Down-regulation of a 67-kDa YIGSR-binding protein upon differentiation of human neuroblastoma cells.
Differentiated human neuroblastoma LA-N1 cells that were exposed to dibutyryl adenosine 3',5'-cyclic monophosphate for 5 days (primed cells) showed increased adhesion to laminin-, fibronectin-, and collagen type I-coated plates as compared to unprimed cells. Moreover, primed cells seemed to adhere best to laminin. The binding site in laminin, mediating cell attachment, was identified as containing the YIGSR sequence, a known cell binding motif, located in the short arm of the B1 chain of laminin. The synthetic peptide amide, C(YIGSR)3-NH2, containing a repeat of this binding motif, inhibited the attachment of neuroblastoma cells to laminin in a competitive manner, and its inhibitory activity was inversely dependent on laminin concentrations. Affinity chromatography of membrane-extracted proteins over an Affi-Gel 10 column conjugated to C(YIGSR)3-NH2, revealed a major YIGSR-binding protein with an apparent molecular mass of 67 kDa. The 67-kDa surface membrane protein was specifically eluted from the column with the soluble C(YIGSR)3-NH2 peptide, but not with an unrelated peptide. Furthermore, no 67-kDa laminin-binding protein was recovered from an unrelated peptide matrix with the free C(YIGSR)3-NH2 peptide. Ligand blot overlay assays with biotin-labeled C(YIGSR)3-NH2 peptide demonstrated that the 67-kDa receptor is indeed a YIGSR-binding protein. This 67-kDa laminin-binding protein appeared to be down-regulated upon differentiation of LA-N1 cells, as indicated by the level of this protein and its mRNA.
Bushkin-Harav
I
I
Department of Neurobiology, Weizmann Institute of Science, Rehovot, Israel.
Garty
N B
NB
Littauer
U Z
UZ
D000595:Amino Acid Sequence; D002448:Cell Adhesion; D002454:Cell Differentiation; D002846:Chromatography, Affinity; D015536:Down-Regulation; D016326:Extracellular Matrix Proteins / Q000378:metabolism; D006801:Humans; D007150:Immunohistochemistry; D007797:Laminin / Q000378:metabolism*; D008969:Molecular Sequence Data; D009447:Neuroblastoma; D009693:Nucleic Acid Hybridization; D009842:Oligopeptides / Q000378:metabolism*; D011485:Protein Binding; D011506:Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D014407:Tumor Cells, Cultured
D016326:Extracellular Matrix Proteins; D007797:Laminin; D009842:Oligopeptides; D011506:Proteins; D012333:RNA, Messenger; C081571:tyrosyl-isoleucyl-glycyl-seryl-arginine