Bio135 Final Exam Lecture Notes.txt

• Phospolipid bilayer
• Proteins
• Cholesterol (animals)

• Peripheral
• Integral (Transmembrane)

The portion facing the extracellular matrix

• Selective barrier
• In eukaryotes, formation of organelles/compartments
• Localization of enzymatic reactions
• Cell-cell communication
• Transmission/reception of signals
• Shape
• Receive stimuli
• Site of ECM attachment

• Golgi
• Cell membrane (i.e. not organelles)

It "evens out" enzymes since most are on cytosol side

A "microdomain" of the plasma membrane which aggregates proteins and phospholpids for transportation.

Via vesicle

• No.
• Glycosylation is for cell-cell recognition

• Useful for specialized enzymatic reactions.
• Greater rate of collisions for reactions to occur.

20 a.a.'s

• Intrinsic or associated enzymatic activity
• Provide cell with shape

• Receptors for soluble ligands
• Channel/gate
• ECM attachment (e.g. integrins)
• Cell-cell attachment (e.g. cadherins)

Hormones or action potentials

• Alpha helices
• Beta barrels

Facing the outside surface of the gate

Aquaporin

Provides membrane fluidity

• Rotation on axis
• Lateral (sideways)
• Flip-flopping (from one side to the other)

• Requires energy and is thus uncommon
• Requires the flippase enzyme
• Proteins are too large for this movement

CO2 and O2

The cell dies

• Change length of hydrocarbon tails
• Change saturation
• Change cholesterol content
• Change temperature

Increases fluidity

Increases fluidity

Decreases fluidity

Decreases fluidity

A signal that is received by the cell usually resulting in a response.

A feature on the cell which receives a ligand

• Intrinsically as an enzyme.
• Associated with an enzyme.
• With the cytoskeleton directly.

• Gene expression
• Metabolism
• Movement

They are proteins activated by intracellular signalling proteins

• Metabolic enzyme
• Gene regulatory protein
• Cytoskeletal protein (altered cell shape or movement)

• Direct contact between transmembrane molecules of two neighboring cells
• Paracrine system (local)
• Synaptic (neurotransmitters)
• Cytonemes (thin cytoplasmic extensions releasing hormones)
• Endocrine (via bloodstream)

Endocrine

• Contact dependent
• Maybe Cytonemes?

• Quick; < 1 hour; protein de/activation; synaptic
• Slow; 18-24 hours; transcription/translation; endocrine
• Single cell or group in development; autocrine

• Survive
• Grow + Divide
• Differentiate
• Die

• Heart muscle - decreased rate and force of contraction
• Skeletal muscle - contraction

• Soluble - interact with transmembrane receptors
• Insoluble - interact with cytoplasmic receptors

transcription

Vitamin D, estrogen, testosterone, cortisol, estradiol, retinoic acid, and thhroxine

• longer-lasting
• their duration in the blood

diffusion

• Early - first 30 min
• Delyated
• Depends on type of protein and timing of synthesis

• Activation
• Repression

Homodimers, heterodimers

Inhibitory

Activate other proteins for a delayed/secondary response.

• Early - male characteristics in development
• Delayed - muscle growth

• Early - female characteristics
• Delayed - retention of bone mass

• Ion-channel-coupled (open channel)
• G-protein-coupled (G activates enzyme)
• Enzyme-coupled (intrinsic and associated enzymatic)

• First messengers - ligands such as hormones
• Intracellular signaling proteins
• Second messengers - not unique to one pathway

cAMP, cGMP, 1,2-diacylglycerol (DAG), IP3, Ca+2

cAMP, cGMP

DAG, IP3

Anchoring, amplifier, integrator, modular, relay, scaffold, transducer

Anchors proteins to a structure at a precise location where needed.

Greatly increases signal they receive.

Combine signals from two or more pathways before forwarding.

Modify the activity of signaling proteins to regulate signal strength.

Pass messages to the next signaling component in the pathway.

Bind to multiple signaling proteins together in a functional complex for quicker and more efficient interaction.

Convert singal to a different form.

• Seven transmembrane spanning domains.
• Large cytoplasmic region that associates/activates with trimeric G proteins

No.

• Adenylyl cyclases
• Phospholipases
• Ion channels
• Protein kinases

alpha, beta, gamma

Different isoforms

The alpha subunit

move and activate other targets.

