Attenuation of 19-9 antigen secretion in human colorectal carcinoma cell cultures by transfection with cDNA encoding novel ADP-ribosylation factor-like proteins.
|
| Abstract |
We have used cDNAs coding for novel ADP-ribosylation factor-like molecules (ARL184 and ARL184Delta) to alter 19-9 antigen glycoprotein secretion in cultured human colorectal carcinoma cells SW1116 by transfection and cloning. This ARL contains a lipophilic N-terminal with an isoleucyl and 3 leucyl residues, 4 functioning consensus sequence GTP binding sites, and 184 total aminoacyl residues. An ARL cDNA was also constructed deleting the codon for the N-terminal glycyl moiety. The resulting cell clones were shown by Northern blots to overexpress ARL mRNA. Electron microscopy-immunocytochemistry also indicated the overexpression of ARL granules subcellularly. Secretion of the tumor-associated 19-9 antigen into apical medium was decreased 3- to 5-fold and the secretion of TCA/PTA precipitable 3H-labeled glycoprotein was decreased by 34% in clone SW1116(ARL184)Delta. Western blot analyses of cell homogenates and media were in agreement with the secretion assays and showed a diminution of 170-200 kDa, 19-9, antigenicity in transfected cells and their media. Apical secretion of 19-9 antigen was diminished 14-fold in cells, SW1116 (ARL184)alpha, transfected with the complete ARL cDNA sequence, suggesting that the glycyl moiety may be required for maximal abatement. However, incorporation of label from [3H]myristate into 22-kDa bands of NP-40 extracts and ARL-antigenic molecules of parent cells was 3-fold greater than that in samples from the two transfectants; thus the transfected cells may not myristylate the overexpressed ARL efficiently. Notwithstanding the N-terminal glycyl moiety undergoing some other modification, we conclude that overexpression of this ARL is sufficient to generate a 19-9-deficient phenotype. These ARLs may eventually disrupt terminal oligosaccharide glycosylation, resulting in an apparent diminished exocytosis of 19-9 glycoprotein carriers by transfected and cloned cells.
|
|---|---|
| Authors | D Eboue, C Icard-Liepkalns, T M Beringer, V A Liepkalns |
| Journal | Archives of biochemistry and biophysics (Arch Biochem Biophys) Vol. 350 Issue 2 Pg. 145-56 (Feb 15 1998) ISSN: 0003-9861 [Print] United States |
| PMID | 9473288 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't) |
| Copyright | Copyright 1998 Academic Press. |
| Chemical References |
|
| Topics |
|
Join CureHunter, for free Research Interface BASIC access!
Realize the full power of the drug-disease research graph!