Cell Bio Exam 4

Front 1

List cytoskeletal systems and functions of bacteria and archaea

Back 1

MreB (like microfilaments) for dna segregation and cell shape

- crescentin makes rod shaped bacteria curved

FtsZ (like microtubules) for regulating division

Front 2

Types of microtubules and their location/funct

Back 2

Cytoplasmic

- found in cytosol

- maintains axons

- maintains cell shape

- forms mitotic/meiotic spindles (chromosome movement)

- moves and places vesicles

Axonemal

- found in cilia, flagella, and the basal bodies they attach to (think sperm)

- forms axoneme, a highly ordered bundle of microtubules that form the central shaft of cilia and flagella

- responsible for cell motility

Front 3

What is the structure of microtubules?

Back 3

Straight hollow cylinders of tubulin with longitudinal arranged polymers called protofilaments

- protofilament = alphabeta-heterodimer noncovalently bonded

-alpha and beta tubulins have almost identical 3d structure but only 40% amino sequence match

- alpha and beta each have n-terminal GTP binding domain, central colchicine binding domain, and c-terminal MAP domain (microtibule-associated proteins)

Front 4

What are tubulin isoforms?

Back 4

alpha and beta tubulin subunits with closely related but not identical Gene's (different isoforms may interact with different proteins)

Front 5

Explain microtubule polarity

Back 5

Dimers that make up microtubules are all oriented in the same way (n-terminus to c-terminus) so the entire microtubule has a positive end and negative end. The plus end grows faster

Front 6

What are singlet, doublet, triplet microtubules?

Back 6

Singlet - cytoplasmic

Doublet and triplet - axonemal

- one 13-protofilament A tubule and one or two 10/11 protofilament B and C tubules

Front 7

Describe microtubule assembly

Back 7

****with gtp and Mg2****

Lag (slow) phase:

Nucleation - dimers aggregate into oligomers

Elongation (fast) phase:

Addition of subunits at either plus or minus end

Plateau phase:

Disassembly due to diminished amt of free tubulin

Front 8

What is critical concentration? When does growth occur?

Back 8

The tubulin concentration at which assembly equals disassembly.

Growth occurs when tubulin concentration is above critical concentration

Front 9

How can microtubule plus and minus ends be visualized?

Back 9

Mix basal bodies (structures at base of cilia) with tubulin heterodimers. Plus end will grow faster

Front 10

What is treadmilling?

Back 10

Plus and minus ends have different critical concentrations.

Treadmilling - plus end is above critical concentration (growing) while minus end is under (disassembling)

Front 11

List drugs that affect microtubule assembly

Back 11

Colchicine - binds to tubulin monomers so they cant assemble into microtubules. Promotes disassembly

Nocodazole - also inhibits assembly but effects are more easily reverse than colchicine

Front 12

Explain dynamic instability (microtubules)

Back 12

each heterodimer has 2 GTP's - one on the alpha tubulin and one on the beta.

Once added to microtubule, beta's gtp hydrolyzes to gdp

If there is plenty of GTP-tubulin, it adds to the plus end to create a gtp cap.

If gtp-tubulin is low, new dimers are added at a slower rate than the hydrolysis of beta's GTP, leading to catastrophe (disassembly stops growth in favor of shrinkage). Reversing this effect (going from shrinkage to growth) is called rescue

See youtube Microtubule Assembly & Disassembly Mechanisms by Medical Sciences Animations

Front 13

Where do microtubules originate?

Back 13

From a microtubule organizing center (MTOC) that anchors and nucleates microtubules

- Usually a centrosome of 2 centrioles surrounded by pericentriolar material. Y-tubulin ring complexes (y-TuRC) nucleate microtubule assembly away from the centrosome with minus ends towards the MTOC (this means growth and shrinkage happens at the plus end)

Front 14

What is the structure of a centriole? Its function?

