Chapter 15. Cytoskeletal Systems. Lectures by Kathleen Fitzpatrick Simon Fraser University Pearson Education, Inc.

Transcription

1 Chapter 15 Cytoskeletal Systems Lectures by Kathleen Fitzpatrick Simon Fraser University

2 Table Microtubules

3 Table Microfilaments

4 Table 15-1 Intermediate Filaments

5 Table 15-3

6 Microtubules Microtubules are the largest of the cytoskeletal components of a cell There are two types of microtubules They are involved in a variety of functions in the cell

7 Two Types of Microtubules Are Responsible for Many Functions in the Cell Cytoplasmic microtubules pervade the cytosol and are responsible for a variety of functions - Maintaining axons - Formation of mitotic and meiotic spindles - Maintaining or altering cell shape - Placement and movement of vesicles

8 Two types of microtubules (MTs) Axonemal microtubules include the organized and stable microtubules found in structures such as - Cilia - Flagella - Basal bodies to which cilia and flagella attach The axoneme, the central shaft of a cilium or flagellum, is a highly ordered bundle of MTs

9 Tubulin Heterodimers Are the Protein Building Blocks of Microtubules MTs are straight, hollow cylinders of varied length that consist of (usually 13) longitudinal arrays of polymers called protofilaments The basic subunit of a protofilament is a heterodimer of tubulin, one a-tubulin and one b- tubulin These bind noncovalently to form an abheterodimer, which does not normally dissociate

10 Figure 15-2

11 Figure 15-2A

12 Figure 15-2B

13 Figure 15-2C

14 Subunit structure a and b subunits have very similar 3-D structure, but only 40% amino acid identity Each has an N-terminal GTP binding domain, a central domain to which colchicine can bind, and a C-terminal domain that interacts with MAPs (microtubule-associated proteins) All the dimers in the MT are oriented the same way

15 MT polarity and isoforms Because of dimer orientation, protofilaments have an inherent polarity The two ends differ both chemically and structurally Most organisms have several closely related genes for slight variants of a- and b-tubulin, referred to as isoforms

16 Microtubules Can Form as Singlets, Doublets, or Triplets Cytoplasmic MTs are simple tubes, or singlet MTs, with 13 protofilaments Some axonemal MTs form doublet or triplet MTs Doublets and triplets contain one 13- protofilament tubule (the A tubule) and one or two additional incomplete rings (B and C tubules) of 10 or 11 protofilaments

17 Microtubules Form by the Addition of Tubulin Dimers at Their Ends MTs form by the reversible polymerization of tubulin dimers in the presence of GTP and Mg 2+ Dimers aggregate into oligomers, which serve as nuclei from which new MTs grow This process is called nucleation; the addition of more subunits at either end is called elongation

18 Microtubule assembly MT formation is slow at first, the lag phase, due to the slow process of nucleation The elongation phase is much faster When the mass of MTs reaches a point where the amount of free tubulin is diminished, the assembly is balanced by disassembly; the plateau phase

19 Figure 15-3

20 Critical concentration Microtubule assembly in vitro depends on concentration of tubulin dimers The tubulin concentration at which MT assembly is exactly balanced by disassembly is called the critical concentration MTs grow when the tubulin concentration exceeds the critical concentration and vice versa

21 Addition of Tubulin Dimers Occurs More Quickly at the Plus Ends of Microtubules The two ends of an MT differ chemically, and one can grow or shrink much faster than the other This can be visualized by mixing basal bodies (structures found at the base of cilia) with tubulin heterodimers The rapidly growing MT end is the plus end and the other is the minus end

22 Figure 15-4

23 Microtubule treadmilling The plus and minus ends of microtubules have different critical concentrations If the [tubulin subunits] is above the critical concentration for the plus end but below that of the minus end, treadmilling will occur Treadmilling: addition of subunits at the plus end, and removal from the minus end

24 Figure 15-5

25 Drugs Can Affect the Assembly of Microtubules Colchicine binds to tubulin monomers, inhibiting their assembly into MTs and promoting MT disassembly Vinblastin, vincristine are related compounds Nocodazole inhibits MT assembly, and its effects are more easily reversed than those of colchicine

26 Antimitotic drugs These drugs are called antimitotic drugs because they interfere with spindle assembly and thus inhibit cell division They are useful for cancer treatment (vinblastine, vincristine) because cancer cells are rapidly dividing and susceptible to drugs that inhibit mitosis

27 Taxol Taxol binds tightly to microtubules and stabilizes them, causing a depletion of free tubulin subunits It causes dividing cells to arrest during mitosis It is also used in cancer treatment, especially for breast cancer

28 Microtubules Originate from Microtubule- Organizing Centers Within the Cell MTs originate from a microtubule-organizing center (MTOC) Many cells have an MTOC called a centrosome near the nucleus In animal cells the centrosome is associated with two centrioles, surrounded by pericentriolar material

29 Centriole structure Centriole walls are formed by 9 pairs of triplet microtubules They are oriented at right angles to each other They are involved in basal body formation for cilia and flagella Cells without centrioles have poorly organized mitotic spindles

30 Figure 15-8A,B

31 Figure 15-8C

32 g-tubulin Centrosomes have large ring-shaped protein complexes in them; these contain g-tubulin (along with gamma tubulin ring proteins: GRiPs) g-tubulin ring complexes (g-turcs) nucleate the assembly of new MTs away from the centrosome Loss of g-turcs prevents a cell from nucleating MTs

