Cloning and characterization of Cbl-associated protein splicing isoforms.
Cbl-associated protein (CAP) is an adaptor protein that plays important roles in both signal transduction and cytoskeleton rearrangement. Alternative splicing of the gene SORBS1 results in multiple isoforms of CAP protein. We report here the cloning of 3 new CAP isoforms, CAP2, CAP3, and CAP4, from mouse adipose tissue. RT-PCR analyses reveal that the isoform mRNAs are differentially expressed. CAP2, CAP3, and CAP4 contain a coiled-coil domain. In addition, CAP4 contains a proline-rich region, part of which exists in CAP3. Coimmunoprecipitation experiments show that CAP4 forms a homodimeric complex. While these new isoforms similarly interact with Cbl, they exhibit varied binding specificity toward vinculin. In contrast to CAP1 and CAP2, CAP4 does not interact with vinculin, and CAP3 binds with low affinity. Immunofluorescence analysis demonstrates differential subcellular localization of Myc-tagged CAP isoforms in 3T3-L1 adipocytes. These results suggest that these new isoforms of CAP might play different signaling roles.
Zhang
Mei
M
Department of Internal Medicine and Physiology, Life Sciences Institute, University of Michigan Medical Center, Ann Arbor, MI 48109, USA.
Kimura
Akiko
A
Saltiel
Alan R
AR
D016475:3T3 Cells; D017667:Adipocytes / Q000378:metabolism*; D017398:Alternative Splicing; D000595:Amino Acid Sequence; D000818:Animals; D019556:COS Cells; D002522:Chlorocebus aethiops; D003001:Cloning, Molecular; D003598:Cytoskeletal Proteins / Q000235:genetics* / Q000378:metabolism; D017931:DNA Primers / Q000737:chemistry; D019281:Dimerization; D005455:Fluorescent Antibody Technique; D051379:Mice; D008969:Molecular Sequence Data; D020033:Protein Isoforms; D012333:RNA, Messenger / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017386:Sequence Homology, Amino Acid; D015398:Signal Transduction; D013347:Subcellular Fractions; D016596:Vinculin / Q000378:metabolism
D003598:Cytoskeletal Proteins; D017931:DNA Primers; D020033:Protein Isoforms; D012333:RNA, Messenger; D016596:Vinculin