L10: Cytokenesis Flashcards
what is cytokenesis and examples where it may not be needed?
Cytokinesis is the process that divides the cytoplasm of the parental cell into two daughter cells, completing the cell division process (following mitosis).
Cytokinesis usually accompanies mitosis in eukaryotic cells, but it doesn’t always occur after every mitotic event.
Cytokinesis Without Mitosis:
In some cases, mitosis can happen without cytokinesis, resulting in cells that have multiple nuclei (a process known as multinucleation). These cells may not fully divide into two separate cells, but the chromosomes still go through mitosis.
Example: Early Fly Embryo:
In the early fruit fly (Drosophila) embryo, after fertilization, there is a specific pattern of nuclear divisions in the absence of complete cell division (cytokinesis):
Anterior bicoid mRNA and posterior nanos mRNA are localized at different regions in the fertilized egg.
These mRNAs direct nuclear divisions that create a syncytium (a single, multinucleate cell).
Following multiple rounds of nuclear division, the nuclei migrate toward the cortex (outer part) of the egg.
Cell boundaries begin to form around the individual nuclei.
Cellularization is completed, forming individual cells.
This process allows for rapid divisions and distribution of cellular components before cellularization happens, which is crucial for the development of the embryo.
Specialized Animal Cells:
Some specialized animal cells, such as hepatocytes (liver cells), also undergo mitosis without cytokinesis, resulting in multinucleated cells. These cells still divide chromosomally, but they do not go through the usual steps of cytokinesis to physically separate into individual cells.
Cytokinesis in Animal Cells:
In most animal cells, cytokinesis involves the formation of a cleavage furrow that pinches the cell into two daughter cells. This occurs through the contractile ring, which is made of actin filaments and myosin motor proteins, ultimately dividing the cell’s cytoplasm and organelles.
Cytokinesis does not require an increase in cell volume — it is the process of physically splitting the cytoplasm, and doesn’t necessitate the cell’s growth during this stage.
actomyosin ring formation in cyotkenesis?
The formation and function of the actomyosin ring are crucial for cytokinesis, which is the final step of cell division. Here’s a breakdown of the process:
Assembly of the Actomyosin Ring:
The actin filaments and myosin II motors assemble at the inner cortex, which is the inner side of the plasma membrane (PM).
The contractile ring forms here, with actin filaments forming a structure that gets attached to the inner plasma membrane.
Myosin II, as a motor protein, interacts with the actin filaments. Myosin moves along the actin filaments, which causes the filaments to slide past one another. This sliding pulls the actin filaments together, resulting in the constriction of the actomyosin ring.
Contraction of the Ring:
As myosin II functions, it causes the actin filaments to slide towards their barbed ends (the plus ends), resulting in ring constriction.
Myosin’s motor activity causes the actin filaments to become shorter, pulling the ring tighter and driving the cleavage furrow to ingress (invaginate) deeper into the cell’s cortex.
Cleavage Furrow Formation and Ingression:
As the ring contracts, it drags the plasma membrane (PM) with it, resulting in the formation of the cleavage furrow.
The cleavage furrow continues to ingresses until it eventually reaches the nuclear midbody, which is the region between the two daughter cells at the final stages of division.
This ingression leads to the formation of the intercellular bridge, which is the final connection between the two daughter cells, before abscission (the final separation
actin, myosin and linker proteins?
Role of Actin, Myosin, and Linker Proteins:
Actin and Myosin generate the force necessary for cytokinesis, where:
Filamentous actin (F-actin) forms the core structure of the ring.
Myosin II motors help contract the ring by interacting with actin, sliding the actin filaments together.
Attachment to the Plasma Membrane (PM):
The actomyosin ring must be attached to the plasma membrane to function correctly. If this attachment doesn’t occur, cytokinesis cannot proceed.
This attachment is facilitated by linker proteins that connect the actin-myosin machinery to the plasma membrane and the medial cortex (the cytoskeleton just beneath the membrane).
Important Linker Proteins:
Formin, septin, and anillin are some of the key linker proteins involved in attaching the actomyosin ring to the cellular cortex.