33 Figure 15-9

34 Figure 15-9A

35 Figure 15-9B

36 MTOCs Organize and Polarize the Micotubules Within Cells MTOCs nucleate and anchor MTs MTs grow outward from the MTOC with a fixed polarity the minus ends are anchored in the MTOC Because of this, dynamic growth and shrinkage of MTs occurs at the plus ends, near the cell periphery

37 Figure 15-10A-C

38 Figure 15-10D

39 Microtubule Stability Is Tightly Regulated in Cells by a Variety of Microtubule- Binding Proteins Cells regulate MTs with great precision Some MT-binding proteins use ATP to drive vesicle or organelle transport or to generate sliding forces between MTs Others regulate MT structure

40 Microfilaments Microfilaments are the smallest of the cytoskeletal filaments They are best known for their role in muscle contraction They play a role in cell migration, amoeboid movement, and cytoplasmic streaming

41 Additional roles of microfilaments Development and maintenance of cell shape (via microfilaments just beneath the plasma membrane at the cell cortex) Structural core of microvilli

42 Actin Is the Protein Building Block of Microfilaments Actin is a very abundant protein in all eukaryotic cells Once synthesized, it folds into a globular-shaped molecule that can bind ATP or ADP (G-actin; globular actin) G-actin molecules polymerize to form microfilaments, F-actin

43 Figure 15-12

44 Figure 15-12A

45 Figure 15-12B

46 Figure 15-12C

47 Different Types of Actin Are Found in Cells Actin is highly conserved, but there are some variants Actins can be broadly divided into muscle-specific actins (a-actins) and nonmuscle actins (b- and g-actins) b- and g-actin localize to different regions of a cell

48 G-Actin Monomers Polymerize into F-Actin Microfilaments G-actin monomers can polymerize reversibly into filaments with a lag phase, and elongation phase, similar to tubulin assembly F-actin filaments are composed of two linear strands of polymerized G-actin, wound into a helix All the actin monomers in the filament have the same orientation

49 Demonstration of microfilament polarity Myosin subfragment 1 (S1) can be incubated with microfilaments (MFs) S1 fragments bind and decorate the actin MFs in a distinctive arrowhead pattern The plus end of an MF is called the barbed end and the minus end is called the pointed end, because of this pattern

50 Polarity of microfilaments The polarity of MFs is reflected in more rapid addition or loss of G-actin at the plus end than the minus end After the G-actin monomers assemble onto a microfilament, the ATP bound to them is slowly hydrolysed So, the growing MF ends have ATP-actin, whereas most of the MF is composed of ADPactin

51 Specific Drugs Affect Polymerization of Microfilaments Cytochalasins are fungal metabolites that prevent the addition of new monomers to existing MFs Latrunculin A is a toxin that sequesters actin monomers and prevents their addition to MFs Phalloidin stabilizes MFs and prevents their depolymerization

52 Figure 15-14A

53 Figure 15-14B

54 Figure 15-15

55 Actin-Binding Proteins Regulate the Polymerization, Length, and Organization of Actin Cells can precisely control where actin assembles and the structure of the resulting network They use a variety of actin-binding proteins to do so Control occurs at the nucleation, elongation, and severing of MFs, and the association of MFs into networks

56 Figure 15-19A

57 Intermediate Filaments Intermediate filaments are the most stable and least soluble cytoskeletal components and are not polarized An abundant intermediate filament (IF) is keratin, an important component of structures that grow from skin in animals IFs may support the entire cytoskeleton

58 Figure 15-22

59 Intermediate Filament Proteins Are Tissue Specific IFs differ greatly in amino acid composition from tissue to tissue They are grouped into six classes

60 Classes of intermediate filament proteins Class I: acidic keratins Class II: basic or neutral keratins Proteins of classes I and II make up the tonofilaments found in epithelial surfaces covering the body and lining its cavities

61 Classes of intermediate filament proteins (continued) Class III: includes vimentin (connective tissue), desmin (muscle cells), and glial fibrillary acidic (GFA) protein (glial cells) Class IV: These are the neurofilament (NF) proteins found in neurofilaments of nerve cells

62 Classes of intermediate filament proteins (continued) Class V: includes the nuclear lamins A, B, and C that form a network along the inner surface of the nuclear membrane Class VI: Neurofilaments in the nerve cells of embryos are made of nestin Animal cells can be distinguished based on the types of IF proteins they contain intermediate filament typing

63 Table 15-4

64 Intermediate Filaments Assemble from Fibrous Subunits IF proteins are fibrous rather than globular All have a homologous central rodlike domain conserved in size, secondary structure, and to some extent, in sequence Flanking the central helical domain are N- and C-terminal domains that differ greatly among IF proteins

65 Figure 15-23

66 Intermediate Filaments Confer Mechanical Strength on Tissues Cellular architecture depends on the unique properties of the cytoskeletal elements working together MTs resist bending when a cell is compressed whereas MFs serve as contractile elements that generate tension IFs are elastic and can withstand tensile forces

67 The Cytoskeleton Is a Mechanically Integrated Structure IFs are important structural determinants in many cells and tissues; they are thought to have a tension-bearing role IFs are not static structures; they are dynamically transported and remodeled The nuclear lamina, on the inner surface of the nuclear envelope, disassemble at the onset of mitosis and reassemble afterward

68 Integration of cytoskeletal elements Plakins are linker proteins that connect intermediate filaments, microfilaments, and microtubules One plakin, called plectin, is found at sites where intermediate filaments connect to MFs and MTs

69 Figure 15-24