Cytoskeletal Microtubules & Microfilaments & Motility PDF

13 Cytoskeletal microtubules & Microfilaments & motility ILOs By the end of this lecture, students will be able to 1. Correlate the molecular organization of microtubules to its dynamic nature. 2. Interpret structural adaptation of MT to their function. 3. Correla...

13 Cytoskeletal microtubules & Microfilaments & motility ILOs By the end of this lecture, students will be able to 1. Correlate the molecular organization of microtubules to its dynamic nature. 2. Interpret structural adaptation of MT to their function. 3. Correlate the relevant motor proteins in cell trafficking to MT. 4. Appraise the importance of microtubules as a target for drug action. 5. Correlate molecular structure of actin to its function. Cytoskeleton The cytoplasm of animal cells contains a cytoskeleton, an intricate three-dimensional meshwork of protein filaments that are responsible for the maintenance of cell shape (Fig 1). Additionally, the cytoskeleton is an active participant in cellular motion, whether of organelles or vesicles within the cytoplasm, regions of the cell, or the entire cell. The cytoskeleton has three components: thin filaments (microfilaments), intermediate filaments, and microtubules. Fig. 1 The cytoskeleton Microtubules Microtubules (MTs) are long, straight, hollow structures that act as intracellular pathways. The centrosome is a region close to the nucleus that houses the centrioles, as well as several hundred ring- -tubulin ring complex molecules that act as nucleation sites for microtubules (fig. 2). Microtubules are dynamic structures that frequently change their length by undergoing growth spurts and then becoming shorter; both processes occur at the plus ends (oriented away from the nucleus), so that the average half-life of a microtubule is only about 10 minutes (fig. 2-B). Molecular structure of MTS Each microtubule consists of 13 parallel protofilaments composed of heterodimers of the globular 1  - -tubulin subunits. Fig. 2 Molecular structure of MTs A B Functions of MTs Provide rigidity and maintain cell shape. Regulate intracellular movement of organelles and vesicles. Establish intracellular compartments. Provide the capability of ciliary and flagellar (tail of the sperm) motion. During cell division, rapid polymerization of existing, as well as, new microtubules is responsible for the formation of the mitotic spindle. Clinical correlation The dynamic process of microtubule formation is disrupted by some drugs which can have different clinical effects on cellular functions e.g. Colchicine: By binding to the tubulin molecules in leukocytes, colchicine prevents its polymerization into microtubules. It thus interferes with leukocyte migration, phagocytosis and further release of inflammatory mediators, and this is the basis for its anti-inflammatory effect in acute gouty arthritis (Joint inflammation due to precipitation of urate crystals). Microtubule-associated proteins Microtubule-associated proteins are motor proteins that assist in the translocation of organelles and vesicles inside the cell. 2  Their primary functions are to prevent depolymerization of microtubules and to assist in the intracellular movement of organelles and vesicles. Movement along a microtubule occurs in both directions and is toward both the plus end and the minus end. The two major families of microtubule motor proteins, the MAPs dynein and kinesin, bind to the microtubule as well as to vesicles and organelles In the presence of ATP, dynein moves the vesicle toward the minus end of the microtubule. Kinesin effects vesicular (and organelle) transport in the opposite direction, toward the plus end. Centrioles Centrioles are small, cylindrical structures composed of nine microtubule triplets; they constitute the core of the microtubule organizing center or the centrosome. They are paired structures, arranged perpendicular to each other, and are located in the microtubule organizing center, the centrosome, in the vicinity of the Golgi apparatus. Functions of the centriole The centrosome assists in the formation and organization of microtubules as well as in its self- duplication before cell division. During cell division, centrioles are responsible for the formation of the spindle apparatus. Additionally, centrioles are the basal bodies that guide the formation of cilia and flagella (motile cell processes). 