Cdc15 and Mid1 are crucial proteins that help link the actomyosin ring to the plasma membrane. In mutants lacking both Cdc15 and Mid1, the actomyosin ring constricts in the cytoplasm rather than at the plasma membrane, meaning cytokinesis fails.
Failure to Attach to the Plasma Membrane:
If the actomyosin ring doesn’t attach to the plasma membrane, cytokinesis cannot occur. This means the cell will not divide properly.
RhoA GTPase?
RhoA GTPase in Cytokinesis:
RhoA Activation:
RhoA is a small GTPase, related to Ras, that exists in two forms:
Inactive form (RhoA-GDP): In its inactive state, RhoA is bound to GDP.
Active form (RhoA-GTP): When RhoA is bound to GTP, it is active.
The activation of RhoA is regulated by RhoGAP (GTPase-activating proteins), which can promote the hydrolysis of GTP to GDP, inactivating RhoA.
RhoA’s Effects When Active:
Once RhoA is active (GTP-bound), it has two major effects on cytokinesis:
Binding to Formin (Actin Nucleation):
Formin is an actin nucleator protein that promotes the formation of actin filaments.
RhoA-GTP binds to formin, which then promotes actin filament nucleation, contributing to the formation of the actomyosin contractile ring. This is crucial for the constriction of the cell during cytokinesis.
Activation of Rho-Activated Kinases (ROCK):
RhoA also binds to and activates Rho-activated kinases (ROCK).
ROCK then phosphorylates the myosin light chain, which activates myosin II.
This activation of myosin II enhances its ability to interact with actin filaments, promoting contraction of the actomyosin ring.
Additionally, ROCK inhibits myosin phosphatase, ensuring that the myosin II remains active during cytokinesis.
RhoA Localization:
The active form of RhoA (RhoA-GTP) is specifically localized to the equator of the cell prior to the formation of the cleavage furrow.
In sea urchin embryos, fluorescent probes that recognize RhoA-GTP have been used to show that RhoA accumulates at the cell equator before the cleavage furrow forms. This localization is important because it determines where the contractile ring will assemble, ensuring that cytokinesis occurs at the correct site in the cell.
RhoA’s Role in Defining Cytokinesis Site:
The accumulation of active RhoA at the equator of the cell defines the location of the contractile ring and, therefore, the site of cytokinesis.
This is a key step because proper site selection for the contractile ring ensures that the cell divides evenly and efficiently.
models for positioning the contractile ring?
Astral Stimulation Model:
Astral microtubules explore inside of the cell and meet at the middle.
These microtubules grow and shrink continuously, and when they meet at the middle, they stabilize.
This stabilization allows for the delivery of RhoA activators to the cortex where the microtubules are, and this defines the middle of the cell for cytokinesis.
This model is more likely in smaller cells, where the microtubules can reach and interact with the cortex more effectively.
Central Spindle Stimulation Model:
RhoA activators can diffuse from the central spindle (at the end of mitosis) and go to the cortex.
The cortex closest to the central spindle becomes the defined “middle” of the cell.
This model is more likely in larger cells, where the central spindle is farther away from the cortex and can influence a larger portion of it.
Astral Relaxation Model:
Astral microtubules explore the poles of the cell and induce a relaxation of the cortex.
This relaxation removes myosin from the poles, which causes it to accumulate in the middle by default, resulting in the formation of the contractile ring at the middle of the cell.
rappaport experiment?
Rappaport’s Experiment: The classic experiment involved manipulating the shape of the cell (in this case, making it a doughnut shape). Initially, when the cell undergoes mitosis, the cleavage furrow forms as expected between the two chromosomes.
First Division: The cleavage furrow forms between the chromosomes as it would in a normal spherical cell.
Horseshoe-shaped Cells: After the first division, the cell adopts a horseshoe shape. When the next mitosis occurs, two spindles assemble, one for each side of the horseshoe.