Back 14

9 triplet microtubules formed at right angles

Funct - helps form basal bodies, which help form cilia and flagella

- no centriole = poorly formed mitotic spindles

Front 15

What are MAPs? List types and funct

Back 15

Microtubule-associated proteins. Affects density of microtubule bundles; one region binds at intervals along microtubule wall and the other end extends out to allow interaction with other cell structures and filaments. Length of extension determines spacing/tightness of bundles

Tau Map - causes microtubules to form tight bundles in axons

Map2 - causes loose bundles in dendrites

Front 16

List proteins that encourage/discourage catastrophe

Back 16

+-TIPs (Tubulin Interacting Proteins) such as EB1 (End binding protein 1) protect growing plus ends

Stathmin/Op18 prevents polymerization by binding to heterodimers

Catastrophins promote subunit peeling at microtubule ends

Katanin cleaves microtubules

Front 17

Functions of microfilaments

Back 17

-Muscle contraction

-Cell migration (cell is made and moves to where it performs funct)

-Amoeboid protozoan movement

-Cytoplasmic streaming (cytoplasm moves)

-develop and maintain cell shape (from cell cortex beneath plasma membrane)

-forms structural core of microvilli

Front 18

How are microfilaments formed?

Back 18

Actin protein enter lag phase and elongation phase to form globular actin/G-actin that binds ATP or ADP (process is reversible)

2 linear G-actin form a helix (F-actin/microfilaments)

***all actin monomers are in the same orientation***

Front 19

List types of actin

Back 19

Alpha actins- muscle specific

Beta and Y actins - nonmuscle actins, (B and Y are localized to different regions of the same cell)

Front 20

Describe microfilament polarity

Back 20

Can be seen by incubating myosin subfragment 1 (S1) onto microfilaments and observing the resulting arrowhead pattern

Barbed end - plus (faster addition and loss)

Pointed end - minus

Like microtubules, atp hydrolyzes as g-actin monomers bind. Growing end has ATP-actin, the rest has adp-actin

Front 21

List drugs that affect microfilament polymerization

Back 21

Cytochalasins - fungal metabolites that block addition of new monomers

Latrunculin A - toxin that sequesters monomers and prevents their addition

Phalloidin - stabilizes microfilament. Prevents depolymerization

Front 22

List structures that microfilaments can form

Back 22

Immobile cells adhered to underlying substratum - thick organized bundles (stress fibers)

Mobile cells in cell cortex beneath membrane - crosslinked gel

Extremely mobile Cells that crawl- parallel straight filopodium and branched lamellipodium

Front 23

How is location and structure of microfilament formation controlled?

Back 23

Via actin-binding proteins at nucleation, elongation, and severing stages

Microfilaments assemble as long as ATP-g-actin is high, but in the cell, it is sequestered (bound) by thymosin B4. Profilin competes with thymosin to bind g-actin and does not sequester it. More profilin = more microfilament formation

Front 24

List microfilament cap proteins

Back 24

CapZ- binds to plus end, prevents addition of subunit

Tropomodulins -binds to minus ends, prevents loss of subunits

Front 25

What is filamin?

Back 25

Protein that cross links microfilaments by splicing them together where they intersect

Front 26

Funct of intermediate filaments

Back 26

Abundant in animal cells as keratin

Supports entire cytoskeleton as most stable and least soluble (more detergent, chemical resistant) cytoskeleton component

Forms nuclear lamina (fibrous mesh in dna replication and cell division)

Involved in cell structure scaffolding

Front 27

List classes of intermediate filaments

Back 27

1. Acidic keratin (epithelial surface)

2. Basic/neutral keratin (epithelial surface)

3. Vimentin (connective tissue)

Desmin (muscle cells)

Glial fibrillary acidic protein (glial cells)

4. Neurofilament proteins (nerve cell)

5. Nuclear lamina A B and C (network along inner surface of nuclear membrane)

6. Nestin (nerve cell neurofilaments of embryos)

Front 28

How are cytoskeletal elements integrated?

Back 28

Spectraplakins - linker proteins that connect MT, MF, IF

-plectin found where IF connects to MT and MF

Front 29

How is the cytoskeleton mechanically integrated?