• Cell-specific receptors, G isoforms, and effectors
• Amount of receptors, G isoforms, and effectors
• Organization of signaling cascades
• Accessory proteins

They regulate the strength, efficiency, and specificity of the transmitted signal.

• GAP-43 - promotes GDP dissociation
• AGS3 - stabilizes G-alpha-GDP interaction
• Tubulin - directly transfers GTP to G-alpha

• Activators of G protein signaling (AGS)
• Regulator of G protein signaling (RGS)
• Inhibitors of GDP dissociation

Can activate G proteins without the use of a receptor

• Accelerate the GTPase activity of specific G-alpha subunits.
• Quick inactivation - hydrolysis

Cell division

• Golgi stability (alternative binding partners)
• Cell polarity in the fruit fly and nematode
• Neurite outgrowth and path-finding

• Adenylyl cyclase
• Phospholipase C-beta

• Increase of cyclic AMP concentration in the cytosol.
• This rise activates PKA.
• PKA enters the nucleus and phosphorylates CREB.
• CREB recruits CBP, and both stimulate gene transcription.

• inositol 1,4,5-trisphosphate (IP3) - releases Ca2+ from the ER
• diacylglycerol (DAG) - helps to activate PKC

• PLC-beta is activated by G protein (via alpha, beta/gamma, or both).
• Two messenger molecules, P3 and DAG, produced from hydrolysis of PIP2.
• IP3 releases Ca2+ from ER
• Ca2+ and DAG activate PKC

Prevents polyspermy by creating fertilization envelope

Delta, ligands

• NECD - Notch extracellular domain
• transmembrane domain
• NID - Notch intracellular domain

ADAM TM-NICD

A desintegrin and metalloprotease

• nucleus
• transcription, cell division, differentiation

• ligands
• family of secreted proteins

development

cancer

sonic, desert, Indian Hh

Patch

• Hip1
• Patched1
• Patched2

Smo

• cubitus interruptus (Ci)
• transcription factor

Desert

Typically by potency - Shh>Ihh>Dhh

cellular proliferation, growth, and axon path finding

• Holoprosencephaly
• Greig cephalopolysyndactyly syndrome
• Pallister-Hall Syndrome
• Gorlin's syndrome

• Basal cell carcinoma
• Rhabomyosarcoma
• Medulloblastoma
• Small cell lung cncer
• Pancreatic cancer

• Patch(Ptc) - membrane receptor
• Smoothened (Smo) - intermembrane protein
• Intracellular Hh Signaling complex (HSC)

• Membrane receptor which activates Smo when bound to Hh.
• After binding, Ptc levels decrease as a result of endocytosis

• Dimer
• Hip

Intermembrane protein that when activated relays signals to HSC

at the cell membrane

• Coastal 2 (Co2) - kinesin-related protein
• Fused (Fu) - Ser/Threo kinase
• Supressor of fused (Su/Fu)
• Cubitus Interruptus (Ci)

Gli1, Gli2, Gli3

truncated

HSC truncates Ci which becomes a repressor

Production of Ci which becomes an activator

In all but cell fate 1 nearest the posterior

In all but cell fate 5 (nearest the Anterior) which results in repressor.

[Hh] is proportional to activation

• Autocleavage
• Binding of cholesterol to C end
• Addition of palmitate to amino terminus

• Critical for target cell intake
• Critical for signal transduction after Hh binds to Ptc
• If binding is inhibited, Hh doesn't work

blocks the phosphorylation and stability of Smo.

G protein coupled receptor

Smo is found in endosomal vesicles

Smo is released and localizes to the cell membrane

high level signaling

• 155
• Weak activator - Ci^act
• Strong activator - Ci*

75

• PKA
• Glycogen synthase Kinase 3-beta
• Casein Kinase 1-gamma

• ubiquitinated
• Slimb (supernumerary limbs
• proteosome for cleavage

• Possibly a microtubule-motor
• Interacts with Smo
• Responsible for moving Smo

• Cos2 binds Fu to Smo
• Their interaction is critical for hh pathway signalling

• cell membrane upon Hh pathway activation.
• Intracellular vessicles in the absence of Hh ligand

• Has kinase activity, i.e. might phosphorylate Cos2 and Su(Fu) upon Hh pathway activation.
• Binds to Cos2 and Su(Fu) via carboxy terminus domain

• May be antagonistic to Fu
• No a.a. homology to known proteins
• Binds Fu and Ci, but not Cos2

Su(Fu)- and Fu- flies yield a wt phenotype

• May inhibit Ci activation
• Nuclear translocation of Ci
• Transcriptional regulation in vertebrates

• Ptc on cell membrane repressing/sequestering Smo at vesicle with HSC-A inactive.
• HSC-R on MT picking up vesicle leading to truncated Ci^75 by Su(Fu).
• Ci^75 to nucleus as repressor

• Hh binds Ptc at cell membrane permitting Smo/HSC-A to go to cell membrane
• HSC-R not at MT
• HSC-A allows Ci^act into Nucleus for low activation with possible low amount of Ci^75.

