Lecture 18 - Key functional proteins in skeletal muscle Flashcards by Cale Dobrosak

What is a sarcomere/

Basic functional unit of striated muscle

repeated units

run between z-discs along the length of the myofibril

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What is a costamere?

Rib-like structures on the surface of the sarcolemma

specialised sites of transmembrane complexes, occuring over the entire sarcolemma, where the transmission of force is concentrated

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What is the role of the costamere?

Force transmission

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What are the protein complexes that transmit the force at the costamere

The Dystrophin-glycoprotein complex (DCG)

The Integrin-Talin-Vinculin complex

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Describe the DCG complex

The DGC is a mutimeric portein complex that acts to link the basal lamina to the actin cytoskeleton in skeletal muscle

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Dystrophin binds to ___ within the cytosol

actin

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Describe Dystrophin and its Function

Large rod-like protein enriched at skeletal muscle sarcomeres

Key intracellular component of the DGC

Contains 2 actin-binding domains

Fucntions:
Acts to link the Sarcolemma to the actin cytoskeleton by interactions with F-actin and Beta-dystroglycan

May also act to modulate signalling pathways through interactions with syntrophin and dystrobrevin

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If Dystrophin is missing, the connection between what is missing

basal lamina and actin cytoskeleton

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What are the different domains/regions of the dystrophin protein?

2 actin-binding domains:
One N-terminal composed of calponin homology domains
One centrally located composed of spectrin-like repeats

24 spectrin-like repeats (which create the rod-like structure)

4 hinge regions (for flexibility)

1 cysteine rich domains (site of protein-protein interactions)
- composed of a WW domain, two EF hand domains and a ZZ domain

1 C-terminal domain (site of interaction with syntropin and dystrobrevin

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What happens when dystrophin is defective or absent?

dissipation of forces throughout the myofiber become disrupted

membranes become permeable

high rate of muscle fiber breakdown and regeneration

DMD/ Becker’s MD (milder)

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In the absense of dystrophin, other DGC members fail to accumulate on the _______ indicating a role for dystrophin in ______ these proteins

In the absense of dystrophin, other DGC members fail to accumulate on the sarcolemma indicating a role for dystrophin in stabilising these proteins

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True or False

The absense of dystrophin results in muscles containing varying muscle fiber size, inflammatory cell infiltrate, accumulating fibrous tissue and membrane permeability

True

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True or False

Dystrophin also regulates multiple signalling pathways

True

via interactions with

Syntrophin/Dystrobrevin - coordinate the assembly of multiple signalling proteins to the DGC

nNOS - produces NO during muscle contraction to facilitate vasodilation

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What composes the Dystroglycan complex?

Laminin, a-dystroglycan, b-dystroglycan

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What is the function of the Dystroglycan complex?

Acts to link the ECM to the plsama membrane

Links the actin cytoskeleton of the muscle fibre by interaction with dystrophin

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The laminin protein family is trimeric, meaning?

It is comprosed of 3 chains needed for its functionality

a chain- necessary for interaction with receptors
y chain
B chain

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What is the function of the laminin protein

Forms a direct link between the ECM and dystroglycan to maintain skeletal muscle structure

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What happens when laminin is defective?

Congenital Musclular Dystrophy (MCD1A)

genetic deficiency of laminin a2 chain

variable muscle fiber size

presence of immature fibers (central nucleation)

Increased connective tissue (fibrosis)

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What is post-translational modification?

A step in protein biosynthesis

INcreases the ofunctional diversity of the proteome

Includes: phosphorylation, glycosylation, ubiquitination, nitrosylation, methylation, acetylation, lipidation, proteolysis

Regulates the activity, localisation and interaction of proteins with other cellular molecules

known to regulate the function of many of the key muscle proteins - only focusing on its role in dystroglycan

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True or False

the extra-cellular component of the Dystroglycan complex (a-dystroglycan) contains muscle sites for glycosylation

True

requires glycosylation for proper function (the receptor for laminin in skeletal muscle)

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True or False

Beta-dystroglycan is a transmembrane protein that binds a-dystroglycan and actin

False

Beta-dystroglycan is a transmembrane protein that binds a-dystroglycan and dystrophin

Function: provides a link between the subsarcolemmal cytoskeleton and ECM

Does Dystroglycan become dysfunctional?

No known association betwen mutations in dystroglycan and human disease

However, mutations in some glycosyltransferaess result in dystroglycan dysfunction as a result of a lack of glycosylation

Lack of glycosylation does not affect dystroglycan expression at the sarcolemma but is required for …

proper ligand binding (laminin)

Phosphorylation of beta-dystroglycn has recently been shown to …

target the b-dystrolgycan protein for destruction of not bound to dystrophin

Phosphorylation of beta-dystroglycn has recently been shown to ...

target the b-dystroglycan protein for destruction of not bound to dystrophin

What composes the Sarcoglycan complex?

alpha-sarcoglycan gamma-sarcoglycan sigma-sarcoglycan beta- sarcoglycan sarcospan

What is the function of the sarcoglycan complex?

