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Dystrophin and the Extracellular Matrix. Eric Niederhoffer SIU-SOM. Protein domain structure Dystrophin glycoprotein complex Contraction unit Cytoskeletal linkages at sarcolemma Muscle proteins and disease Creatine kinase and CKI In situ staining and Western blots.

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dystrophin and the extracellular matrix

Dystrophin and the Extracellular Matrix

Eric Niederhoffer

SIU-SOM

  • Protein domain structure
  • Dystrophin glycoprotein complex
  • Contraction unit
  • Cytoskeletal linkages at sarcolemma
  • Muscle proteins and disease
  • Creatine kinase and CKI
  • In situ staining and Western blots
dystrophin domain structure

Dystrophin Domain Structure

Domains of the dystrophin molecule. The N terminus contains the primary actin-binding site, whereas the C terminus contains the -dystroglycan, dystrobrevin, and syntrophin-binding sites. The N- and C-terminal domains are connected by 24 spectrin-like repeats, some of which have been shown to bind actin. The four "hinge" regions are denoted H1-H4.

http://www.pnas.org/cgi/content/full/97/25/13464/F2

dystrophin glycoprotein complex
Dystrophin Glycoprotein Complex

Schematic representation of the dystrophin-associated glycoprotein complex. The N-terminal, actin-binding domain of dystrophin in purple is associated with the cortical actin. The C-terminal domain associates with b-dystroglycan and with a- and b-syntrophin and dystrobrevin. nNOS is known to interact with the syntrophins as well as with caveolin.

http://www.pnas.org/cgi/content/full/97/25/13464/F1

contraction units
Contraction Units

http://cellbio.annualreviews.org/cgi/content/full/18/1/637

cytoskeletal linkages at sarcolemma
Cytoskeletal Linkages at Sarcolemma

http://cellbio.annualreviews.org/cgi/content/full/18/1/637

muscle proteins and diseases
Muscle Proteins and Diseases

http://www-ermm.cbcu.cam.ac.uk/0200488Xh.htm

creatine kinase and cki

contents spill out

detected in blood

Creatine Kinase and CKI
  • CKtotal 0-220 U/L CKMB 0-5 ng/mL
  • Muscle damage CKtotal elevated
  • CK index (CKI) = CKMB/CKtotal (x 100 for %)
  • Skeletal origin CKI <0.03 (3%) or use CKMM
  • Cardiac origin CKI >0.06 (6%)
  • If 0.03 (3%) > CKI <0.05 (5%) follow with Tn biomarkers

Cell damage

in situ dystrophin
In Situ Dystrophin
  • Normal dystrophin staining around the rim of muscle fibers

Absent dystrophin: Duchenne muscular dystrophy

  • Left: No staining around the rim of muscle fibers.
  • Right: No staining of most muscle fibers. One "revertant" fiber with dystrophin staining.

http://www.neuro.wustl.edu/neuromuscular/pathol/dmdpath.htm

western blots
Western Blots
  • Lane 3: Normal; Dystrophin has normal size and amount.
  • Lane 1: Becker dystrophy; Dystrophin has reduced abundance but normal size.
  • Lane 2: Becker dystrophy; Dystrophin has reduced size and abundance.
  • Lane 4: Duchenne dystrophy; Almost no protein is present.
  • Lane 5: Duchenne outlier; Dystrophin has severely reduced abundance.

Western blot of dystrophin from dystrophinopathies.

http://www.neuro.wustl.edu/neuromuscular/pathol/dmdpath.htm

review questions
Review Questions
  • What is dystrophin (domains, characteristics)?
  • Where is dystrophin found (interactions, role)?
  • How do you assess muscle damage?
  • How do you assess dystrophin levels?