• Hh sequesters Ptc to vesicle permitting Smo Multimer/HSC-A to go to cell membrane
• Fu phosphorylates Cos2/SuFu
• SuFu leaves HSC
• Cos2 drives Ci to Smo
• HSC-R not present/inactivated.
• HSC-A allows Ci* to be processed untruncated into Nucleus by SuFu for high activation

• Formation of tumors
• Survival of tumors

• Gli forms MT-attached complex with Fu and SuFu
• Gli remains in cytoplasm

• Hh binds to Ptc
• Smo is activated (no longer supressed)
• Processing of Gli is activated
• Gli is translocated to nucleus

Ptch, Hip, Gli

• Cyclins D1 and D2 (mammalian) -> mitosis
• cMyc

• Cyclin B - Mitotic P Factor (MPF) - nuclear translocation
• P21 inhibition - tumor supression
• PDGF pathway activation (MAPK) - cell division

• LOF for tumor supression - Ptc and SuFu
• Overespression of oncogenes - Shh and Smo

medulloblastoma

cell nevus carcinoma

develop tumors

binding of Hh

Smo, Gli

• Cyclopamine - binds to Smo, but difficult to synthesize and toxic
• KAAD - modified cyclopamine - less toxic

• Forskolin - PKA activator - used in different pathways as well
• RNAi - Not feasible for treatment

ECM

cytoskeletons of the cells themselves (linked by anchoring junctions)

• Provides scaffolding and support for tissues and cells.
• Signal transduction.

proteoglycans, collagen, laminim, fibronectin, and vitronectin.

"filler" substance. Traps water. Binds cations.

Most abundant ECM component. Connective tissue.

Forms network of weblike structures that resist tensile forces.

Glycoproteins. Maintains cell shape.

Glycoprotein. Promotes cell adhesion and spreading.

• Cells must be attached to each other.
• Cells must be attached to a scaffold.

Protein-protein interaction between cells.

• Cells secrete proteins and carbs which make up the ECM.
• Intermembrane proteins connect the ECM with the cell's cytoskeleton.

Cadherins, selectins, integrins, Ig family

Tissue specific; dimerize; Ca+2 dependent; regulate cell shape and migration.

cadherin

Indirect binding

• Cell adhesion molecules that bind to sugars.
• Type of lectin.

Weak adhesion and rolling.

• strong adhesion and emigration
• Lets white blood cells exit capillary

• Adhesive
• Tight
• Gap

• Desmosomes and adherens.
• Hold cells together in fixed positions w/in tissues.
• Ca+2 dependent.

Desmosomes and adherens (both Ca+2 dependent)

• Keratin intermediate filaments connected to plaque.
• Plaque composed of anchor proteins.
• Transmembrane cadherin adhesion proteins attached to plaque.

desmoglein and desmocollin.

• desmoplakin
• plakoglobin
• plakophilin

structural support (not movement)

• Seal space between cells.
• Prevent flow of molecules and ions thru EC space.
• Important for organs that store liquids.

claudin and occluding

• Most common type of junction between animal cells.
• Form open channels between cells allowing ions and small molecules to pass.
• Useful for cell-cell communication.
• Open at low Ca+2 and low pH

calmodulin (also binds to calcium)

small (e.g. cAMP)

Secreted by the cells

• Used in cell-cell adhesion.
• Serve as attachment to ECM.
• Bind to specific ECM proteins.

• Procollagen triple-helix formed in ER/Golgi complex.
• Single procollagen molecule out via secretory vessicle.
• Cleavage of propeptides.
• Thousands of collagen molecules form fibril in ECM.
• Aggregation of fibrils form collagen fiber.

• Trap water and provide elasticity (e.g. skin).
• "Filler" substance.
• Hold ECM in place.

