The cytoplasmic domain of C-CAM is required for C-CAM-mediated adhesion function: studies of a C-CAM transcript containing an unspliced intron

Abstract

Cell-CAM105 (also named C-CAM) is a cell surface glycoprotein involved in intercellular adhesion of rat hepatocytes. It has four extracellular immunoglobulin (Ig) domains, a transmembrane domain and a cytoplasmic domain and therefore is a member of the Ig supergene family. We have characterized multiple cDNAs of the C-CAM genes in rat intestine. Sequence analyses showed that rat intestine contained not only the previously reported L-form and S-form C-CAMs (renamed C-CAM1 and C-CAM2 respectively) but also a new isoform, C-CAM3. The C-CAM3 transcript codes for a polypeptide with a truncated C-terminus that lacks 65 amino acids from the previously reported C-CAM1 cytoplasmic domain. Unlike C-CAM1, C-CAM3 did not mediate cell adhesion when expressed in insect cells using the baculoviral expression system. Thus the extra 65 amino acids in the cytoplasmic domain of C-CAM1 are important for adhesion phenotype when expressed in insect cells. Although C-CAM1 and C-CAM2 are encoded by different genes, sequence analysis suggests that C-CAM3 is probably derived from alternative splicing of the C-CAM1 gene. To examine this possibility, we have determined the exon organization of the C-CAM1 gene. C-CAM3 differed from C-CAM1 by the presence of a single unspliced intron which contained a stop codon immediately after the regular splice junction. As a result, translation of C-CAM3 terminates at the point where C-CAM1 and C-CAM3 sequences diverge. To investigate the expression of C-CAM1, C-CAM2 and C-CAM3 in different tissues, we used an RNAase-protection assay to simultaneously assess the levels of expression of these transcripts. Using total RNA prepared from various tissues, we showed that expression of C-CAM3 was tissue-specific, and the C-CAM3 transcript accounted for about 25% of the transcripts derived from the C-CAM1 gene. However, further analysis revealed that C-CAM3 transcript was not present in cytosolic RNA, rather it was enriched in nuclear RNA prepared from hepatocytes. Although C-CAM3 cDNA contains the polyadenylation signal and is polyadenylated, these results indicate that C-CAM3 is probably an incomplete spliced product of C-CAM1 gene.

C-CAM1 expression: Differential effects on morphology, differentiation state and suppression of human PC-3 prostate carcinoma cells

“…Previous studies in myoblast culture systems (Rojas et al, 1996) and studies of C-CAM expression during liver development (Cheung et al, 1993b;Mowery and Hixson, 1991;Thompson et al, 1993) have shown that C-CAM (or BGP) accelerates di erentiation of myoblasts in vitro and is associated with the functional di erentiation of hepatocytes in the developing liver, respectively. However, C-CAM1 expression appeared to induce a less organized or di erentiated state in the PC-3 nude mouse tumors, evidenced by the decrease in number or absence or glandular/ductular and cord-like structures found in control PC3-V tumors.…”

“…C-CAM1, the rat homolog of human biliary glycoprotein I (BGPI), is a member of the carcinoembryonic antigen (CEA) family of Ig-like cell adhesion molecules (Cheung et al, 1993b;Culic et al, 1992). In rat liver, C-CAM1, also known as L-form, is the longer of the two major splice variants that vary in the length of their cytoplasmic domains (71 versus 10 amino acids).…”

Association of an 80 kDa protein with C-CAM1 cytoplasmic domain correlates with C-CAM1-mediated growth inhibition

“…The cytoplasmic domain of C-CAM1 contains 71 amino acids that are encoded by four exons, i.e., exons 6 ± 9 (Cheung et al, 1993a;Najjar et al, 1993). The membrane-proximal 6 amino acids together with the transmembrane domain is encoded by exon 6, while the rest of the cytoplasmic sequence was encoded by exons 7, 8 and 9 (each with 18, 10 and 35 amino acids, respectively).…”

Expression and androgen regulation of C-CAM cell adhesion molecule isoforms in rat dorsal and ventral prostate

“…While messages for both L-form and S-form C-CAM were detected in the DLP and the presence of C- (Cheung et al, 1993a) and the region used in the preparation of the antisense RNA probe. The restriction map and exon arrangement of the C-CAM1 genomic clone are shown.…”

“…L-and Sform C-CAM, the predominant isoforms in the rat, are generated from a single gene by alternative splicing (Cheung et al, 1993a;Najjar et al, 1993). L-form C-CAM has a 71-amino acid cytoplasmic domain.…”