Related Experiment Videos
Transactivating potential of the 3' open reading frame of murine mammary tumor virus
1Department of Retrovirology, Radiobiological Institute, Rijswijk, The Netherlands.
Abstract:
The procaryotic chloramphenicol acetyltransferase (CAT) gene controlled by the murine mammary tumor virus (MMTV) promoter shows reduced activity in a rat mammary tumor cell line after infection with MMTV but to a considerably lesser extent than the CAT gene controlled by a heterologous promoter, indicating trans-acting regulation of promoter activity by MMTV. Cotransfection of vectors capable of expressing RNA from the 3' open reading frame (orf) of MMTV with the CAT-MMTV construct resulted in enhanced CAT activity, suggesting that orf carries a transactivating potential. Since transactivation was also found with a vector containing only orf and part of the viral env gene, it was concluded that a separate transcriptional unit exists for the orf message.
Insights
Murine mammary tumor virus (MMTV) regulates gene activity. A specific MMTV gene region (orf) shows transactivating potential, enhancing chloramphenicol acetyltransferase (CAT) gene expression, indicating a separate MMTV transcriptional unit.
Area of Science:
- Molecular Biology
- Virology
- Gene Regulation
Background:
- The murine mammary tumor virus (MMTV) is a retrovirus known to influence host gene expression.
- Understanding MMTV's regulatory mechanisms is crucial for studying viral pathogenesis and host-virus interactions.
Purpose of the Study:
- To investigate the trans-acting regulatory potential of MMTV components on gene expression.
- To determine if specific MMTV regions, particularly the 3' open reading frame (orf), influence promoter activity.
Main Methods:
- Utilized the chloramphenicol acetyltransferase (CAT) gene as a reporter under the control of the MMTV promoter.
- Infected rat mammary tumor cells with MMTV and analyzed CAT gene activity.
- Performed cotransfection experiments with vectors expressing MMTV RNA, including the 3' orf, to assess transactivation.
Main Results:
- MMTV infection led to reduced CAT gene activity, but less significantly than with a heterologous promoter, suggesting MMTV-mediated trans-acting regulation.
- Cotransfection with vectors expressing the MMTV 3' orf enhanced CAT activity, indicating a transactivating role for this region.
- Transactivation was observed even with vectors containing only the orf and part of the viral env gene.
Conclusions:
- MMTV possesses trans-acting regulatory elements that modulate gene expression.
- The 3' open reading frame (orf) of MMTV exhibits transactivating potential.
- Evidence suggests the existence of a separate transcriptional unit for the MMTV orf message.