Back 29

MT resist bending during compression

MF contracts and generates tension

IF are elastic and resist tension

Front 30

Describe intermediate filament structure/assembly

Back 30

Fibrous (rather than globular) dimers with N and C terminals that differ greatly among IF proteins

1. Two parallel IF polypeptides form a coiled coil dimer

2. Two dimers align to form a tetrameric protofilament

3. Protofilaments overlap to build an intermediate filament

Front 31

List signaling molecule classifications

Back 31

Classified by distance:

Long - endocrine signals produced far from target tissue; travel via circulatory system

Short - paracrine signals that are diffusable

Physical contact between sending and receiving cells - juxtacrine signals

Signals sent and received on same cell - autocrine signals

Front 32

What are receptors and ligands?

Back 32

Receptor - area on cell surface that binds a messenger; has a binding site or binding pocket

Ligand - a messenger that binds to the target cell

Front 33

What are 2nd messengers?

Back 33

1. Ligand binds to receptor

2. Binding induces production of additional molecules/ions in the cell (2nd messengers) that relay signals from one location in cell to the interior, causing a cascade of changes

Front 34

What is signal transduction?

Back 34

Ability of a cell to respond to ligand-recrptor binding by changing behavior or gene expression

Front 35

How are receptors altered after ligand binding?

Back 35

They change shape or cluster together.

Then, the receptor starts a preprogrammed chain of signal transduction events (cell changes behavior or gene expression), depending on cell's history, that lay dormant until they are triggered

Front 36

What are agonists and antagonists?

Back 36

Synthetic ligands that bind more tightly or selectively than real ones.

Agonist - receptor activators

Antagonist - receptor blockers

Front 37

What gives ligand-receptor binding its specificity?

Back 37

Binding site/binding pocket fits the messenger closely, like a puzzle piece

Amino acid side chains positioned to form chem bonds with the messenger

Front 38

How do cells shut down signaling?

Back 38

- reduce amount of ligands

- reduce receptor sensitivity (receptor desensitization) or amount.

-Cells sense changes in ligand concentration. Cell will stop responding to a certain ligand when receptors are already preoccupied for a long time (2 ways) so ligand concentration must increase to restimulate the cell

1.lower density of receptors by removing them via receptor-mediated endocytosis

2. Biochem changes that reduce affinity for ligand (ex: add or remove phosphate groups to amino acids)

Front 39

What is signal amplification? Examples?

Back 39

A small amt of ligand causes target cell to undergo a cascade of events. At each intermediate, more product is made.

Ex: epinephrine causes glycogen breakdown in liver cells. 1 epinephrine releases hundreds of millions of glucose from glycogen

Front 40

List cell-cell signal pathways

Back 40

1.Hydrophilic transmembrane receptors - Ligand-gated ion channels, proteins, small peptides, aminos and derivatives, nucleotides or nucleosides

2. G protein-coupled receptors - hydrolysis of GTP

3. cytosolic enzyme receptors - protein kinases

4. Hydrophobic Nuclear receptor - receptor bound to ligand (steroids and retinoids) enters nucleus to regulate gene transcription

Front 41

What do g protein-coupled receptors cause? Example?

Back 41

1. Ligand binding changes conformation change of receptor, which activates guanine nucleotide binding protein

2. Activated g-protein binds a target protein and changes its activity

Ex: opioid receptors

Front 42

What is the structure of a g protein-linked receptor?

Back 42

7 transmembrane alpha helices connected with alternating cytosolic or extracellular loops and a unique extracellular messenger binding site.

All have similar structure but different amino acid sequences

Front 43

How does phosphorylation regulate g protein-linked receptors?

Back 43

G protein-linked receptor kinase phosphorylates cytosolic amino acids on activated receptors.