3  Fig. 5 Cilia & Flagellum Actin Filaments (microfilaments) Thin filaments (microfilaments) are composed of two chains of globular subunits (G-actin) coiled around each other to form a filamentous protein, F-actin. Thin filaments are 6-nm thick and possess a faster-growing plus end and a slower-growing minus end. Functional forms of actin 1. Contractile bundles: Their actin filaments are arranged loosely, parallel to each other, with the plus and minus ends alternating in direction. They form cleavage furrows (contractile rings) during mitotic division. Movement of organelles and vesicles within the cell. Cellular activities, such as exocytosis and endocytosis, as well as the extension of filopodia and cell migration (fig. 6-D). 2. Gel-like networks: provide the structural foundation of much of the cell cortex (fig. 6-C). 3. Bundles: form the core of microvilli (apical cell projections) (fig. 6-A). 4. Focal points: points of contact between the cell and the extracellular matrix (fig 6-B). 4  Fig. 6- Functional forms of actin A B Focal points F C D 5  14 Cell Cycle control & Mitosis ILOs By the end of this lecture, students will be able to 1. Interpret the significance of different phases of the cell cycle. 2. Correlate the phases of mitosis to normal and abnormal division outcomes. Cell cycle control The capability of the cell to begin and advance through the cell cycle is governed by the presence and interactions of a group of related proteins known as cyclins, with specific cyclin-dependent kinases (CDKs). Thus: Cyclin D, synthesized during early G1 phase, binds to CDK4 as well as to CDK6. Additionally, in the late G1 phase cyclin E is synthesized and binds to CDK2. These three complexes, through other intermediaries, permit the cell to enter and progress through the S phase. Cyclin A binds to CDK2 and CDK1 and these complexes permit the cell to leave the S phase and enter the G2 phase and induce the formation of cyclin B. Cyclin B binds to CDK1, and this complex allows the cell to leave the G2 phase and enter the M phase.(Figure 23-2) Fig 23-2. Control of the cell cycle 1 Fate of cyclins Once the cyclins have performed their specific functions, they enter the ubiquitin- proteasome pathway, where they are degraded into their component molecules. Cell cycle check points The cell also employs quality control mechanisms, known as checkpoints, to safeguard against early transition between the phases. These checkpoints ensure the meticulous completion of essential events, such as adequate cell growth, correct DNA synthesis, and proper chromosome segregation, before permitting the cell to leave its current phase of the cell cycle. The cell accomplishes such delays in the progression through the cell cycle by activating inhibitory pathways and/or by suppressing activating pathways. (check the details in Figure 23-3) Factors stimulating the cell to enter the cycle: The triggers inducing the cell to enter the cell cycle may be: (1) A mechanical force (e.g., stretching of smooth muscle), (2) Injury to the tissue (e.g., ischemia), and (3) Cell death. All of these incidents cause the release of ligands by signaling cells in the involved tissue. Frequently, these ligands are growth factors that indirectly induce the expression of proto-oncogenes, genes that are responsible for controlling the proliferative pathways of the cell. Clinical Hint Oncogenes and cancer: Normal cell proliferation and differentiation are controlled by a group of genes called protooncogenes; altering the structure or expression of these genes promotes the production of tumours. The expression of proto-oncogenes must be very strictly regulated to prevent unwanted and uncontrolled cell proliferation. Mutations in proto-oncogenes that enable the cell to escape control and divide in an uncontrolled way are responsible for many cancers. Such mutated proto-oncogenes are known as oncogenes. Altered oncogene activity can be induced by a change in the DNA sequence (mutation), an increase in the number of genes (gene amplification), or gene rearrangement. The Chromosomes Chromosomes are chromatin fibers that become so condensed and tightly coiled during mitosis and meiosis so that they are visible with the light microscope. 2  Each chromosome is formed of two sister chromatids attached at a point called the centromere. As the cell leaves the interphase stage and prepares to undergo mitotic or meiotic activity, the chromatin fibers are extensively condensed to form chromosomes, carrying the duplicated DNA (In which phase of the cell cycle?) Fig. 23-3. Structure of the chromosome 3 Types of cells in the human body 1. Somatic cells They are the body cells that are produced by mitosis and contain 46 chromosomes (diploid number), representing 23 homologous pairs of chromosomes. One member of each of the chromosome pairs is derived from the maternal parent; the other comes from the paternal parent. Of the 23 pairs, 22 are called autosomes; the remaining pair, which determines gender, are the sex chromosomes. The sex chromosomes of the female are two X chromosomes (XX); those of the male are the X and Y chromosomes (XY). Only one of the two X chromosomes in female somatic cells is transcriptionally active. The inactive X chromosome, randomly determined early in development, remains inactive throughout the life of that individual. The number of chromosomes in somatic cells is specific for the species and is called the genome, the total genetic makeup. 