Identification of a Candidate Human Spectrin Src Homology 3 Domain-binding Protein Suggests a General Mechanism of Association of Tyrosine Kinases with the Spectrin-based Membrane Skeleton

• Published in: The Journal of biological chemistry 1998-05, Vol.273 (22), p.13681-13692
• Main Authors: Ziemnicka-Kotula, Dorota, Xu, Jiliu, Gu, Hong, Potempska, Anna, Kim, Kwang Soo, Jenkins, Edmund C., Trenkner, Ekkhart, Kotula, Leszek
• Format: Article
• Language: English
• Subjects: 3T3 Cells; Adaptor Proteins, Signal Transducing; Amino Acid Sequence; Animals; Base Sequence; Binding Sites; Carrier Proteins - chemistry; Carrier Proteins - genetics; Carrier Proteins - metabolism; Cell Membrane - enzymology; Chromosome Mapping; Chromosomes, Human, Pair 10; Cytoskeletal Proteins; DNA, Complementary; Humans; Mice; Molecular Sequence Data; Protein-Tyrosine Kinases - metabolism; Recombinant Fusion Proteins - chemistry; Recombinant Fusion Proteins - genetics; Recombinant Fusion Proteins - metabolism; Saccharomyces cerevisiae - genetics; Sequence Homology, Amino Acid; Spectrin - chemistry; Spectrin - genetics; Spectrin - metabolism; src Homology Domains
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.273.22.13681

Spectrin is a widely expressed protein with specific isoforms found in erythroid and nonerythroid cells. Spectrin contains an Src homology 3 (SH3) domain of unknown function. A cDNA encoding a candidate spectrin SH3 domain-binding protein was identified by interaction screening of a human brain expression library using the human erythroid spectrin (αI) SH3 domain as a bait. Five isoforms of the αI SH3 domain-binding protein mRNA were identified in human brain. Mapping of SH3 binding regions revealed the presence of two αI SH3 domain binding regions and one Abl-SH3 domain binding region. The gene encoding the candidate spectrin SH3 domain-binding protein has been located to human chromosome 10p11.2 → p12. The gene belongs to a recently identified family of tyrosine kinase-binding proteins, and one of its isoforms is identical to e3B1, an eps8-binding protein (Biesova, Z., Piccoli, C., and Wong, W. T. (1997)Oncogene14, 233–241). Overexpression of the green fluorescent protein fusion of the SH3 domain-binding protein in NIH3T3 cells resulted in cytoplasmic punctate fluorescence characteristic of the reticulovesicular system. This fluorescence pattern was similar to that obtained with the anti-human erythroid spectrin αIΣI/βIΣI antibody in untransfected NIH3T3 cells; in addition, the anti-αIΣI/βIΣI antibody also stained Golgi apparatus. Immunofluorescence obtained using antibodies against αIΣI/βIΣI spectrin and Abl tyrosine kinase but not against αII/βII spectrin colocalized with the overexpressed green fluorescent protein-SH3-binding protein. Based on the conservation of the spectrin SH3 binding site within members of this protein family and published interactions, a general mechanism of interactions of tyrosine kinases with the spectrin-based membrane skeleton is proposed.