Characterization of proteins purified from a beta(1) integrin affinity column: A novel ecm molecule and a muscle-specific integrin alpha subunit

Bao, Zhengzheng

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https://hdl.handle.net/2142/20539 Copy

Description

Title

Characterization of proteins purified from a beta(1) integrin affinity column: A novel ecm molecule and a muscle-specific integrin alpha subunit

Author(s)

Bao, Zhengzheng

Issue Date

1993

Doctoral Committee Chair(s)

Horwitz, Alan F.

Department of Study

Biochemistry

Discipline

Biochemistry

Degree Granting Institution

University of Illinois at Urbana-Champaign

Degree Name

Ph.D.

Degree Level

Dissertation

Keyword(s)

• Biology, Cell
• Chemistry, Analytical

Language

eng

Abstract

• Previous work on the distribution of the $\beta$1 integrins on skeletal muscle revealed that integrin occupied all of the major junctional regions on muscle. It also showed interesting, anomalous proteins on SDS-PAGE from $\beta$1 affinity purifications from muscle tissue. The possibility that these anomalous proteins represented novel integrin-associated proteins or even new $\alpha$ subunits motivated this work which was to characterize two of the major proteins with the molecular weights at 220 kD and 70 kD, respectively.
• "The first protein (220 kD) we isolated and characterized is a novel extracellular matrix molecule, as determined by its subunit composition, N-terminal amino acid sequence, and in vivo localization. It is abundant in many adult tissues. It has the unusual property that it is expressed initially very late in development. However, the protein does not bind to integrin directly as expected. The co-purification with integrin is because it binds to the carbohydrate support in Sepharose. Hemagglutination assays with mono and disaccharides show that it functions as a lectin with galactoside-binding specificity. This protein is also found to bind specifically to laminin, but does not bind to fibronectin or type IV collagen. We call this novel protein ""LBL"" for laminin binding lectin."
• The 70 kD protein which is also present in the $\beta$1 integrin affinity purification from skeletal muscle, was characterized next. Immunization against this protein has resulted in an antibody directed against the avian $\alpha\sb7$ integrin subunit. The specificity of the antibody was established by patterns of tissue staining and cross reactivity with antibodies against the rat $\alpha\sb7$ cytoplasmic domain. On sections of adult skeletal muscle the $\alpha7$ integrin was enriched in the myotendinous junction (MTJ). This localization was unique as neither the $\alpha\sb1$, $\alpha\sb3$, $\alpha\sb5$, $\alpha\sb6$, nor $\alpha\sb{\rm v}$ subunit localizes in the MTJ. The expression of the $\alpha\sb7$ subunit in the developing MTJ is first apparent around embryo day 14. The time of appearance of the $\alpha\sb7$ in the MTJ correlates with the insertion of myofibrils into subsarcolemmal densities and folding of the junctional membrane, suggesting a role for the $\alpha\sb7$ integrin in this process. The presence of the $\alpha\sb7$ in the MTJ and the $\alpha\sb5$ in the adhesion plaque demonstrates a molecular difference between these two adherens junctions. It also points to possible origins of junctional specificity on muscle. Differences between these two junctions were developed further using an antibody against phosphotyrosine. The focal adhesion and NMJ, but not the MTJ contained proteins phosphorylated on tyrosine.

Type of Resource

text

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http://hdl.handle.net/2142/20539

Copyright and License Information

Copyright 1993 Bao, Zhengzheng

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