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Heterogeneous RNA-binding protein M4 is a receptor for carcinoembryonic antigen in Kupffer cells
O V Bajenova1, R Zimmer, E Stolper
1Department of Surgery, Boston University School of Medicine, Boston, Massachusetts 02118, USA.
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. Both techniques resulted in the identification of the rat heterogeneous RNA-binding protein (hnRNP) M4 gene. The rat ortholog cDNA sequence has not been previously described. The open reading frame for this gene contains a 2351-base pair sequence with the polyadenylation signal AATAAA and a termination poly(A) tail. The mRNA shows ubiquitous tissue expression as a 2.4-kilobase transcript. The deduced amino acid sequence comprised a 78-kDa membrane protein with 3 putative RNA-binding domains, arginine/methionine/glutamine-rich C terminus and 3 potential membrane spanning regions. When hnRNP M4 protein is expressed in pGEX4T-3 vector system in Escherichia coli it binds (125)I-labeled CEA in a Ca(2+)-dependent fashion. Transfection of rat hnRNP M4 cDNA into a non-CEA binding mouse macrophage cell line p388D1 resulted in CEA binding. These data provide evidence for a new function of hnRNP M4 protein as a CEA-binding protein in Kupffer cells.
Insights
Researchers identified the rat heterogeneous nuclear ribonucleoprotein M4 (hnRNP M4) gene, a novel protein that binds to carcinoembryonic antigen (CEA). This discovery reveals a new function for hnRNP M4 in Kupffer cells, impacting CEA interactions.
Area of Science:
- Molecular Biology
- Immunology
- Cell Biology
Background:
- Carcinoembryonic antigen (CEA) is implicated in various cellular processes.
- Kupffer cells (KC) are macrophages in the liver with roles in immune surveillance.
- The specific receptor mediating CEA interaction with KC was previously unidentified.
Purpose of the Study:
- To isolate and identify the gene encoding the CEA receptor in rat Kupffer cells.
- To characterize the function and binding properties of the identified protein.
Main Methods:
- Screening of a rat Kupffer cell cDNA library using a specific antibody against CEA.
- Employing the yeast two-hybrid system to detect protein-protein interactions with CEA.
- Expressing the identified gene in Escherichia coli and transfecting it into mouse macrophage cell lines.
Main Results:
- Both screening methods identified the rat heterogeneous nuclear ribonucleoprotein M4 (hnRNP M4) gene.
- The deduced hnRNP M4 protein is a 78-kDa membrane protein with RNA-binding domains and membrane-spanning regions.
- Recombinant hnRNP M4 protein demonstrated calcium-dependent binding to CEA, and its expression in cells conferred CEA-binding ability.
Conclusions:
- The study identifies hnRNP M4 as a novel CEA-binding protein in Kupffer cells.
- This finding suggests a new role for hnRNP M4 in cellular interactions involving CEA.
- The characterized rat hnRNP M4 cDNA sequence provides a valuable resource for further research.