SFOM Podcast 2.1 Intercellular Motors

*Objective 1.

If given a direction of motor protein movement on a microtubule, be able to state if the protein is more likely a dynein or kinesin.

*Objective 4.

Outline the function of axonemal dynein and chromokinesin.

Cytoplasmic dynein

was purified much later and is responsible for minus end directed movement of cargo in the cytoplasm. - Dynein is much larger than kinesin and includes 2-3 heavy chains and a variable number of light and intermediate chains. - Head domains also bind microtubules and ATP. - Light and intermediate chains bind cargo. - Cargo bound to dynein moves toward the center of the cell.

*Objective 3.

Diagram kinesin 1 and dynein and identify those parts of the protein that bind microtubules, hydrolyze ATP and bind to cargo.

*Objective 2.

Diagram the polarity of microtubules within a typical cell during interphase and the movement of kinesin and dynein on these microtubules.

Dynein Types

There are two major types of dyneins. 1) Axonemal dynein 2) Cytoplasmic dynein

Anaphase B

- Anaphase B is synchronous with anaphase A but involves the position of the spindle poles. - Polar microtubules overlap at the midline and during anaphase they separate from each other - Via plus-end-directed kinesins that bind one microtubule and travel along the other. - The spindle moves towards the cell membrane by the action of dyneins anchored to the cell membrane. - These dyneins pull the poles toward the cell membrane. Anaphase B involves the positioning of the spindle poles. Polar microtubules make sure that the two microtubules don't overlap and just making sure one goes below the other and shove the two poles away from each other. Dyneins help move spindles toward cell center.

Anaphase

- In anaphase, the chromosomes separate. - Anaphase can be broken down into anaphase A and B. - Anaphase A: - Movement of chromosomes along the spindle. - Kinetochore microtubules depolymerize (break down) via a depolymerizing kinesin. - Chromosomes move toward the poles via an unclear mechanism. - Anaphase A involves the positioning of the mitotic spindle and specifically it breaking down/depolymerizing Depolymerization (or depolymerisation) is the process of converting a polymer into a monomer or mixture of monomers. All polymers depolymerize at high temperatures, a process driven by an increase in entropy.

Kinesin 1

- Kinesin 1 is plus end directed motor and is the classic kinesin. - 45 different kinesins have been identified in humans. Kinesin 1 has two heavy chains and two light chains. - The motor portion is within the heavy chains in the head domains and interact directly with the microtubules. - Head domains also bind and hydrolyze ATP to power protein movement. - Kinesin and myosin are structurally similar. - The opposite end of the protein consists of the light chains and the carboxy termini of the heavy chains. - Cargo being transported by the kinesin binds to this region. - Kinesin 1 moves cargo toward the periphery of the cell. - A few kinesins go the opposite direction. - Different kinesins are thought to bind different cargo. - There are three kinesins that don't move at all but act like anchors.

Motor Proteins

- Motor proteins can be divided into two large families: 1) Dyneins and 2) Kinesins - Both types move along microtubules within the cytoplasm. - All known dyneins move toward the minus ends of microtubules. - Most but not all kinesins move toward the plus ends of microtubules. - Microtubule minus ends are within the centrosome. - Plus ends are at the cell periphery. - Microtubules grow and shrink at their plus ends. Pic slide 4

1) Dynein Movement

- Move towards the minus (-) ends of microtubules - Dyneins move toward the centrosome.

2) Kinesin Movement

- Move towards the plus (+) ends of microtubules - Since most kinesins move toward the plus end of microtubules, they travel toward the cell periphery.

Vesicular Transport

- Movement of vesicles between different membrane bound organelles requires motor proteins. - For example, vesicle movement between the ER and the Golgi. - Vesicle movement is dictated by the motor protein associated. - ERGIC- ER golgi intermediate/intermediary compartment/complex - Reverse transport can occur where vesicles can be transported from ERGIC to center of cell

Motor Protein Direction

- Since most kinesins move toward the plus end of microtubules, they travel toward the cell periphery. - Dyneins move toward the centrosome. Pic slide 5 million

Positioning of Organelles

- The ER in a typical cell extends throughout the cytoplasm. - Microtubule inhibitors cause retraction (to pull away) toward the cell center. - Kinesin I motors appear to pull the ER toward the cell periphery. - Lysosomes are also positioned to the periphery of the cell via kinesins. - What about Golgi positioning? Via dyneins - This is kind of interesting b/c the diagram on the right is not very accurate- the purple part is the golgi, but the golgi tend to localized by dyneins and are typically positioned close to the centrosome and the nucleus for this reason.

Motors and M phase

- The microtubule network of an interphase cell disassembles as a cell enters M phase. - Microtubules then reorganize as the mitotic spindle. - Microtubules are more numerous (5-10X) and more dynamic (10x) in the mitotic spindle. - mitotic splindle = microtubules plus centrosomes

Cargo Transport

- Transport along microtubules has been extensively studied in nerve cells. - Ribosomes are absent from the axon so cytoplasmic proteins are synthesized in the cell body and then must be transported from the cell body to the axon. - Also organelles and vesicles must also move from the cell body to the axon. - Vesicles moving from the Golgi to the membrane of an axon would be carried by Kinesin 1 or Dynein? --> Kinesin 1 - In neurons, microtubules are not constricted to/bound to centrosomes. In a dendrite can have + and - ends of microtubules facing opposite directions so kinesin can travel toward the nucleus when the plus end is facing the center of the cell body. Axons= normal movement b/c aligned with - end of microtubule facing inwards and positive end of microtubule facing outwards. - Plus-end-directed kinesins transport vesicles and organelles to the periphery. - Minus-end-directed kinesins and all dyneins transport vesicles and organelles to the central regions of the cell.

Chromosome Movement

- Within the mitotic spindle, specialized microtubules called kinetochore microtubules interact with the chromosomes. - Kinetochore microbutules are stabilized by their association with kinetochores at the centromeres. - Chromosomal microtubules attach to the ends of chromosomes via a + end directed kinesin called chromokinesin. - Chromokinesin pushes chromosomes toward the midline - Chromokinesin only pushes chromosomes in towards the cell and the cell and to the midline of the mitotic spindle when they move outwards and the two forces balance each other. It's important that they push the kinetochores towards the center before mitosis begins. Not 100% understood. - If you look at a mitotic spindle- numbered differently. Kinetochore mts interact with kinetochores which are located at the centromere.

Axonemal dynein

- was the first isolated and is a critical component of cilia. - Axonemal dynein proteins are responsible for the movements associated with both cilia and flagella. - Movement of the dynein head along the B tubule toward the negative end causes the microtubule doublets to bend in the same direction. - Coordination of motor activity results in precise movement.

Intracellular Movements

A number of different processes within the cell involve intracellular movements. - Intracellular transport - Positioning of organelles - Positioning of vesicles near the cell membrane - Separation of Chromosomes - Cilia and Flagella Function - This presentation will focus on the role of motor proteins in these processes.