The 67-kDa enzymatically inactive alternatively spliced variant of beta-galactosidase is identical to the elastin/laminin-binding protein.
Our previous studies showed immunological and functional similarities, as well as partial sequence homology, between the enzymatically inactive alternatively spliced variant of human beta-galactosidase (S-gal) and the 67-kDa elastin/laminin-binding protein (EBP) from sheep. To define the genetic origin of the EBP further, a full-length human S-gal cDNA clone was constructed and subjected to in vitro transcription/translation. The cDNA was also transfected into COS-1 cells and into the EBP-deficient smooth muscle cells (SMC) from sheep ductus arteriosus (DA). In vitro translation yielded an unglycosylated form of the S-gal protein, which immunoreacted with anti-beta-galactosidase antibodies and bound to elastin and laminin affinity columns. S-gal cDNA transfections into COS-1 and DA SMC increased expression of a 67-kDa protein that immunolocalized intracellularly and to the cell surface and, when extracted from the cells, bound to elastin. The S-gal-transfected cells displayed increased adherence to elastin-covered dishes, consistent with the cell surface distribution of the newly produced S-gal-encoded protein. Transfection of DA SMC additionally corrected their impaired elastic fiber assembly. These results conclusively identify the 67-kDa splice variant of beta-galactosidase as EBP.
Privitera
S
S
Division of Cardiovascular Research, The Hospital for Sick Children, Toronto, Ontario M5G 1X8, Canada.
Prody
C A
CA
Callahan
J W
JW
Hinek
A
A
D017398:Alternative Splicing; D000595:Amino Acid Sequence; D000818:Animals; D019556:COS Cells; D002448:Cell Adhesion; D002478:Cells, Cultured; D018076:DNA, Complementary / Q000235:genetics; D004549:Elastin / Q000302:isolation & purification / Q000378:metabolism; D005353:Fibronectins / Q000302:isolation & purification; D006801:Humans; D007797:Laminin / Q000378:metabolism; D008969:Molecular Sequence Data; D009131:Muscle, Smooth, Vascular / Q000166:cytology; D011485:Protein Binding; D014176:Protein Biosynthesis; D011498:Protein Precursors; D011956:Receptors, Cell Surface / Q000235:genetics* / Q000378:metabolism; D017474:Receptors, Laminin / Q000235:genetics* / Q000378:metabolism; D012756:Sheep; D013045:Species Specificity; D014158:Transcription, Genetic; D001616:beta-Galactosidase / Q000235:genetics* / Q000378:metabolism
D018076:DNA, Complementary; D005353:Fibronectins; D007797:Laminin; D011498:Protein Precursors; D011956:Receptors, Cell Surface; D017474:Receptors, Laminin; C054901:elastin-binding proteins; D004549:Elastin; D001616:beta-Galactosidase