Analysis of factors determining replication of BK virus (BKV) episomal vectors in human cells showed that vector copy number was related to the level of BKV T antigen expression. T antigen was synthesized efficiently, as assessed by indirect immunofluorescence, in vector-transfected primary embryonic fibroblasts undergoing neoplastic transformation. Surprisingly, transfected continuous cell lines (143 B, HeLa and KB), kept under biochemical selection or tested in transient assays, produced negligible amounts or no T antigen, revealed only by a sensitive ELISA test, suggesting that in these cells vector amplification was under the control of cellular factors. Presence or absence of BKV late region sequences, BKV strain, orientation of the inserted genes and presence or absence of selection were not relevant for vector replication. Type of biochemical selection, however, was important, since BKV vectors containing the thymidine kinase gene replicated better than those containing the neo gene. Despite great variability, vector copy number increased in transfected clones of adenovirus 5-transformed 293 cells, in the absence of immunofluorescence detectable T antigen. These cells express adenovirus immediate early proteins E1A and E1B which may directly or indirectly activate BKV origin of replication.

Factors affecting amplification of BK virus episomal vectors in human cells. Brief report.

CAPUTO, ANTONELLA;
1988

Abstract

Analysis of factors determining replication of BK virus (BKV) episomal vectors in human cells showed that vector copy number was related to the level of BKV T antigen expression. T antigen was synthesized efficiently, as assessed by indirect immunofluorescence, in vector-transfected primary embryonic fibroblasts undergoing neoplastic transformation. Surprisingly, transfected continuous cell lines (143 B, HeLa and KB), kept under biochemical selection or tested in transient assays, produced negligible amounts or no T antigen, revealed only by a sensitive ELISA test, suggesting that in these cells vector amplification was under the control of cellular factors. Presence or absence of BKV late region sequences, BKV strain, orientation of the inserted genes and presence or absence of selection were not relevant for vector replication. Type of biochemical selection, however, was important, since BKV vectors containing the thymidine kinase gene replicated better than those containing the neo gene. Despite great variability, vector copy number increased in transfected clones of adenovirus 5-transformed 293 cells, in the absence of immunofluorescence detectable T antigen. These cells express adenovirus immediate early proteins E1A and E1B which may directly or indirectly activate BKV origin of replication.
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Utilizza questo identificativo per citare o creare un link a questo documento: https://hdl.handle.net/11577/2502179
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