Dynamic DissonantsCell and Developmental Biology
• New York UniversityDavid ScicchitanoMark SiegalKris Gunsalus
• University of HawaiiSteve RobinowAthula Wikramanayake
• University of Wisconsin, MadisonBrian Manske
• Part I: structure, assembly and dynamics of microfilaments
• Part II: the cytoskeleton organizes the cytosol and supports the function of other cellular organelles and processes
• Part III:cytoskeletal filaments form higher-order structures that perform diverse functions
Actin Cytoskeleton Teachable Unit
• Chemical principles govern the behavior of biological macromolecules.
• Cellular phenomena reflect the collective action of populations of molecules.
• The same molecule can have multiple, diverse functions within cells.
Teaching Challenges
Diseases of the cytoskeleton• Muscular Dystrophy Dystrophin
– muscular degeneration
• Kartagener Syndrome
Axonemal dynein intermediate chain– Infertility due to defective sperm motility – Situs inversus
• Usher Syndrome myosin VII– deafness
• Epidermolysis bullosa and bullosa pemphigoid keratin mutation or autoimmune disorder
– skin blistering
Actin
Actin
Microtubules
Intermediate filaments
Are microfilaments the same as F-actin?
A. YesB. No
What polymerizes to form microfilaments?
A. alpha-tubulin
B. intermediate filaments
C. beta-tubulin
D. G-actin
The cytoskeleton is composed of:
A. microtubules
B. microfilaments
C. intermediate filaments
D. A and B
E. A and C
F. B and C
G. A, B, and C
Are microfilaments the same as F-actin?A. YesB. No
What polymerizes to form microfilaments?
A. alpha-tubulin
B. intermediate filaments
C. beta-tubulin
D. G-actin
The cytoskeleton is composed of:
A. microtubules
B. microfilaments
C. intermediate filaments
D. A and BE. A and CF. B and CG. A, B, and C
Are microfilaments the same as F-actin?A. YesB. No
What polymerizes to form microfilaments?
A. alpha-tubulin
B. intermediate filaments
C. beta-tubulin
D. G-actin
The cytoskeleton is composed of:
A. microtubules
B. microfilaments
C. intermediate filaments
D. A and BE. A and CF. B and CG. A, B, and C
synonymous to
Learning Goals
• To understand the structure, assembly and dynamics of microfilaments
• Apply chemical principles to enhance understanding of actin polymerization
• Predict and understand a biological model of actin polymerization
• Predict and interpret experimental results
G-actin: globular monomerF-actin: filamentous polymer
G-actin F-actin
Experiment
G-Actin
+
ATP
G-Actin
I
Results
F-actin is present in I only (at steady state)
II
Additional Experiment
G-Actin
+
ATP*
G-Actin
+
ADP
A. No F-actin in eitherB. F-actin in bothC. F-actin in ID. F-actin in II
ATP* cannot be hydrolyzed to ADP
I II
Actual Results
G-Actin
+
ATP
G-Actin
G-Actin
+
ATP*
G-Actin
+
ADP
Amount of F-actin at steady state+++ - +++ ++
Conclusion
For G-actin to form F-actin:
A. ATP hydrolysis is necessary.
B. ATP is necessary.
C. ADP is necessary.
D. Either ATP or ADP is necessary.
E. Neither ATP nor ADP is needed.
APPP
APP
F-actin
G-actin
Fast Slow
Summary
1.G-Actin has four lobes and an ATP binding site
2.In a test tube, add magnesium, sodium, potassium and either ATP or ADP:
G-actin F-actin
3. ATP binding to G-actin provides the best conformation to promote F-actin assembly
4. F-actin has directionality (the two ends are not the same)
G-actin ↔ F-actin ATP
Actin Dynamics
ATP
G-actin ↔ F-actin ATP
Summary
• G-actin and F-actin will reach a steady state.
• The nucleation step is slow.
Acknowledgements
Thanks to our facilitators!
• Randy Phillis
• Lauren Gollahon
