
Mechanisms of sister chromatids separation
Control of G1 progression
G1 is held by active Sic1 and Hct1-APC
Mechanisms of S-phage initiation
DNA damage arrests the cell cycle
Phosphorylation of p53 leading to transcription of CKI p21
Overview of the cell-cycle
What growth factors control?
Example?
Regulation of cell growth or division.
Proliferation of cells.
Survival.
Migration
Physiological function of cells.
e.g. mitogen->RAS->myc
Cell determination
Cells retain a record of signals their ancestors received in early embryonic environment.
Cell determination before differentiation
Two ways of making sister cells different
Asymmetric cell division
Inducitve interactions (also concentration dependent; called morphogens); limited time and space; patterning by sequential induction)
Two ways to create a morphogen gradient
Gradient of inducer source
Gradient of inhibitor source
How embryo can be polar?
Fertilization triggers 2 types of intracellular movements.
Cell cortex rotation through 30 degree relative to the core of the egg in a direction determined by sperm entry.
Active transport of Dishevelled protein, a component of the Wnt signaling pathway.
The resultant Dorsal concentration of Dishevelled protein defines the dorsoventral polarity.
Stem cells
It is not itself terminally differentiated = Self renewal
It can divide without limit
Potency = The capacity to differentiate into any specialized cell.
When it divides, each daughter has a choice: it can either remain a stem cell, or it can embark on a course that commits it to terminal differentiation
4 cell types in the gut
- Absorptive cells - brush border cells or enterocytes.
- Goblet cells (as in respiratory epithelium) secrete mucus.
- Paneth cells form part of the innate immune defense system.
- Enteroendocrine cells, of more than 15 different subtypes, secrete serotonin and cholecystokinin (CCK).
Wnt/Notch signalling can alter formation of secreting or absorbin cell
High Energy Compounds values
Esters (amide)
Thiol-Esters (acetyl CoA)
Anhydrides (ATP)
Guanidinium phosphate (CP)
Enoyl Phosphate (PEP)
NADH
FADH2
Esters (amide): -3 kcal/mole
Thiol-Esters (acetyl CoA): -6 to -8 kcal/mole
Anhydrides (ATP): -7 to -10 kcal/mole
Guanidinium phosphate (CP): -10 kcal/mole
Enoyl Phosphate (PEP): -14 kcal/mole
NADH oxidation (to NAD+) -15 kcal/mole
FADH2 oxidation (to FADH+) -15 kcal/mole
Sources / Fates of Pyruvate?
Soruces / Fates of Acteyl CoA?
Order of substrates and enzymes in Krebs cycle
At which step is GTP / FADH2 / NADH produced?
Which steps are irreversible?
Which is the rate limiting step?
Which moleucules of Krebs cycles are exported out of the mitochondria and used as a regulatory signals?
Where and what signals control Krebs cycle?
Electron flow
Flavoproteins
examples?
of electrons that they can accept?
Ubiquinone
nickname?
of electrons it can accept?
location?
Coenzyme Q or just Q
1-2 electrons
Freely diffusible within inner membrane billayer
Which part is F1 and which F0?
Which one is rotating and which one synthesizes ATP?
The top is F1, and the bottom F0
F0 rotating
F1 ATP synthesis
How NADH is transported to mitochondria from cystol? (in liver, kidney, and heart)
Malate-Aspartate shuttle
How NADH is transported to mitochondria from cystol? (in muscle and brain)
glycerol 3 phosphate shuttle
Molecule that is found in brown adipose tissue
Thermogenin
Uncouples reacitons
Define mutation scanning (exon scanning)
When use it?
Examples?
Exons (coding regions within a gene) are subjected to physical tests to confirm the presence of a mutation before sequencing is used to delineate the exact mutation
When use it: If a gene has many possible mutations; reduces amount of DNA to be sequenced
Methods used include: conformation sensitive gel electrophoresis (CSGE), single-stranded conformational-polymorphism (SSCP), and denaturing gradient gel electrophoresis (DGGE).
Two ways that sickle cells can be detected
Protein electrophoresis
Allelic specific nucleotides
Hemoglobin lepore:
Testing?
It is a type of β thalassemia allele
Unequal crossing over between d and b genes generates hybrid db and bd genes.
Hb lepore (db) is functionally active, but expressed at low levels due to fetal promotor
1) Targeted mutation analysis (population appropriate),
2) Mutation scanning or sequence analysis.
