lect 11: cytoskeleton Flashcards
what are vertebrate skeletons composed of? what does the human skeleton do?
vertebrate skeleton: composed of hardened elements (bones)
-human skeleton: supports the structure of our bodies and facilitates bodily movement
what are the learning objectives?
-the basic elements and functions of the eukaryotic cell cytoskeleton
-the structure and functions of intermediate filaments (IF)
-the structure and functions of actin filaments (F-actin)
-actin-binding proteins
-cell motility
-muscle contraction in relation to F-actin
what is the definition of the cytoskeleton?
cytoskeleton: “skeletal system” of eukaryotic cells is an intricate network of protein filaments that extends throughout the cytoplasm
-composed of a network of three filamentous structures
-together, they form interactive and dynamic network with different structures, properties and functions
what are the three filamentous structures that the cytoskeleton is composed of?
-intermediate filaments (IF)
-actin filaments (F-actin or microfilament)
-microtubules (MT)
what are the functions of cytoskeletal components?
- provide structural support that determines cell shape
- direct the movement of cargo and organelles within the cell
- generate forces needed for cellular locomotion
- position various organelles in the cell (spatial organization of cell organelles)
- make up an essential part of the cell division machinery responsible for DNA separation
what are intermediate filaments?
-intermediate filaments are strong, unbranched and ropelike
-IF strengthens cells against mechanical stress (epithelial, neurons, muscle)
-in specific cell types, such as epithelial cells, IF connect cells to other cells or the extracellular matrix via desmosomes or hemidesmosomes
-chemically heterogeneous group of structures (70 different genes)
what is common intermediate filament (IF) architecture?
- each monomer has a pair of globular terminal domains separated by a long fibrous alpha-helical region (polar)
- pairs of monomers are associated in parallel orientation to form dimers (polar)
- dimers associate in an anti-parallel, staggered fashion to form tetramers (nonpolar)
- 8 tetramers associate to form a unit length of the IF (nonpolar)
- these unit lengths associate with one another in an end-to-end fashion to form the elongated intermediate filament (nonpolar)
what is the first part of intermediate filament architecture?
IF monomer is polar
what is the second part of intermediate filament architecture?
IF dimer is polar
what is the third part of intermediate filament architecture?
IF tetramer is nonpolar
what is the fourth and fifth part of intermediate filament architecture?
IF is nonpolar
what are the four major classes of intermediate filaments?
these classes can include numerous subtypes. Humans, for example, have more than 50 keratin genes
what are keratin containing IFs? what are their functions?
keratin containing IF-structural proteins of epithelial cells
-IF radiate through the cytoplasm, tethered to the nuclear envelope in the center of the cell, and anchored at the outer edge of the cell by desmosomes and hemidesmosomes
-IF are also connected to microtubules and F-actin via plectins
-they organize and maintain cellular architecture and absorb mechanical stress
what is epidermolysis bullosa simplex?
a mutant form of keratin makes skin more prone to blistering. The defective protein assembles with the normal keratins and thereby disrupts the keratin filament network in the skin
what are neurofilaments (NFs)? what are their functions?
neurofilaments: bundles of IF located in the cytoplasm of mature neurons-oriented parallel to the axon to provide strength and stability
-aggregation of NFs is seen in several human neurodegenerative disorders such as parkinson’s disease
-these NF aggregated may block axonal transport on microtubules leading to neurons death
what is the assembly and disassembly of intermediate filaments?
-none of the IF assembly steps require the direct involvement of either ATP or GTP
-instead, assembly and disassembly of IFs are controlled primarily by subunit phosphorylation and dephosphorylation (ex. phosphorylation of vimentin filaments by protein kinase A leads to their disassembly)
-once assembled, the subunits are not incorporated at one end of the filament but into the filament’s interior in a dynamic fashion
what is the relationship between intermediate filaments and linker proteins?
-IF are often interconnected to other IF and cytoskeletal filaments by thin, wispy cross-bridges consisting of a dimeric protein called plectin (has various isoforms)
-each plectin dimer has a binding site for an IF at one end and a binding site for another IF, F-actin, or microtubule at the other end
what is the structure of actin filaments?
-actin filament (F-actin, microfilament) is a thin and flexible helical filament composed of actin monomers
actin protein is:
-one of the most abundant proteins in almost all cell types
-a major contractile muscle protein
-F-actin is a two-stranded helix with a plus end and a minus end (polar filament)
what is actin filament assembly and disassembly?
-actin filaments can grow by the addition of monomers at either end, but their rate of growth is faster at the plus end (10 times) than at the minus end
-before it is incorporated into a filament, an actin monomer binds an ATP molecules, and therefore, Actin is an ATPase
-the initial nucleation event in filament formation occurs slowly in vitro, whereas the subsequent stage of filament elongation occurs much more rapidly
-the ATP associated with actin monomer is hydrolyzed to ADP at some time after it is incorporated into the end of a growing filament
what is the first and second step of actin assembly/disassembly in vitro?
- preformed actin filaments (seed) are added in the presence of ATP-actin
- if the concentration of ATP-actin monomers is very high, subunits are added to both ends
what is the third step of actin assembly/disassembly in vitro?
- as ATP-actin monomers are consumed by addition to the ends of the filaments the concentration of free ATP-actin drops, until a point is reached where net addition of monomers continues at the plus end by stops at the minus end
what is the fourth step of actin assembly/disassembly in vitro?
- as filament elongation continues, the free monomer concentration drops, such that monomers continue to be added to the plus ends of the filaments, but a net loss of subunits occurs at their minus ends
what is the fifth step of actin assembly/disassembly in vitro?