Unexpected Outcome: Instead of dividing simply at the “middle” (which would be expected if the central spindle determined cleavage), the cells actually divided into four parts, following the locations where the astral microtubules were positioned. In the Rappaport experiment, cleavage occurs not just between the centrosomes linked by the central mitotic spindle, but also between adjacent centrosomes. This leads to the formation of 4 daughter cells instead of 2. This shows that astral microtubules, not just the central spindle, can guide where cleavage happens, meaning multiple centrosomes can influence the cleavage plane.
Conclusion: This suggests that astral microtubules play a crucial role in defining where the division occurs, even in the absence of a clear central spindle. Astral microtubules from the poles appear to guide the formation of the contractile ring, determining where the cleavage furrow forms.
activating rho A?
Astral stimulation model: RhoA is activated by machinery in the spindle, and it’s also delivered by the astral microtubules to the cortex. This ensures RhoA activation is localized at the equator of the cell, where cleavage is needed.
Centralspindlin complex: The complex contains CYK-4 (a RhoA GAP, which activates RhoA) and MKLP1 (a kinase that helps in this process). CYK-4 activates RhoA by binding to its GTP-bound form, starting a cascade that leads to activation of Myosin II and nucleation of actin filaments.
RhoA activation cycle: RhoA must continuously cycle between active and inactive forms to regulate the process properly. If RhoA were always active, the cell couldn’t properly regulate the mechanics of cleavage.
The machinery involved sits in the microtubules at the middle of the spindle, coordinating the localization of RhoA activity at the correct site for cleavage.
activating rhoA + dyenin?
Activation of RhoA:
In anaphase, cells are in a non-contractile state because endogenous RhoA activation is prevented. However, RhoA can be exogenously activated by UV light.
Engineering System for RhoA Activation:
By shining light, the sensor activates RhoA. RhoA is activated in the middle of the cell, triggering an interaction with the cortex. This is sufficient to initiate actin-myosin constriction.
Dynein-Dependent Transport of Myosin II:
Dynein-mediated transport of myosin II promotes polar relaxation, which allows constriction to occur in the middle of the cell.
Polar Relaxation and Furrow Formation:
Dynein-mediated removal of myosin II from the cell cortex drives polar relaxation, bidirectional cortical flow, and furrow formation.
Cell Identity Variations:
The positioning of the myosin ring may vary depending on cell identity.
the midbody?
Midbody Formation: Dividing mammalian cells are joined at the midbody. After the cortex ingresses, the tubular cells still collide or connect with each other and remain joined by the midbody. The midbody is a remnant of the central spindle.
Structure of the Midbody: The midbody is composed of overlapping antiparallel interpolar microtubules in the center. These microtubules interdigitate (intertwine) and occupy space within the midbody. region of interiditated interpolar microtubules in midbody.
Secretion at the Midbody: There is a significant amount of secretion or vesicular activity occurring in the midbody region, which plays a role in the process of abscission (final separation of the daughter cells).
mibody and cleavage?
Cleavage Process: To cleave the cell, the plasma membrane must fuse to separate the individual cells. Before this, the spindle needs to be disassembled, and the cytoplasm must remain intact for the separation process to occur.
Central Spindle Components and Plasma Membrane: Central spindle components link the plasma membrane to the underlying cytoskeleton. The complex that activates RhoA also plays a role in organizing the midbody. These components crosslink microtubules and keep them close together in the midbody while interacting with the plasma membrane.
Mechanical Stability of the Furrow: The crosslinking and interaction between midbody components and the plasma membrane provide mechanical stability to the furrow. This is essential because, without it, the furrow may become unstable and fail to properly ingress.
Midbody and Cortical Stability: The midbody is crucial for ensuring that the cellular cortex remains ingressed. Additionally, other mechanisms involving microtubules, actin-interacting proteins, and membranes further contribute to maintaining the mechanical stability of the ingressed furrow during cleavage.
Central Spindle and Plasma Membrane Link: Central spindle components link the plasma membrane (PM) to the underlying cytoskeleton. Additional mechanisms, involving microtubules, actin-interacting proteins, and membranes, provide the mechanical stability necessary for the ingressed furrow.
Midbody Constriction: The midbody progressively constricts as cell division continues. At a certain point, the midbody begins to narrow from both sides. Eventually, one or both sides of the midbody fully constrict, completing the process of cellular division.
