Cytoskeleton

Question 1

Microfilaments:

1. Localization
2. Function
3. Subunits and Filament
4. Polarity
5. Nucleation and Polymerization
6. Associated proteins
7. How to they generate motility?

Answer 1

1. location: filipodia at villi projections from cells; stress fibers across cells, amorphous gels (at cortical surface of the cell)
2. cell periphery (stress fibers); cell motility (lamellipodia, filopodia); contraction (w/myosin in contractile ring, sarcomere)
3. Subunits: G-actin. ATP-Mg2++ bound as a cofactor required for polymerization
4. +/- end with growth at + end
5. Nucleating factors (Arp 2/3 for lamellipodia, formin for filopodia) bind G-actin (which is bount to ATP) to make a trimer when high conc of ATP/G actin are present (rate limitng step). A long filament assembles (rapid event). Hydrolysis of bound ATP causes dissociation of monomers. Profilin promotes polymerization and Thymosin promotes dissociation of monomers. Rac controls polymerization for lamellipodia: Rac–> wASP–> Arp2/3–> lamellipodia. For filopodia, CDC42–> formin–> filopodia.
6. Myosin: most go to + end (except myo. 6)
7. 1. actin polymerization causes extension of free lamellipodium at leading pole 2. integrins root down which bind middle of cell 3. actin depolymerization and myosin contraction in the rear moves nucleus forward.

Question 2

Rho and CDC42 are __________, so an increase in GEF means an increase in _____, and increase in GAP means an increase in____. Rho-GTPases control____.

Answer 2

Rho-GTPases, so an increase in GEF means an increase in Rho-GTP (active)

Increase in GAP means an increase in Rho-GDP (inactive)

Rho-GTPases control nucleation factors to regulate formation of distinct actin arrays.

Question 3

Microtubules:

1. Localization
2. Function
3. Subunits and fIlament
4. polarity
5. Nucleation and polymerization
6. Associated proteins/ Cell movement

Answer 3

1. “skeleton” of the cell
2. move cili and flagella; move chromosomes during cell division; move organelles; provide tracks for intracellular transport; especially important for neuronal transport down axons.
3. dimer of alpha and beta tubulin subunts. Both bind ATP, but Beta hydrolyzes it. B is the + end. 13 protofilaments make a hollow MT. Nucleated at MTOC by gamma tubulin rings.
4. +/- end with growth at + end
5. MTOC = centrosome contains rings of gamma tubulin where MT build. MT undergo dynamic instability due to their GTP cap which can be hydrolyzed to GDP during “catastrophe” (cell shrink). TIP proteins (EB-1) are found at ends of MT and bind to RhoGTPase which anchors MT in place and stops dynamic instability.
6. kinesin kicks cargo out - to + (No kinesin, ER collapse). Dynein drags deliveries in: + to - . Axoneme in flagella and cilia is 9 doublets with dynein arms connected by nexin

Question 4

chemotherapy drugs vincristine and vinblastine, along w/colchicine, garlic, etc. are ______, while taxol is a _______

Answer 4

MT destabilizers

MT stabilizers

Question 5

IFs:

1. Functions/Location
2. Subunits and filament
3. Polarity
4. Associated proteins

Answer 5

1. Nuclear lamina; structural integrity (desmosomes, nucleus) (IFs do not cross plasma memb. into another cell)
2. tissue specific: keratins in epithelia; vimentin: connective tissue; desmin: muscle; GFAP: glia; neurofilaments: neurons; lamins: nucleus
3. no polarity
4. no polarity –> no motor proteins; plakins crossbridge IFs to MTs and actin; IFs get stronger w deforming force

Question 6

Epidermal bullosa

Answer 6

defect of IF insertion into plasma membrane junctions results in blistering in newborns

Question 7

when lamins are mutated, _____

Answer 7

can cause mislocatin of nuclei in muscle and bizzare aging phenotypes

Question 8

Lissencephaly

1. mechanism
2. symptoms
3. treatment
4. MOI

Answer 8

1. LIS 1 typically associates with dynein and allows it to apply a sustained force to the nucleus, resulting in neuronal migration. In Lissencephaly, neurons fail to migrate to normal layered arrangement at basal surface, disrupting function and connectivity of the brain.
2. faliure of the brain to develop gyrations during development, resulting in a “smooth brain” and therefore cognitive and functional impairment
3. ?
4. sporadic LIS 1 mutations

Question 9

Wiskott-Aldrich Syndrome (WAS)

1. Mechanism
2. Symptoms
3. Treatment
4. MOI

Answer 9

1. Mutations in the WASp gene, an effector of Arp 2/3. Typically loss of function: no activation of Arp 2/3, no actin polymerization, no release of megakaryocytes (low platelets–> defects in clotting ), no phagocytosis of bacteria. Leads to defective movement of WBCs (decreased immune function b/c WBCs can’t migrate across capillaries to sites of infection thru podosomes, made of lamellipodial structures). Sometimes, gain of function mutation prevents inhibitory domain of WASp from binding, so WASp is very active. Leads to decreased WBCs.
2. defect in clotting, immune function defects, low platelets decreased # WBCs (in gain of function mutation)
3. bone marrow transplant to replace dysfunctional cells
4. X-linked (typically found in males)

Question 10

Function of WAS in healthy person:

Answer 10

1. CDC42 GTPase activated by GEF.
2. CDC42 (GTPase) and PiP bind to WASp autoinhibitory domain, allowing WASp to become free and active.
3. Free WASp activates Arp 2/3, inducing actin polymerization

Question 11

Effects of defective Lis-1

Answer 11

1. Faliure of nuclear/cellular movements in neural progenitors and hence smooth brain defects of the cortex.
2. fungal organisms fail to to move nuclei along MTs
3. centrisome and nucleus fail to move and provide cell motility
4. abnormal brain progenitors because a huge force is needed to move these progenitors to the cortex

Question 12

Platelets are deficient in WASp disease because:

Answer 12

progenitor cells (megakaryocytes) fail to insert pseudopods into blood vessels