• 95% carbs by weight.
• glycosaminoglycan (GAG) is main component.
• Single polypeptide with hundreds of GAGs.

Linkages of proteoglycans to cell membranes.

• Receptors.
• Binding to ECM.
• Integrins binding to proeins in ECM.

fibronectin and laminin

Provides/maintains cell shape.

• Two large polypeptides (not identical) linked by disulfide bridges.
• Some domains bind to ECM.
• Other domains bind to membrane receptors.

By the a.a.'s flanking the RGD motif

• Receptors that mediate attachment between cells and ECM/other cells.
• Critical for growth, hemostasis, and host defense.
• Interact with cytoskelton.

• Heterodimeric with alpha and beta subunits.
• Variable subunits - mammals have 18 alpha, 8 beta
• e.g. melanoma: alpha-v, beta-3

• Binds to soluble and attached ligands.
• Binding changes conformation of the dimer.
• Binding is Mn+2 dependent.
• Clustering occurs with other integrin receptors upon ligand binding.

• Outside-in - info from outside to cell
• Inside-out - info from cell to outside

same target, cluster

Cells cease to be bound to ECM

Cell death, cell migration, cell shape change, cell division

Affects cell migration and differentiation.

• Angiogenesis and metastasis
• Focal adhestion tyrosine kinase -> cell survival
• MAP Kinase -> differentiation, cell growth, apoptosis

G1, S, G2, M

G0

G1/S and G2/M and in the middle of Mitosis

G1

S (synthesis)

G2

M

• Hormones
• Physical space

• Unhealthy cell - e.g. starving.
• Heavily damaged DNA

• Suicide
• Senescence

• Health
• Replicated DNA
• No more than 2 copies of chromosomes

Chromosomes must be attached to spindles

• Temp senstive mutants.
• Biochemical experiments with frogs.
• In vitro studies with tissue culture.
• BrdU pulse labeling.
• Flow cytometry.

• BrdU is a thymidine analog.
• Anti-BrdU Abs are used to stain chromosomal DNA
• Provides estimate of duration of each phase

S and G2 (when chromsomes are duplicated)

• Look at copies of chromsomes if they're labelled.
• Look for mitotic spindles.

G1

• Available space.
• Organ size.
• Signals/hormones.
• Proteins (e.g. cyclins)

• M -> M
• S -> M
• G1 -> S -> G2 (some plants and insects)

Something in S and M activate G1 (G1 is pushed into either of those phases)

• In the cytoplasm.
• Found by removing cytoplasm from M cell and placing in G1 cell.

• fusion
• cyclical

• Nuclear envelope breakdown.
• Chromosome condensation.
• Spindle fiber formation.

• high
• at the end of mitosis

a protein called cyclin

• p34 - CdK - phosphorylates
• p45 - cyclin - activates CdK

• G1/S - progression thru start.
• S - stimulate DNA replication.
• M - part of the MPF.

At the beginning of mitosis.

CdKs, but not CKIs (inhibitors)

• Wee1/Myt1 kinases.
• p27

Ubiquination

Cdc25

Proteolysis by APC/C and cdc20 and ubiquitin

phosphorylation/dephosphorylation

Adding two inhibitory phosphates.

Removing an inhibitory phosphates.

• an activating phosphate
• G2
• mitosis

• Phosphorylation
• Cyclin concentration

Figure 17-21 - overview of cell-cycle control system.

activates G1-CdK

inhibits G1/S-CdK, S-CdK, M-CdK

inhibits M-CdK

inhibits APC/C

• Activates S-Phase.
• inhibits DNA re-replication by initiating degradation of Cdc6.

• inhibits DNA re-replication.
• activates M-Phase.

allows passage thru mid-mitotic checkpoint.

• Mitogen binds to mitogen receptor which activates Ras.
• Ras activates MAP kinase pathway.
• Myc is produced which leads to expression of cyclin genes including G1-CdK.

• G1-CdK inactivates Rb which activates E2F
• E2F leads to S-phase gene transcription including G1/S-cyclin and S-cyclin.
• G1/S-cyclin and S-cyclin lead to active S-CdK which leads to DNA synthesis.

• E2F provides positive feedback for itself.
• G1/S-CdK and S-CdK further phosphorylate Rb providing pos feedback for E2F.

• inactivation
• cells to enter the S-phase

• Tumor suppressor protein.
• Trans-acting repressor that inhibits transcription of genes for S-phase.
• Becomes deactivated by phosphorylation by G1-CdK.
• Forms heterodimer with E2F protein.

