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Published on: March 30, 2018
Characterization of DP103, a novel DEAD box protein that binds to the Epstein-Barr virus nuclear proteins EBNA2 and
A T Grundhoff1, E Kremmer, O Türeci
1Institut für Medizinische Mikrobiologie und Hygiene, Abteilung Virologie, Universitätskliniken des Saarlandes, Germany.
Abstract:
The Epstein-Barr virus-encoded nuclear antigens EBNA2 and EBNA3C both interact with the cellular transcription factor RBP-Jkappa and modulate the expression of several shared target genes, suggesting a tight cooperation in latently infected cells. In a survey for additional cellular factors that bind to EBNA2 as well as EBNA3C, we have isolated and characterized DP103, a novel human member of the DEAD box family of putative ATP-dependent RNA helicases. The interaction with DP103 is mediated by amino acids (aa) 121-213 of EBNA2 and aa 534-778 of EBNA3C, regions that are not involved in binding of the viral proteins to RBP-Jkappa. The DP103-cDNA encodes a protein of 824 aa that harbors all of the common DEAD box motifs. Monoclonal antibodies raised against DP103 detect a protein of 103 kDa in mammalian cells that resides in high molecular weight complexes in vivo. We have detected an ATPase activity intrinsic to or closely associated with DP103. By subcellular fractionation, we find DP103 in both a soluble nuclear fraction as well as in the insoluble skeletal fraction. Whereas the protein and its mRNA are uniformly expressed in all tested cell lines, we observed differential expression of the mRNA in normal human tissues.
Insights
Researchers identified DP103, a novel human DEAD box protein, that binds to Epstein-Barr virus nuclear antigens EBNA2 and EBNA3C. This discovery sheds light on viral protein interactions and cellular factor involvement in infected cells.
Area of Science:
- Molecular Biology
- Virology
- Cell Biology
Background:
- Epstein-Barr virus (EBV)-encoded nuclear antigens EBNA2 and EBNA3C interact with RBP-Jkappa, influencing target gene expression in infected cells.
- Investigating cellular factors interacting with both EBNA2 and EBNA3C is crucial for understanding EBV latency.
Purpose of the Study:
- To identify and characterize novel cellular factors that bind to EBV nuclear antigens EBNA2 and EBNA3C.
- To elucidate the interaction domains and functional properties of the newly identified factor.
Main Methods:
- Protein-protein interaction assays to identify binding partners of EBNA2 and EBNA3C.
- Molecular cloning and characterization of the novel human DEAD box protein, DP103.
- Antibody generation and Western blot analysis to detect DP103 protein.
- Biochemical assays to determine ATPase activity.
- Subcellular fractionation to localize DP103 within the cell.
Main Results:
- DP103, a novel human DEAD box protein, was identified as a binding partner for both EBNA2 and EBNA3C.
- Specific interaction domains were mapped: EBNA2 aa 121-213 and EBNA3C aa 534-778.
- DP103 encodes an 824 amino acid protein with conserved DEAD box motifs and exhibits ATPase activity.
- DP103 protein (103 kDa) exists in high molecular weight complexes and is found in both soluble nuclear and insoluble skeletal fractions.
- DP103 mRNA and protein expression are ubiquitous in cell lines, but mRNA shows differential expression in human tissues.
Conclusions:
- DP103 is a novel cellular factor interacting with EBV proteins EBNA2 and EBNA3C, independent of their RBP-Jkappa binding sites.
- DP103's characteristics suggest a role as an ATP-dependent RNA helicase involved in cellular processes potentially exploited by EBV.
- The differential tissue expression of DP103 warrants further investigation into its specific biological functions and relevance in EBV infection.
