Lecture 2 Cards Flashcards
Dimensions of an actin filament (aka microfilament)
4 - 6 nm diameter; 10 nm to over 100 micrometers long
Where are actin filaments most highly concentrated?
Cell cortex (just beneath the plasma membrane) - note they are also found throughout the cytoplasm, in lower numbers
Describe the structure of an actin filament
Polar structure with fast growing, plus (barbed) and slow growing, minus (pointed) ends; G-actin polymerized into F-actin; 2-stranded helical structure (see Figure 2.47, p.59)
How many actin genes are there? How many families (describe their distribution)?
Six genes in three families: alpha (skeletal muscle), beta and gamma (muscle and nonmuscle cells)
Describe the globular actin protein (size and enzymatic activity)
43 kDa (375 aa); has ATPase similar to that of hexokinase and hsp70
List six things that actin-binding proteins serve to do
Stabilize filaments, bundle or cross link filaments, fragment filaments, attach filaments to the membrane, bind actin monomers and increase actin microfilament dynamics
What does tropomyosin do?
Binds the groove of microfilament (stabilizes filaments)
What do fimbrin and alpha actinin do?
Link adjacent actin filaments into bundles
What does filamin do?
Creates actin microfilament gels
What does gelsolin do?
Severs actin filaments into shorter filaments
What does spectrin do?
Anchors actin filaments to membrane surfaces by binding microfilaments and membrane proteins
What does profilin do?
Binds soluble g-actin monomers, but in response to signals can release actin for rapid polymerization
What do capping proteins do? What are examples of capping proteins?
Increase actin microfilament dynamics, via regulation of cell shape, adhesion and motility; members of the Ena/VASP are capping proteins
List eight structural and motile functions of microfilaments
Membrane cytoskeleton, vesicle movements (streaming in plants, secretion in animal cells), cytokinesis (form contractile ring), cell locomotion, cell shape, cell adhesion, muscle contractility, metabolic compartmentalization
How do phalloidin and phallacidin act?
These phallotoxins from the deadly Amanita phalloides mushroom bind and stabilize F-actin
How does latrunculin A act?
This toxin isolated from a Red Sea sponge disrupts microfilament organization in cultured cells
How do cytochalasins act?
These toxins are a family of fungal metabolites that bind to the fast growing, barbed end to block assembly and disassembly at that end
How do bacteria such as Listeria and Shigella and viruses such as Vaccinia use human cytoskeletal elements?
They may use actin-based motility to spread and to move within cells, as a way of spreading an infection across tissue barriers like epithelia
What does the WASP protein do? What can result when it is mutated?
It is critical for rapid assembly of actin filaments at the cell surface. When mutated, Wiskott-Aldrich Syndrome (multiple defects in cellular immunity, lymphoid development, and maturation/function of myeloid monocytic cells) results.
What disease is correlated with mutations in the profilin 1 gene?
Some familial forms of Amyotrophic Lateral Sclerosis (Lou Gehrig?s disease or motor neuron disease) are caused by mutations in the profilin 1 gene
What diseases are associated with mutations in alpha cardiac actin?
Familial Hypertrophic Cardiomyopathy (FHC) - if mutation affects contraction; Idiopathic Dilated Cardiomyopathy (IDC) if the mutation affects force transmission from the sarcomere
What is the association between cancer and the actin cytoskeleton?
Cancer cells exhibit dramatic changes in the actin cytoskeleton, especially in metastatic cells (converting from a stationary cell phenotype to a motile form). Also, oncogenes such as Src and Rac are modulators of the actin cytoskeleton.
What is Celiac disease, and what is the link between Celiac disease and the cytoskeleton?
A lifelong autoimmune disease of the gut; components of the actin cytoskeleton are a common autoantigen in autoimmune diseases including this disease
Dimensions of a microtubule
25 nm outer diameter, a few micrometers to hundreds of micrometers long
Describe the structure of a microtubule
Made up of tubulin heterodimers (alpha and beta) - usually 13 heterodimers (can be 12-15) laterally arranged to form a hollow tube
Describe the size and genetics of tubulin.
50 kD; about half a dozen alpha genes, half a dozen beta genes and one gamma gene; highly conserved with significant sequence homology between the gene families
What enzymatic activity is associated with tubulin? Differentiate between alpha and beta tubulin.
