{"id":{"repo_id":"lmu-germany","oai_identifier":"oai:edoc.ub.uni-muenchen.de:2168"},"canonical_url":"https://search.dev.ndltd.org/etd/lmu-germany/oai:edoc.ub.uni-muenchen.de:2168","repository":{"repo_id":"lmu-germany","name":"Ludwig Maxmilians Universität München","base_url":"https://edoc.ub.uni-muenchen.de/cgi/oai2"},"display":{"title":"RBP-J dependent and independent signalling of EBNA-2","abstract":"EBNA-2 is a multifunctional viral oncogene involved in the immortalisation of B-cells by EBV. EBNA-2 regulates transcription of viral and cellular genes in the proliferative phase of the viral life cycle, which in vitro results in the outgrowth of EBV positive B-cells into lymphoblastoid cell lines (LCLs). EBNA-2 transcriptional signalling is mediated by cellular DNA-binding proteins, such as RBP-J and PU.1, since EBNA-2 does not contain its own DNA-binding domain. In order to better characterise EBNA-2 signalling we conducted a mutational analysis of the viral LMP-1 promoter that is strongly induced by EBNA-2 in the EBV-immortalised B-cells. Our mutational analysis of the LMP-1 promoter confirmed that the PU.1 binding site is important for transactivation of the LMP-1 promoter by EBNA-2, whereas RBP-J binding to the LMP-1 promoter leads to repression and EBNA-2 binding to RBP-J is not required for transactivation. These results imply that EBNA-2 transactivates the LMP-1 promoter preferentially by an RBP-J independent mechanism. We further characterised EBNA-2 signalling by dissection of promoter targeting domains in the EBNA-2 protein. Two EBNA-2 mutants, the CR4del and WW mutant, preferentially activated RBP-J dependent and independent signalling indicating that EBNA-2 uses at least two separate signalling pathways. We introduced the characterised EBNA-2 mutants into the EBV genome and produced recombinant viruses carrying specific mutations in the EBNA-2 genes. Primary B-cells were infected with increasing titres of recombinant EBVs lacking the EBNA-2 ORF or carrying the WW or CR4del mutant. Viruses lacking the EBNA-2 ORF or carrying the WW mutant were not able to immortalise primary B-cells even at high viral titres. The CR4 region of EBNA-2 strongly influenced B-cell immortalisation efficiency and growth rate of the immortalised B-cells. These results indicate that EBNA-2 and the RBP-J signalling of EBNA-2 are absolutely essential for B-cell immortalisation by EBV. In contrast, the CR4 EBNA-2 region mediating RBP-J independent signalling is critical, but not absolutely essential for the process of EBV immortalisation.","abstract_html":"EBNA-2 is a multifunctional viral oncogene involved in the immortalisation of B-cells by EBV. EBNA-2 regulates transcription of viral and cellular genes in the proliferative phase of the viral life cycle, which in vitro results in the outgrowth of EBV positive B-cells into lymphoblastoid cell lines (LCLs). EBNA-2 transcriptional signalling is mediated by cellular DNA-binding proteins, such as RBP-J and PU.1, since EBNA-2 does not contain its own DNA-binding domain. In order to better characterise EBNA-2 signalling we conducted a mutational analysis of the viral LMP-1 promoter that is strongly induced by EBNA-2 in the EBV-immortalised B-cells. Our mutational analysis of the LMP-1 promoter confirmed that the PU.1 binding site is important for transactivation of the LMP-1 promoter by EBNA-2, whereas RBP-J binding to the LMP-1 promoter leads to repression and EBNA-2 binding to RBP-J is not required for transactivation. These results imply that EBNA-2 transactivates the LMP-1 promoter preferentially by an RBP-J independent mechanism. We further characterised EBNA-2 signalling by dissection of promoter targeting domains in the EBNA-2 protein. Two EBNA-2 mutants, the CR4del and WW mutant, preferentially activated RBP-J dependent and independent signalling indicating that EBNA-2 uses at least two separate signalling pathways. We introduced the characterised EBNA-2 mutants into the EBV genome and produced recombinant viruses carrying specific mutations in the EBNA-2 genes. Primary B-cells were infected with increasing titres of recombinant EBVs lacking the EBNA-2 ORF or carrying the WW or CR4del mutant. Viruses lacking the EBNA-2 ORF or carrying the WW mutant were not able to immortalise primary B-cells even at high viral titres. The CR4 region of EBNA-2 strongly influenced B-cell immortalisation efficiency and growth rate of the immortalised B-cells. These results indicate that EBNA-2 and the RBP-J signalling of EBNA-2 are absolutely essential for B-cell immortalisation by EBV. In contrast, the CR4 EBNA-2 region mediating RBP-J independent signalling is critical, but not absolutely essential for the process of EBV immortalisation.","abstract_has_math":false,"creators":["Grabusic, Kristina"],"institution":"Ludwig-Maximilians-Universität","degree_name":null,"degree_level":"thesis.doctoral","degree_discipline":null,"degree_department":null,"school":null,"contributors":[],"advisors":[],"committee_chairs":[],"committee_members":[],"year":2004,"date_issued":"2004-05-10","date_published":"2004-05-10","updated_at":"2026-07-24T02:51:56Z","subjects":[],"languages":[],"rights":[],"rights_urls":[],"identifier_entries":[]},"links":{"outbound_url":"https://edoc.ub.uni-muenchen.de/2168/","outbound_label":"Repository record","outbound_source":"source_url"},"metadata_groups":[{"id":"people","label":"People","entries":[{"key":"dc:creator","label":"Author","values":["Grabusic, Kristina"]}]},{"id":"academic_context","label":"Academic