Lecture 14 Flashcards
It was known for many years that material could move _______________ in cells?
Bi-directionally
What had already been discovered by the time kinesis were being examined?
Myosins had already been discovered; they used actin filaments as their cystokeletal framework
What had also already been discovered by the time kinesis were being examined?
Axonemal Dynein, a microtubule-associated motor had been shown to move flagella using microtubules
How did kinesin discovery start?
The discovery of kinesis really started with work from competitors/colleagues Robert Allen and Shinya Inoue
What did they use to greatly enchance the resolution during LIVE imaging?
The both used video cameras attached to microscopes that used polarized light to greatly enhance the resolution during LIVE imaging.
What was said in Inoue’s paper?
In this paper where he used polarized light to more clearly identify cellular components. This is of a Diatom
What was Inoue able to capture using his video capture technique?
He was able to capture fine detailed structures in real time…live! This is a 50nm acrosomal extension from Thyone sperm.
What does it mean to “optically” copy a video monitor image with another video camera?
This enabled them to ultimately substract one image (that was out of focus) from another image (that had the material they were interested in imaging). This enables the fine detail to be documented without the additional “junk” in the sample.
What technique did Robert Allen event?
Robert Allen invented a technique called “allen Video enchanced Contrast, Differential Intereference Contrast (AVEC-DIC) Microscopy”.
What does this technique show?
This technique could document microtubule-based motility of material in living things.
What kind of marine worms did Allen use?
Allen used the marine worm called Allogromia laticollaris for his studies.
When did Allen and Brady come together?
Allen and Brady came together when Brady worked with Ray Lesak as a postdoctoral fellow.
What did Brady use AVEC-DIC on?
There he used AVEC-DIC on Axoplasm from squid giant Axons
What is Axoplasm?
Axoplasm is essentially all of the stuff in the axon that is squished out onto a glass slide.
What did Robert Allen and Scott Brady study?
Robert Allen and Scott Brady had been studying the movement of material in squid giant axons for years. They documented the movement of material the sizes of small vesicales (30nm) all the way up to 5000nm (the sizes of organelles).
Who was Ron Vale?
Ron Vale was a graduate student at the time and went to Brady’s lab to learn his techniques of imaging axoplasm from squid giant axons. Discovered Kinesin in back to back papers in Cell
How did Vale acknowledge Brady, Allen and Lasek?
Barely acknowledged Brady, Aleen and Lasek
What did Brady do in return?
Brady (on his own) published 2 papers in Nature on the identification and partial characterization of a 130kDa ATPase (it was kinesis) that had properties expexted for fast axonal transport and caused organelles to bind to microtubules. But it was too late. They were submitted when Vale’s were in press
What do Kinesins use to move?
Use ATP hydrolysis to move primarily towards the + end of the microtubules
What is anterograde movement?
Moving towards the + ends of microtubules
What is retrograde movement?
Movement towards the - ends of the microtubules
What is Kinesin-1?
It is also known as Conventional Kinesis. Moves processively (they can do continuous movement without falling off of the microtubule). It moves cargo (like organelles, vesicles, etc…)
What is the structure of Kinesin-1?
Head(s): ~10nm in size (less than half the size of myosin heads), bind microtubules, catalyzes ATP hydrolysis.
There is a neck linker (between the head and tail).
Stalk (Tail): is a coiled-coil, binds cargo, has light chains (green) attached to it.
How do myosin heads compare to kinesin heads?
Myosin heads are bigger than kinesin heads, one binds actin and one microtubules. As a whole they are structurally very similar and are thought to have a common origin
What is the movement of kinesins?
It walks… takes 8nm steps usually on the SAME microtubule protofilament. That is the exact spacing of tubulin dimers. But, kinesin heads are only 10nm each. It can take these steps because of its neck. Moves at ~0.8um/s (so it steps once every 10 milliseconds)