Tubulins are GTPases; the beta tubulin GTP is readily exchangeable and hydrolyzes, while the alpha tubulin GTP is only poorly exchangeable
How does the nucleotide bound to the tubulin dimer correlate with assembly or disassembly?
The GTP form of the tubulin dimer favors assembly at the plus end; the GDP form favors disassembly
Where are microtubules generally found? From where do they originate?
Generally found in cytoplasm; originate from microtubule organizing center (MTOC) aka centrosome
Describe the structure of the MTOC, its placement within the cell and the orientation of microtubules with respect to the MTOC.
Centrioles surrounded by pericentriolar material, out of which cytoplasmic (and mitotic spindle) microtubules are nucleated by rings of gamma-tubulin
Usually found near the nucleus
Minus end of the microtubule remains attached; plus end is the growing end directed toward the plasma membrane
(see Figure 2.52, p. 64)
What is the structural basis of centrioles and basal bodies?
9 sets of circumferentially arranged triplet microtubules; each is a paired structure with two cylinders oriented orthogonally to each other (see Figure 2.57, p. 67)
How are cilia and flagellae formed?
They grow directly out of basal bodies (see Figure 2.54, p. 66)
Describe the structure of cilia and flagella
Axoneme is cytoskeletal core of cilia and flagella, consisting of 9 sets of circumferentially arranged doublet microtubules plus a pair of centrally located single microtubules
Differentiate between likely location of flagella versus cilia
Flagella are found in free-swimming cells, such as sperm
Cilia are found on cells which are part of solid tissues
Describe how cilia and flagella operate in movement
Both cilia and flagella beat, or gyrate, by a mechanism that depends upon dynein-induced microtubule sliding
Describe the atypical microtubules in basal bodies, centrioles, cilia and flagella
In cilia and flagella (doublet microtubules) and basal bodies and centrioles (triplet microtubules), one of the microtubules has the normal tubular form, but the others are incomplete tubules in which the wall of the adjacent microtubule forms part of their wall
What are two possible in vivo functions of microtubule-associated proteins (MAPs)?
Link microtubules to other cellular structures
Serve as scaffolds for proteins like kinases and phosphatases
What are the major MAPs in brain?
Tau (60-70 kD), MAP1 and MAP2 (high molecular weight, 250 kD)
What MAPs are commonly found in nonneuronal cells?
MAP4 and other high molecular weight MAPs
Name two things microtubule modifying proteins do
Enzymatic or structural proteins - introduce posttranslational modifications of tubulins (like acetylation or detyrosination) that may affect the interactions of microtubules with other cell structures
Modify microtubule dynamics
What do katanin and spastin do?
Cleave microtubules
What roles do cytoplasmic microtubules have in the cell during interphase?
Act as highways for transport of membrane-bounded organelles (secretory and endocytotic vesicles; transport intermediates between the ER and Golgi; lysosomes; mitochondria); determine cell polarity and cell shape; consolidate cell movements
What roles do cytoplasmic microtubules have in the cell during mitosis and meiosis?
Establish the spindle for segregation of chromosomes (karyokinesis)
As an aside, the major stages of mitosis are: prophase (nuclear membrane breakdown and chromosome condensation); metaphase (formation of spindle; chromosomes align at center of the spindle); anaphase (chromosomes separate); telophase (cleavage furrow forms, and cytokinesis occurs; nuclear membranes form around newly formed daughter nuclei)
How does the drug Vincristine act?
Microtubule destabilizing; chemotherapeutic agent for cancer - interferes with spindle formation and progression through mitosis, thereby blocking cell division of malignant cells
How does the drug Taxol act?
Microtubule stabilizing; chemotherapeutic agent for cancer - interferes with spindle formation and progression through mitosis, thereby blocking cell division of malignant cells
What happens when the microtubule-associated protein Tau aggregates?
Alzheimer?s Disease; Tau aggregates form neurofibrillary tangles, whose density and distribution correlate with loss of synaptic contacts and dementia
What happens when the gene for the microtubule-associated protein tau is mutated?
