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Kinetics and regulation of transcription of bacteriophage Mu
Abstract:
Mu transcription was analyzed by hybridization of [3H]uridine pulse-labeled RNA from heat-induced Mu lysogens to Mu DNA restriction fragments on nitrocellulose blots. Based on their time of appearance and dependence on Mu functions, we have defined three classes of transcripts: early, middle, and late. Replication-defective prophages containing A or B amber mutations or a deletion of the beta (right) end produced only early RNA derived from the left-most 8 to 10 kb of the Mu genome. A replication-proficient C amber mutant exhibited similar early transcription but at later times also produced middle transcripts from a region including C, which encodes the activator of late transcription. The C mutant did not produce late transcripts from the right-most 26 kb of the Mu genome encoding genes involved in phage morphogenesis and release. These results indicate that Mu DNA replication is required for efficient expression of middle RNA, which is itself required for expression of late transcripts. Amber mutations in essential genes other than A, B, and C had no significant effect on transcription except for polarity of one E mutation. Uninduced Mu c+ and Mu cts prophages produced very low levels of Mu-specific RNA derived from several regions including the c (immunity) gene and the region between genes B and C.
Insights
Bacteriophage Mu transcription occurs in three stages: early, middle, and late. Mu DNA replication is essential for middle and late transcript expression, which are required for phage development.
Area of Science:
- Molecular Biology
- Virology
- Bacteriophage Genetics
Background:
- Bacteriophage Mu is a temperate bacteriophage known for its transpositional activity.
- Understanding Mu gene expression is crucial for deciphering its lytic and lysogenic cycles.
Purpose of the Study:
- To investigate the temporal regulation of Mu transcription.
- To determine the role of Mu DNA replication and specific Mu genes in transcript production.
Main Methods:
- Analysis of Mu transcription using [3H]uridine pulse-labeled RNA.
- Hybridization of RNA to Mu DNA restriction fragments on nitrocellulose blots.
- Utilizing various Mu mutants (amber, deletion) to assess gene function.
Main Results:
- Three classes of Mu transcripts were identified: early, middle, and late, based on timing and gene dependence.
- Replication-defective mutants produced only early transcripts.
- A replication-proficient mutant defective in gene C produced early and middle transcripts but lacked late transcripts.
- Mu DNA replication is required for efficient middle RNA expression, which in turn is necessary for late transcript production.
Conclusions:
- Mu transcription is tightly regulated and proceeds sequentially through early, middle, and late phases.
- Mu DNA replication is a critical regulatory step, enabling the transition from early to middle and subsequently to late gene expression.
- The activator of late transcription (encoded by gene C) requires middle transcripts for its function, highlighting a cascade regulatory mechanism.