Ex: amino acid such as B-adrenergic receptor is phosphorylated, then B-arrestin binds to it and inhibits it from interacting with g-protein

Ex: protein kinase A is activated by g protein-mediated signalling but then it phosphorylates (inhibits) other amino acids on the receptor; this is negative feedback

Front 44

How are g proteins activated or inactivated? List structure of g proteins

Back 44

G proteins turn on and off depending on whether they are bound to GTP or gdp

Structure - small monomeric proteins or large heterotrimeric proteins with Ga, Gb, and Gy subunits

-Ga is largest and binds to gdp or GTP

Front 45

Describe g protein activation/inactivation cycle

Back 45

1. Ligand binds to g protein-linked receptor

2. Receptor changes conformation and a g protein binds and releases its gdp

3.Ga binds to new Gtp molecule and detaches

4. Depending on g protein, either Ga or Gby initiates signal transduction

5. G protein stays active until Ga hydrolyzes GTP to GDP and rejoins Gyb (some Ga's are not efficient at hydrolysis)

6. If not efficient, regulators of g protein signaling (RGS) proteins are GTPase activating proteins that bind Ga to stimulate hydrolysis

Front 46

What is the most important g protein function?

Back 46

Formation of 2nd messengers, identified by the signaling process, such as cyclic AMP

Front 47

Example of Gby subunit signaling

Back 47

1.Acetylcholine binds muscarinic acetylcholine receptor

2. Gby subunit causes potassium channels to open, and close once acetylcholine is no longer present

Front 48

Describe regulation of cyclic AMP by g proteins

Back 48

1. Ligand binds to receptor and g protein attaches, losing gdp

2. Ga attaches to GTP and leaves to bind adenylyl cyclase

3. Adenylyl cyclase makes cAMP (a second messenger) from cytosolic ATP until Ga hydrolyzes and rejoins Gby

4. Remaining cAMP is degraded by phosphodiesterase

Front 49

Main function of cAMP?

Back 49

Binds to regulatory subunits of protein kinase A, causing them to dissociate from catalytic subunits, which are then free to phosphorylate target cell proteins on serine or threonine residues using ATP

Front 50

Diseases caused by disruption of g protein?

Back 50

Cholera -gs can't hydrolyze

whooping cough - pertussis keeps gi from inhibiting adenylyl cyclase

Front 51

How do g proteins act through inositol?

Back 51

1. Ligand binds receptor and activates Gq protein

2. Gq activates Phospholipase C (Cbeta), which cleaves PIP2 into IP3 second messengers and diacylglycerol (DAG)

3. IP3 diffuses through cytosol and binds ligand-gated calcium channel (IP3 receptor channel)

4. Calcium is released into cytosol

5. Calcium and DAG activate protein kinase C, which phosphorylate serine and threonine residues on various target proteins

Front 52

How do protein kinase-associated receptors function as kinases?

Back 52

Ligand binding stimulates tyrosine or serine/threonine kinase activity, and they transmit signals through phosphorylation cascade

Front 53

What are growth factors? List the ones that stimulate receptor tyrosine kinases

Back 53

Cells in vitro wont grow without blood serum with growth factor messengers

Insulin

Insulin-likengrowth factor 1

Fibroblast growth factor

Epidermal growth factor

Nerve growth factor

Front 54

What is the structure of a tyrosine kinase?

Back 54

1 polypeptide chain with 1 transmembrane segment.

Single - Ligand-binding domain outside, tyrosine kinase domain inside

Double - receptor and kinase are separate proteins. Non receptor tyrosine kinase binds to receptor and activates once ligand binds

Front 55

Describe autophosphorylation of tyrosine kinases

Back 55

1. Ligand binds

2. Some (fibroblast gf) receptors dimerize and phosphorylate each other

3. Some (epidermal gf) receptors cluster and the tyrosine kinase of each receptor phosphorylates its neighboring receptors

Front 56

Describe Ras signaling

Back 56

1. Autophosphorylation

2. Receptor recruits cytosolic proteins with SH2 amino acid domain to phosphotyrosine

3. GRB2 protein from the SH2 domain binds to Sos, a guanine-nucleotide exchange factor

4. Sos stimulates Ras G protein to release gdp and bind gtp

***some receptor tyrosine kinases activate phospholipase Cgamma***

5. Ras triggers a series of phosphorylations, starting with Raf

6. Raf phosphorylates serinenand threonine residues in MEK protein kinase

7. MEK phosphorylates threonine and tyrosine residues in mitogen-activated protein kinases (MAP kinases), activated to grow and divide by mitogens

8. Map kinases phosphorylate transcription factors

9. GTPase activating protein facilitates Gtp hydrolysis. GDP-bound Ras inactivates