Frontotemporal Dementia and Parkinsonism (FTDP-17) associated with degeneration of cortical neurons
What does spastin normally do? What does a mutation in spastin cause?
Fragments microtubules; most frequently mutated gene in Hereditary Spastic Paraplegia
What are three proteins whose mutations cause lissencephaly?
Lis1, tubulin alpha 1a and doublecortin
What happens to the brain in lissencephaly?
Alterations in cortical layering due to altered neuronal migration - a thickening of the cortex and smooth surface, lacking the folds and ridges of a normal brain; chages in microtubule function suggested to be underlying cause
Distinguish between the effects of the mutations that cause lissencephaly
LisI mutants (similarly in tubulin alpha 1a) - neurons fail to migrate so the layering of the cortex fails to form, leading to severe retardation and early death
Hemizygous doublecortin mutants in males have a classic lissencephaly
Heterozygous mutations in females have double cortex
What are the effects of mutations in Beta3 tubulin?
Congenital fibrosis of the extraocular muscles type 3
May lead to cognitive and behavioral disorders, facial paralysis and a late onset sensory polyneuropathy
Dimensions of an intermediate filament
10 nm in diameter; 10-100 micrometers long
Describe the structure of an intermediate filament
core domain of coiled-coil alpha helix most conserved across families; rod shaped structure confers structural stablility; flexible and mechanically strong
What is the common name of Type I and Type II intermediate filaments, and where are they found?
Keratins - epithelial cells
Name three Type III intermediate filaments and where are they found
Vimentin - many cell types including fibroblast and early differentiation
Glial fibrillary acidic protein (GFAP) - astrocytes
Desmin - smooth muscle cells
What is the common name of Type IV intermediate filaments, and where are they found?
Neurofilaments - all vertebrate neurons
What is the common name of Type V intermediate filaments, and where are they found?
Lamins - nuclear envelope
Name some features of intermediate filaments that distinguish them from actin filaments and microtubules
Higher degree of cell specificity; flexible even when bundled; most stable; no role in motility (only structure); nonpolar filaments
What is BPAg1e, and what can occur when it is mutated?
Intermediate filament-associated protein; 230kD splice variant of BPAG1 in epithelial cells; binds keratin filaments to hemidesmosomes; mutants cause a blistering disease
What is BPAg1n, and what can occur when it is mutated?
Intermediate filament-associated protein; 280 kD splice variant of BPAG1 in sensory neurons; cross-links neurofilaments to membrane cytoskeleton; mutants have an axonopathy
What is filaggrin?
37kD intermediate filament-associated protein; found in cornified epithelia; aggregates keratin
Where are lamin-associated proteins found?
N uclear envelope (note they are 57-75 kD)
What is emerin, and what can occur when it is mutated?
34kD intermediate filament-associated protein; found in inner nuclear membrane; mutants develop Emery-Dreifuss muscular dystrophy
What is plectin, and what can occur when it is mutated?
Intermediate filament-associated protein; 500 kD homodimer; links keratin to other cytoskeletal elements; mutant causes blistering disease
What disease is associated with keratins in the skin?
Epidermolysis Bullosa Simplex (EBS) (diseases of skin blistering: differ in location and severity with gene affected)
What disease is associated with the IF associated proteins plectin and BPAG 1?
EBS variants (with muscular or sensory neuron involvement)
What disorder is associated with keratins in various epithelia?
Stress induced degeneration (hair, nails, esophagous, and cornea)
Name three diseases/disease classes associated with neurofilaments
Amyotrophic Lateral Sclerosis and some other neurodegenerative diseases exhibit accumulations of neurofilaments in the initial segment of the axon
Mutations in neurofilaments may be associated with some forms of ALS-like motor neuron disease or some forms of Charcot-Marie-Tooth peripheral neuropathies
What diseases are associated with lamin mutations?
Progeria (a rare, premature aging syndrome), cardiomyopathy, muscular dystrophy
Describe three cytoskeletal element-related abnormalities linked to familial Amyotrophic Lateral Sclerosis (ALS, or Lou Gehrig?s Disease)
Mutations in the profilin 1 gene (actin-associated protein)
Mutations in the alpha-tubulin gene TUBA4A (microtubule component)
Accumulations of neurofilaments in the initial segment of the axon (intermediate filament)
