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An RNA polymerase II transcription factor inactivated in poliovirus-infected cells copurifies with transcription
1Department of Microbiology and Immunology, School of Medicine, University of California, Los Angeles, 90024-1747.
Abstract:
Inhibition of host cell RNA polymerase II-mediated transcription by poliovirus infection was studied in vitro. Whole-cell extracts prepared from poliovirus-infected HeLa cells at 3 h postinfection were shown to be deficient in a factor required for specific transcription from the adenovirus major late promoter. Three lines of evidence suggest that transcription factor TFIID is deficient in poliovirus-infected cells. First, the activity required to specifically restore transcription in poliovirus-infected cell extracts was shown to copurify with TFIID through three chromatographic steps. Second, transcription reactions reconstituted with phosphocellulose-derived chromatographic fractions revealed a fourfold decrease in the specific activity of the TFIID-containing fraction prepared from poliovirus-infected cells compared with that of the same fraction prepared from mock-infected cells. Finally, TFIID and the activity required to specifically restore transcription in virus-infected cell extracts were shown to have the same kinetics of heat inactivation. Together, these results suggest that inactivation of TFIID is an early event in the inhibition of host cell RNA polymerase II transcription by poliovirus.
Insights
Poliovirus infection inactivates transcription factor TFIID, a key component of RNA polymerase II transcription. This early inactivation disrupts host cell transcription, impacting viral replication.
Area of Science:
- Molecular Biology
- Virology
- Biochemistry
Background:
- Poliovirus infection inhibits host cell RNA polymerase II transcription.
- The precise mechanism of this inhibition is not fully understood.
Purpose of the Study:
- To investigate the molecular mechanisms underlying poliovirus-induced inhibition of host cell transcription.
- To identify the specific host factors affected by poliovirus infection.
Main Methods:
- In vitro transcription assays using whole-cell extracts from infected and mock-infected HeLa cells.
- Chromatographic purification to isolate and identify the deficient transcription factor.
- Heat inactivation kinetics to compare the stability of TFIID and the transcription-restoring activity.
Main Results:
- Poliovirus-infected cell extracts showed a deficiency in a factor essential for transcription from the adenovirus major late promoter.
- This deficient factor copurified with transcription factor TFIID through multiple chromatographic steps.
- The TFIID-containing fraction from infected cells exhibited a fourfold decrease in specific activity compared to mock-infected cells.
- TFIID and the transcription-restoring activity displayed similar heat inactivation kinetics.
Conclusions:
- Transcription factor TFIID is inactivated early during poliovirus infection.
- Inactivation of TFIID is a key event contributing to the inhibition of host cell RNA polymerase II transcription by poliovirus.