1. Chapter 15 Cytoskeleton: Regulation by Accessory Proteins

2. g-tubulin ring complex (g-TuRC): minus end
Microtubules are nucleated by a protein
complex containing g-tubulin
Dynamics of
cytoskeletal
filaments:
Length
Stability
Number
Geometry
g-tubulin ring complex
(g-TuRC): minus end
Subcomplex:
accessory proteins
bound to g-tubulins
In a living cell,
cytoskeletal filaments
are more dynamic:
accessory proteins! By
Covalent modification
of subunits&
Accessory proteins

3. Microtubule-organizing
center (MTOC)
Centrisomes
star-like, “astral”
2 Centrioles
(L-shaped)
Centrosome matrix
(50 g-TuRCs)
Become basal
bodies, mitotic
spindles
Fungi (spindle pole
body) and plants (near
nuclear envelope) don’t
have centrioles but use
g-tubulin to nucleate

4. Actin filaments are often nucleated at the plasma membrane
“Cell cortex”:
shape and movement
of the cell surface
Spiky bundles: microvilli
or filopodia
Protrusive veils: lamellipodia
Two Actin-related proteins,
ARPs, nucleate actin filaments
45% identical to actin
The ARP complex
(Arp2/3 complex):
minus end
Attach to the side of another
actin filament:branches
Grow new filaments!

5. Plus end Profilin promotes polymerization
Thymosin keeps actins in monomer form
50% is soluble 50% in filament
Associates with plus end!
Plus end

6. One stathmin binds to two tubulin heterodimers and
prevents polymerization
Phosphorylation of stathmin
inhibits stathmin and increases
the rate of elongation and suppress
dynamic instability

7. MAPs (bind along the sides of MTs) in axon and dendrites
Of a neuron (MAP2 orange) (unphosporylated tau green)
Stablize MT against disassembly

8. Tropomyosin stabalizes actin filaments
Cofilin destablizes actin filaments by twisting the actin
Filaments a little more tightly--mechanical stress weakens
The contacts between actin subunits:brittle and ADP actin
Dissociate from minus end easier. Shorter lived than in test tube.
Aka
actin depolymerization
factor
Rapid turn over:
Binds to ADP actin

9. Capping proteins change filament dynamics by binding to ends
End-binding proteins can affect dynamics at low [ ]
Inactivate plus end
Minus end either capped
by ARP2/3 or uncapped
CapZ (muscle Z band)
Stable actin filament in muscle:
Plus end-CapZ
Minus end-tropomodulin(only
binds when coated by tropomyosin)
Inositol phospholipid, PIP2, uncaps plus ends

10. Proteins that bind to microtubule ends
Dynamic instability
Catastrophins:
kinesin-related
proteins
Capping proteins
Also control
Positioning:
Minus end--g-TuRC
Plus end--EB1, which
binds to Kar9
Directs spindle MTs into
the growing bud during mitosis

11. Actin filament organizations
Organization of MT by
Stabilizing proteins:
MTOC and MAPs
Actin organizing
is mediated by
a separate class of
Proteins (crosslinking)
Actin bundles
Weblike
Bundling proteins
Gel-forming proteins

12. (RBC) Web-like:spectrin & filamin Bundles! 3D gels!
Four actin-crosslinking proteins
Web-like:spectrin &
filamin
Bundles!
3D gels!
Stress fiber
Parallel filaments
Actin bundling proteins
Focal contacts

13. Filamin cross-links actin filaments to a 3-D network
Pigment-cell cancers
Melanoma cells
that have low
levels of filamin:
blebbing can’t
metastasize
Lamellipodia
is normal and
highly metastatic
when filamin is
restored
lamellipodia

14. Severing proteins regulate the length and kinetic behavior
The number of ends
increases when severed
Faster increase and
Decreast of filament mass
Gelsolin superfamily
Serves at high Ca++.
No energy requirement
katanin (sword)
breaks 13 protofilaments
Requires ATP

15. ERM family (ezrin, radixin and moesin) attaches actin filaments
to the plasma membrane
Regulated by
intracellular and
extracellular
signals

16. Focal contacts and stress fibers in a cultured fibroblast
Attachments between the actin filaments and ECM that
allows cells to pull on the substratum to which they are bound
Also relays signals: focal adhesion kinase (FAK)

17. “cytoskeleton?” The dramatic effects of Rac, Rho and Cdc42 on
actin organization in fibroblasts
Stress fibers
Focal
contacts
Lamellipodia
Membrane
ruffles
Filapodia
Microspikes
“cytoskeleton?”

18. Summary
Nucleation, polymerization
Proteins that bind to the sides
End binding proteins
Crossing-linking proteins
Severing proteins
Anchoring to the membrane
Rho-family GTPases and regulation by signals