Context","entries":[{"key":"dc:publisher","label":"Institution","values":["Universitätsbibliothek der Ludwig-Maximilians-Universität"]},{"key":"dc:type","label":"Dc Type","values":["doctoralThesis"]},{"key":"thesis:degree_level","label":"Degree Level","values":["thesis.doctoral"]},{"key":"thesis:institution_name","label":"Thesis Institution Name","values":["Ludwig-Maximilians-Universität"]}]},{"id":"additional","label":"Additional Metadata","entries":[{"key":"dc:description.abstract","label":"Abstract","values":["EBNA-2 is a multifunctional viral oncogene involved in the immortalisation of B-cells by EBV. EBNA-2 regulates transcription of viral and cellular genes in the proliferative phase of the viral life cycle, which in vitro results in the outgrowth of EBV positive B-cells into lymphoblastoid cell lines (LCLs). EBNA-2 transcriptional signalling is mediated by cellular DNA-binding proteins, such as RBP-J and PU.1, since EBNA-2 does not contain its own DNA-binding domain. In order to better characterise EBNA-2 signalling we conducted a mutational analysis of the viral LMP-1 promoter that is strongly induced by EBNA-2 in the EBV-immortalised B-cells. Our mutational analysis of the LMP-1 promoter confirmed that the PU.1 binding site is important for transactivation of the LMP-1 promoter by EBNA-2, whereas RBP-J binding to the LMP-1 promoter leads to repression and EBNA-2 binding to RBP-J is not required for transactivation. These results imply that EBNA-2 transactivates the LMP-1 promoter preferentially by an RBP-J independent mechanism. We further characterised EBNA-2 signalling by dissection of promoter targeting domains in the EBNA-2 protein. Two EBNA-2 mutants, the CR4del and WW mutant, preferentially activated RBP-J dependent and independent signalling indicating that EBNA-2 uses at least two separate signalling pathways. We introduced the characterised EBNA-2 mutants into the EBV genome and produced recombinant viruses carrying specific mutations in the EBNA-2 genes. Primary B-cells were infected with increasing titres of recombinant EBVs lacking the EBNA-2 ORF or carrying the WW or CR4del mutant. Viruses lacking the EBNA-2 ORF or carrying the WW mutant were not able to immortalise primary B-cells even at high viral titres. The CR4 region of EBNA-2 strongly influenced B-cell immortalisation efficiency and growth rate of the immortalised B-cells. These results indicate that EBNA-2 and the RBP-J signalling of EBNA-2 are absolutely essential for B-cell immortalisation by EBV. In contrast, the CR4 EBNA-2 region mediating RBP-J independent signalling is critical, but not absolutely essential for the process of EBV immortalisation."]},{"key":"dc:format.medium","label":"Dc Format Medium","values":["application/pdf"]},{"key":"dc:title","label":"Title","values":["RBP-J dependent and independent signalling of EBNA-2"]}]}],"canonical_facts":{"dc:creator":["Grabusic, Kristina"],"dc:description.abstract":["EBNA-2 is a multifunctional viral oncogene involved in the immortalisation of B-cells by EBV. EBNA-2 regulates transcription of viral and cellular genes in the proliferative phase of the viral life cycle, which in vitro results in the outgrowth of EBV positive B-cells into lymphoblastoid cell lines (LCLs). EBNA-2 transcriptional signalling is mediated by cellular DNA-binding proteins, such as RBP-J and PU.1, since EBNA-2 does not contain its own DNA-binding domain. In order to better characterise EBNA-2 signalling we conducted a mutational analysis of the viral LMP-1 promoter that is strongly induced by EBNA-2 in the EBV-immortalised B-cells. Our mutational analysis of the LMP-1 promoter confirmed that the PU.1 binding site is important for transactivation of the LMP-1 promoter by EBNA-2, whereas RBP-J binding to the LMP-1 promoter leads to repression and EBNA-2 binding to RBP-J is not required for transactivation. These results imply that EBNA-2 transactivates the LMP-1 promoter preferentially by an RBP-J independent mechanism. We further characterised EBNA-2 signalling by dissection of promoter targeting domains in the EBNA-2 protein. Two EBNA-2 mutants, the CR4del and WW mutant, preferentially activated RBP-J dependent and independent signalling indicating that EBNA-2 uses at least two separate signalling pathways. We introduced the characterised EBNA-2 mutants into the EBV genome and produced recombinant viruses carrying specific mutations in the EBNA-2 genes. Primary B-cells were infected with increasing titres of recombinant EBVs lacking the EBNA-2 ORF or carrying the WW or CR4del mutant. Viruses lacking the EBNA-2 ORF or carrying the WW mutant were not able to immortalise primary B-cells even at high viral titres. The CR4 region of EBNA-2 strongly influenced B-cell immortalisation efficiency and growth rate of the immortalised B-cells. These results indicate that EBNA-2 and the RBP-J signalling of EBNA-2 are absolutely essential for B-cell immortalisation by EBV. In contrast, the CR4 EBNA-2 region mediating RBP-J independent signalling is critical, but not absolutely essential for the process of EBV immortalisation."],"dc:format.medium":["application/pdf"],"dc:publisher":["Universitätsbibliothek der Ludwig-Maximilians-Universität"],"dc:title":["RBP-J dependent and independent signalling of EBNA-2"],"dc:type":["doctoralThesis"],"thesis:degree_level":["thesis.doctoral"],"thesis:institution_name":["Ludwig-Maximilians-Universität"]},"updated_at":"2026-07-24T02:51:56Z"}