Ubiquitin ligase.

• Represses transcription.
• Stimulates Skp2 removal at G0 resulting no S-phase.

• Rb is phosphorylated.
• Skp2 transcription increases.
• Skp2 proteolysis decreases resulting in cell division.

pre-replicative complexes.

• Formation of pre-initiation complex and initiation.
• Replication forks.
• Elongation.

• Chromosome segregation.
• Mitosis.

• APC/C activation.
• CdK inactivation.
• Assembly of new pre-replicative complexes at origins.

• 1. Gyrase - unwinds supercoils.
• 2. Helicase - unwinds dsDNA.
• 3. Polymerase - adds nucleotides.
• 4. Primase - adds primers (attachment points).
• 5. Ligase

Figure 17-23 - Control of initiation of DNA replication.

• Cdc6 and Cdt1 recruit 6 proteins and form the pre-Replicative Comples (pre-RC).
• S-Cdk stimulates assembly of the pre-initiation complex.
• DNA polymerase et. al are recruited to the origin.
• Mcm protein rings are activated as DNA helicases.
• DNA unwinds.
• Replication begins.

An APC/C target which inactivates Cdt1.

• S-Cdk triggers destruction of Cdc6 and inactivation of ORC.
• Cdt1 is inactivated by geminim (an APC/C target).
• Thus, a new pre-RC cannot be formed until end of mitosis.

• After the pre-replication complex is formed.
• Before S-CdK triggers S-phase.

The pre-initiation complex.

Prevent replication.

At the end of M-phase.

Receiving a signal.

• Remove receptor of start signal.
• Remove enzymes in signal transduction pathway.
• Remove CdK.

• Transcriptional activation of CKIs promotes G0 phase.
• Anaphase promoting complex (APC) and ubiquiting ligase promote G0 arrest.
• Remodeling of the chromatin.

Members of the bHLH family including MyoD and Hesl

• APC binds to Cdc20 or Cdh1 which activates ubiquiting ligase.
• S cyclins are removed.
• Skp2 is also degraded.
• This results in Rb binding to E2F proteins.

• Rb recruits histone deacetylase (HDAC) complex.
• HDAC associates with SWI/SNF ATP-dependent nucleosome remodeling complex.
• Rb binds to enzyme that methylates H3 histones.
• HP1 is recruited and keeps chromatin in repressive state.

Packs DNA together.

Repackages DNA tightly for activation of replication.

Blocks access to chromatin especially in areas that contain cyclin and promoter genes.

There is no such pathway.

Those cells can't divide and can't lead to cancer.

Water

• protein kinases
• initiate a signaling pathway that causes cell cycle arrest.

ATM or ATR

Chk1/Chk2

Phosphorylates p53 preventing Mdm2 binding resulting in CKI p21 production.

Binds to G1/S-CdK & S-CdK and inactivates them arresting cell in G1.

activation of Arf.

Binds and inhibits Mdm2 thereby increasing P53 levels.

Cell-cycle arrest or apoptosis depending on the cell.

metabolic pathway

• Increase in life span in C. elegans, Drosophila, and mice.
• Keeps mutant animals healthy with less age-related problems.

in the adult and not during developmental stages.

Smaller

C. elegans

• Age-1 - Phosphoinositide-3-kinase (PI3K)
• Daf-2 - insulin receptor
• Daf-10 - forkhead transcription factor

• InR - Insulin receptor
• Chico - fruit fly receptor

IRS1 - insulin receptor substrate 1

• age-1, daf-2
• age-1(-/-), daf-2(-/-)
• rescues

are twice as long.

• Insulin-signaling needs to be appropriate.
• Amount of insulin in resto of body (w.r.t. neurons) needs to lowered.

• inversely proportional.
• Exposure to oxidative stress.
• DNA damage

• InR
• extend

• dPTEN
• dFOXO

• secondary
• Insulin/Insulin-like Signaling (IIS)

Related to cardiovascular disease

ablation

young ovaries

• cancer
• cardiac problems
• Alzheimer's disease

• ligands
• invertebrate

• 1
• 4

mild growth defect in males

• short lifespan, diabetes (insulin resistance) in males.
• longer lifespan if brain-specific

extended lifespan in females

• Downstream mechanisms (e.g. oxidative stress).
• Multi-gene effects.