Not fully understood appear to play both mechanical and non-mechanical roles that mediate interactions among the ACM, the sarcolemma and the cytoskeleton forms a lateral associated with the dystroglycan complex may function to stabilise the DGC at the sarcolemma

True or false Absense of sarcoglycans leads to alterations in membrane permeability, ultimately resulting in cell death

true genetic deletion of the sarcoglycan subunits of Limb-girdle muscular dystrophy reffered to as "sarcoglycanopathies'

What is the function of the Vinculin-talin-Integrin complex?

Focal adhesion complex of skeletal msucle Co-localises with the DGC at the costamere composed of integrin, talin and viculin force transducer

Integrin has to form a _____ to be functional

heterodimers composed of a and b subunits 22 heterodimers have been identified the a7B and Beta1D isoforms are the most common integrins

Which isoforms are the most common integrins found in adult skeletal muscle?

the aplha7B and Beta1D isoforms are the most common integrins

What is the function of integrin?

To mediate the processses of cell adhesion and migration Regulate the intracellular organisation of the actin cytoskeleton Important role in many signalling processes

Mutations in alpha7 integrin result in...

early onset CMD

What is Talin and how many isoforms exist?

Two isoforms (talin1 and talin2) Talin 1 is central for integrin signalling - interacts with the cytoplsmic domain of beta-1 integrin and with focal adhesion kinase and vinculin Binds to F-actin to establish a link between Beta1-integrins and the cytoskeleton

True or False Talin 2 is expressed at higher levels in skeletal muscle that Talin1

True but both Talin 1 and 2 are essential for skeletal muscle development

What does loss of the talin 1 gene in skeletal muscle lead to...

A progressive myopathy caused by mechanical failure of MTJs

Mice lacking talin 2 are viable and fertile but develop...

a myopathy with centrally located nuclei that is assocaited with defects in the maintenance of MTJs

Loss of both Talin 1 and 2 causes...

severe defects in myoblast fusion and sarcomere assembly

True or False Mutations in Talin have been identified

False

What is Vinculin?

membrane - cytoskeletal protein involved in linkage of integrin to the actin cytoskeleton

What is the Muscle specific splice variant of Vinculin?

metavinculin Mutations in metavinculin are associated with idiopathic dilated cardiomyopathy (DCM) in the human population

Why do we need two costameric protein complexes?

Loss of expression of proteins in each complex causes disease compromosed integrity of both complexes accelerates the develpoment of a severe muscular dystrophy Loss of dystrophin reults in a compensatory increase of integrin in dystrophic mouse models and DMD patients Therefore, the DGC and integrin complexes have both overlapping and compensatory functions

What is Desmin? | Where is it expressed and what is its function

intermediate filament protein - muscle specific type III Expressed in skeletal, cardiac and smooth muscle Function: links adjacent myofibres at the level of the Z disc and binds myofibrils to the sarcolemma at the level of the costameres

True or False Desmin is required for normal muscle development

False

Desmin is essential for the maintenance of the myofibril integrity following ____

stress

Is Desminopathy common and what causes it?

Very rare (only 60 patients to date) Mutation in desmin gene preventing the formation of desmin filaments. This results in misalignment of sarcomeres, disorganisation of muscle fibers, and muscle cell death

What is Plectin?

A large 500kDa protein located in nearly all mammalian cells Structural component of striated muscle Acts to link actin microfilaments, microtubules and intermediate filaments Approx. 12 different isoforms of plectin - 1, 1b, 1d and 1f most prominent

Where are the following isoforms of plectin localised: 1 and If 1d 1b

1 and If - localised to costameres 1d - associated with the z-disc 1b - localised to mitochondria

Plectin mutations in Epidermolysis bullosa simplex (EBS) and with MD affect the interaction between...

The plasma membrane and the cytoskeleton in skin and muscle resulting in blistering of the skin and late-onset muscle weakness

What is titin?

Largest known protein in nature Anchored at the Z and M lines of the sarcomere Maintains rigid contact with the thick filament along the A band Flexible along the I band ''Molecular bungee cord'of skeletal msucle - functions to protect muscle fibres from damage due to overs stretching

What are the 4 main roles of Titin in skeletal muscle?

1. Provide a structural framework through association with sarcomeric proteins 2. Centreing the thick filament within the sarcomere during contraction 3. Act as a molecular spring 4. Signalling mediator

What is the effect of Tininopathy?

Altered structure and function of Titin May disrupt Titin's interactions with other sarcomeric proteins Disrupts normal muscle contraction, resulting in msucle weakness and wasting over time Not known why disease is confined to lower leg muscles