2. Germ cells They are either the sperm in male or ovum in the female and produced by meiosis. They contain 23 chromosomes (haploid number) and one sex chromosome either X or Y. Mitosis (M) occurs at the conclusion of the G2 phase and thus completes the cell cycle. Mitosis is the process whereby the cytoplasm and the nucleus of the cell are divided equally into two identical daughter cells. First, the nuclear material is divided in a process called karyokinesis, followed by division of the cytoplasm, called cytokinesis. The process of mitosis is divided into four distinct stages: prophase, metaphase, anaphase, and telophase Mitosis Mitosis is the process of cell division that results in the formation of two identical daughter cells. I- Prophase During prophase, the chromosomes condense, and the nucleolus disappears. Each chromosome consists of two parallel sister chromatids. As chromosomes condense, the nucleolus disappears. The centrosome also divides into two regions, each half containing a pair of centrioles and a microtubule-organizing center (MTOC), which migrate away from each other to opposite poles of the cell. 4  Fig. 23-4 Centrosome structure & prophase From each MTOC, astral rays and spindle fibers develop, giving rise to the mitotic spindle apparatus. It is thought that the astral rays (microtubules that radiate out from the pole of the spindle) may assist in orienting the MTOC at the pole of the cell. Those microtubules that attach to the centromere region of the chromosome are the spindle fibers, which assist in directing the chromosome alignment on cell equator and later migration of chromatids to the cell pole. In the absence of centrioles, the microtubule-nucleating material is dispersed within the cytoplasm with the result that astral rays and spindle fibers do not form properly, and mitosis does not proceed in the appropriate manner. (How is related to the cell cycle?) Spindle fibers bind to the kinetochore in preparation for chromatid migration to effect karyokinesis. II- Metaphase The nuclear envelope disappears early at this phase. The newly duplicated chromosomes align themselves on the equator of the mitotic spindle, being directed by the spindle fibers, and become maximally condensed. Each chromatid is oriented parallel to the equator, and spindle microtubules are attached to its kinetochore (a protein on the centromere that helps in movement of chromatids), radiating to the spindle pole (fig. 22-5). 5  Fig. 23-5. Metaphase & kinetochore III- Anaphase During anaphase, the sister chromatids separate and begin to migrate to opposite poles of the cell, and a cleavage furrow begins to develop (fig. 22-6). The spindle/kinetochore attachment site leads the way, with the arms of the chromatids simply moving along, contributing nothing to the migration or its pathway. In late anaphase, a cleavage furrow begins to form at the plasmalemma, indicating the region where the cell will be divided during cytokinesis. Figure 23-6 Anaphase & telophase IV- Telophase Telophase, the terminal phase of mitosis, is characterized by cytokinesis, reconstitution of the nucleus and nuclear envelope, disappearance of the mitotic spindle, and unwinding of the chromosomes into chromatin. 6. The chromosomes uncoil and become organized into heterochromatin and euchromatin of the interphase cell. Nucleolus reappears. Cytokinesis is the division of the cytoplasm into two equal parts during mitosis. The cleavage furrow continues to deepen (fig. 22-6). The remaining microtubules are surrounded by a contractile ring, composed of actin and myosin filaments attached to the plasma membrane. Constriction of the ring helps in separation of the two new cells. Each daughter cell resulting from mitosis is identical in every respect, including the entire genome, and each daughter cell possesses a diploid (2n) number of chromosomes. Clinical correlations Understanding of mitosis and the cell cycle has greatly aided cancer chemotherapy, making it possible to use drugs at the time when the cells are in a particular stage of the cell cycle. For example, vincristine and similar drugs disrupt the mitotic spindle, arresting the cell in mitosis. Colchicine , another plant alkaloid that produces the same effect, has been used extensively in studies of individual chromosomes and karyotyping. 7