Heterogeneous RNA-binding Protein M4 Is a Receptor for Carcinoembryonic Antigen in Kupffer Cells

Abstract

Here we report the isolation of the recombinant cDNA clone from rat macrophages, Kupffer cells (KC) that encodes a protein interacting with carcinoembryonic antigen (CEA). To isolate and identify the CEA receptor gene we used two approaches: screening of a KC cDNA library with a specific antibody and the yeast two-hybrid system for protein interaction using as a bait the N-terminal part of the CEA encoding the binding site. The liver is a common site for metastasis from various forms of primary malignancies. Both experimental and clinical results reveal that the presence of carcinoembryonic antigen (CEA) 1 enhances liver metastasis from colorectal carcinoma cells (1, 2). CEA is a highly characterized, cell surface glycoprotein overexpressed by various tumor cells and provides a tool for tumor tissue-specific targeting. Increasing amounts of CEA in the serum correlates with the development of metastatic recurrence after surgical removal of the primary tumor (3). Earlier we have shown that CEA production of human colorectal cancer cell lines directly correlates with the metastatic potential (4). Poorly metastatic colon cancer cell lines become highly metastatic when transfected with the cDNA coding for CEA (5, 6). As a member of the immunoglobulin supergene family, CEA is involved in intercellular recognition and may facilitate attachment of colorectal carcinoma cells to sites of metastasis. In an experimental metastasis model of colorectal carcinoma in athymic nude mice, systemic injection of CEA enhanced experimental liver metastasis and implantation in liver by weakly metastatic tumor cells (7,8).To successfully treat cancers, it is necessary to prevent the development of metastasis after the treatment or removal of the primary malignancy. Therefore, it is important to elucidate the mechanism by which CEA enhances metastatic potential. The molecular basis by which CEA can influence metastasis is only partly understood. We have shown that CEA is rapidly cleared from the circulation of experimental animals, accumulates in the liver, and is endocytosed in vitro by Kupffer cells. This initiates a series of signaling events that leads to tyrosine phosphorylation on at least two intracellular proteins (9) and is followed by induction of IL-1␣, IL-6, IL-10, and tumor necrosis factor-␣ cytokines (10). CEA uptake by Kupffer cells is independent of its carbohydrate composition and is mediated by an 80-kDa binding protein (11). CEA is recognized by this binding protein through a 5-amino acid sequence, Pro-Glu-Leu-Pro-Lys (PELPK), located at the hinge region (amino acids 108 -112) between the N-terminal and the first immunoglobulin loop domain (12). Molecular modeling studies have suggested that this region is exposed on the surface of the molecule. 2 We have recently shown in a subset of colorectal cancer patients that mutations in the PELPK region of CEA results in the accumulation of large amounts of CEA in the serum (13). To further study the interaction between the peptide sequence, PELPK, and rat Kupffer cells, the...

The Role of the Carcinoembryonic Antigen Receptor in Colorectal Cancer Progression

“…СЕА production occurs in many types of epithelial cancers and can be observed in 9-15% of testicular, ovarian, pancreatic, gastric and thyroid cancers, as well as in 50-90% metastasizing colorectal and breast cancers [10,31]. To study metastasis it is especially important to have an adequate biological model to elucidate the genes that are involved in metastasis in vitro and in vivo.…”

“…HnRNPM is a very abundant nuclear shuttling protein that has at least 4 protein isoforms [10]. However, only 2 mRNA transcripts have been experimentally validated: the full-length protein (isoform 1) and the short isoform 2 that has a 39 amino acid deletion between the RNA binding domains 1 and 2 [10]. We initially identified isoform 2 as a CEA-binding protein in Kupffer cells though both isoforms are capable of binding CEA.…”

“…We identified a novel CEA binding protein (CEAR) on liver macrophages, (Kupffer cells), who's activity alters the liver microenvironment such that implantation and survival of tumor cells increases [9]. Previously this protein was identified as an isoform of the hnRNPM protein [10]. HnRNPM belongs to a large family of 20 heterogeneous nuclear RNA-binding proteins (hnRNPs A-U), also called "the histones of RNA" [11].…”

“…RBD3 overlaps with the domain that binds CEA [16]. HnRNPM is a very abundant nuclear shuttling protein that has at least 4 protein isoforms [10]. However, only 2 mRNA transcripts have been experimentally validated: the full-length protein (isoform 1) and the short isoform 2 that has a 39 amino acid deletion between the RNA binding domains 1 and 2 [10].…”

“…We initially identified isoform 2 as a CEA-binding protein in Kupffer cells though both isoforms are capable of binding CEA. [10]. HnRNPM is a multifunctional protein that is involved in mRNA processing [17], splicing [18], stress response [19], mRNA transport and stability of exosomes [20].…”