Bovine aortic endothelial cells express a variant of the very low density lipoprotein receptor that lacks the O-linked sugar domain.
The very low density lipoprotein (VLDL) receptor is a member of the low density lipoprotein supergene family of receptors in which differential splicing of mRNA has been reported. We present several lines of evidence showing that bovine aortic endothelial cells exclusively express a VLDL receptor isoform that lacks the O-linked sugar domain i) Western and receptor-associated protein (RAP) ligand blotting gave a single band of about 99 kDa in membrane extracts of bovine aortic endothelial cells (BAEC). ii) Screening of the BAEC cDNA library with the previously characterized human VLDL receptor cDNA as a probe gave several C-terminal-positive clones; all lacked the 84 nucleotides corresponding to exon 16. Polymerase chain reaction (PCR) confirmed that VLDL receptor cDNA encoding exon 16 was absent from the library. iii) Reverse transcription (RT)-PCR analysis of the BAEC mRNA using a pair of oligonucleotide primers that flank the deletion gave only one band of 136 nt. iv) Semiquantitative RT-PCR analysis showed that only the non-O-glycosylated variant was expressed in BAEC. Cell-binding studies with antibodies against the N-terminal domain showed that the BAEC VLDL receptor is present at the plasma membrane, suggesting that the non-glycosylated variant could be functional. In addition, RT-PCR performed in bovine tissues showed that the variant containing the O-linked sugar domain is preferentially expressed in heart, brain, and skeletal muscle, whereas the non-O-glycosylated spliced variant is found in all tissues analyzed. Taken together these results suggest that the differential splicing of the VLDL receptor is cell- and tissue-specific and that the functions of the receptor could depend on the cell type.
Magrané
J
J
Department of Cellular Biology, Faculty of Biology, University of Barcelona, Avda. Diagonal, 645, E-08028 Barcelona, Spain.
Reina
M
M
Pagan
R
R
Luna
A
A
Casaroli-Marano
R P
RP
Angelin
B
B
Gåfvels
M
M
Vilaró
S
S
D017398:Alternative Splicing; D000595:Amino Acid Sequence; D000818:Animals; D001011:Aorta; D001483:Base Sequence; D015153:Blotting, Western; D019556:COS Cells; D002241:Carbohydrates / Q000737:chemistry*; D002417:Cattle; D002478:Cells, Cultured; D018076:DNA, Complementary / Q000737:chemistry / Q000302:isolation & purification; D004730:Endothelium, Vascular / Q000378:metabolism*; D006801:Humans; D008024:Ligands; D008856:Microscopy, Fluorescence; D008969:Molecular Sequence Data; D011973:Receptors, LDL / Q000096:biosynthesis / Q000737:chemistry* / Q000235:genetics; D013045:Species Specificity; D014162:Transfection
D002241:Carbohydrates; D018076:DNA, Complementary; D008024:Ligands; D011973:Receptors, LDL; C036053:VLDL receptor