- a point is reached where the two reactions at opposite ends of the filaments are balanced, such that the rate of monomer addition (at the plus end) equals that of monomer loss (at the minus end), the filament remains the same length (assembly=disassembly)-> this leads to F-actin moving in the direction of assembly, known as ‘treadmilling’
what is important in actin filament assembly and disassembly?
by controlling the assembly/disassembly, the cell can rapidly reorganize its actin cytoskeleton when and where is needed for dynamic processes such as:
-cell locomotion
-changes in cell shape
-phagocytosis
-cytokinesis
-assembly/disassembly rate can be influenced by several actin-binding proteins
what are actin-binding proteins?
-patterns of F-actin in living cells include various types of bundles as well as crosslinked and branched networks (but not in vitro)
-the organization and behavior of actin filaments inside cells are determined by the interaction of actin with a variety of actin-binding proteins
what are the categories of actin-binding proteins?
actin-binding proteins can be divided into categories based on their presumed function in the cell:
1. nucleating
2. monomer-sequestering
3. bundling
4. end-blocking (capping)
5. cross-linking
6. filament-severing
7. actin filament depolymerizing
what is essential in cell motility (crawling)?
the actin cortex is essential for cell locomotion
-three interrelated processes are known to be essential for cell motility
what are the three interrelated processes that are known to be essential for cell motility?
- the cell sends out protrusions at its “front”, or leading edge (by actin polymerization);
- these protrusions adhere to the surface over which the cell is crawling (forming focal adhesions); and
- the rest of the cell drags itself forward by traction on these points of anchorage
what does the graph of polymerization of F-actin in cell motility (crawling) look like?
what is the graph of what cell motility (crawling) looks like?
what are myosins?
the molecular motors of F-actin
-a protein superfamily of many classes
generally divided into two broad groups:
-myosin II: muscle cells (discovered first)
-other myosins (such as myosin I): most other cells
what is the function and structure of myosins?
-function as molecular motors that operates in conjunction with actin filaments
-move towards the plus end of an actin filament (with one exception)
-all myosins share a characteristic motor (head) domain
-whereas the head domains of various myosins are similar, the tail domains are highly divergent
the head domain contains two sites:
-a site that binds an actin filament
-a site that binds and hydrolyzes ATP to drive the myosin motor
what is myosin I?
-the head domain of myosin I binds to an actin filament and has the ATP-hydrolyzing motor activity that enables it to move along the filament in a repetitive cycle of binding, detachment and rebinding
-it moves towards the plus end of F-actin
-the tail binds to vesicles or the plasma membrane leading to their translocation
what are type II myosins?
-myosins II are primary motors for muscle contraction and also found in a variety of nonmuscle cells
-these proteins form dimers, with two globular ATPase heads at one end and a single coiled-coil tail at the other
-like most other myosins, myosin II heads bind F-actin and move toward the plus end of the filament
what do myosin II consist of?
myosin II consists of:
1. a pair of globular heads that contain the catalytic site of the molecular (binds to ATP)
2. a pair of necks
3. a single, long, rod-shaped tail formed by the intertwining of long alpha-helical sections
what does interacting F-actin and myosin II lead to?
muscle contraction
-the fibrous tail of myosin II plays a structural role so it can form filaments
-clusters of myosin II molecules binds to each other through their coiled-coil tails, forming a bipolar myosin filament
-because they are bipolar, the myosin heads at the opposite ends of a myosin filament can interact and pull actin filaments toward one another
what is muscle organization and contraction?
-a skeletal muscle fiber contains hundreds of cylindrical strands called myofibrils
-each myofibril consists of a chain of contractile units called sarcomeres
what is the organization of sarcomeres?
-sarcomeres are highly organized assemblies of two types of filaments: actin filaments (thin filaments) and myosin II filaments (thick filaments)
-the myosin II filaments are centrally positioned in each sarcomere
-the thin actin filaments extend inward from each end of the sarcomere and are anchored by their plus ends
what is the sliding filament model of muscle contraction?
what is the attached state of the sliding filament model of muscle contraction?
attached
-at the start of the cycle shown in this figure, a myosin head lacking a bound ATP or ADP is attached tightly to an actin filament in a rigor configuration (so named because it is responsible for rigor mortis, the rigidity of death)
-in an actively contracting muscle, this state is very short-lived, being rapidly terminated by the binding of a molecule of ATP to the myosin head
what is the released state of the sliding filament model of muscle contraction?
a molecule of ATP binds to the large cleft on the “back” of the myosin head (that is, on the side farthest from the actin filament) and immediately causes a slight change in the conformation of the domains that make up the actin-binding site
-this reduces the affinity of the head for actin and allows it to let go of the actin filament
-the space drawn here between the head and actin emphasizes this change, although in reality the head probably remains very close to the actin
what is the cocked state of the sliding filament model of muscle contraction?
the cleft closes like a clam shell around the ATP molecule, triggering a large shape change that causes the head to be displaced along the actin filament by a distance of about 5nm
-hydrolysis of ATP occurs, but the ADP and inorganic phosphate (P) produced remain tightly bound to the myosin head
-dashed lines show the position of the myosin head prior to ATP hydrolysis
what is the rebinding and power stroke state of the sliding filament model of muscle contraction?
weak binding of the myosin head to a new site on the actin filament causes release of the inorganic phosphate produced by ATP hydrolysis
-this release allows myosin to bind more tightly and triggers the power stroke-the force-generating change in shape during which the head regains its original conformation
-in the course of the power stroke, the head loses its bound ADP, thereby returning to the start of a new cycle
what is the attached state of the sliding filament model of muscle contraction?
at the end of the cycle, the myosin head is again bound tightly to the actin filament in a rigor configuration
-note that the head has moved to a new position on the actin filament, which has slid to the left along the myosin filament
