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Synthesis of the adenovirus-coded DNA binding protein in infected cells
Abstract:
Synthesis of the 75K (75K indicates a moleculatr weight of 70,000 to 75,000) DNA binding protein, an early virus-coded protein in adenovirus 2-infected KB cells, and its regulation were studied by using a radioimmune precipitation inhibition assay. The protein was first detected at 4 h postinfection and accumulated at an expoential rate. An arrest of further synthesis (accumulation) was observed at 10 to 11 h postinfection, coinciding with the onset of synthesis of late virion proteins. In contrast, when the infected cells were treated with 25 mug of arabinosyl cytosine per ml to block viral DNA replication, the synthesis of 75K protein did not cease but continue for up to 36 h postinfection. The synthesis of 75K protein in cells after release from a cycloheximide block (2 to 9 h postinfection) was analyzed. Increased amounts of early adenovirus-specific mRNA accumulate in infected cells during a cycloheximide block (Parsons and Green, 1971). However, cycloheximide treatment did not produce increased levels of 75K protein, and an abrupt arrest of 75K protein formation was again observed at the time of synthesis of late virion proteins. Partition of the 75K protein between the nuclear and cytoplasmic fractions during the course of infection was studied. The 75K protein appeared first in the cytoplasm and then in the nucleus after a slight lag. Accumulation of the 75K protein continued both in the cytoplasm and nucleus, with higher levels being found in the cytoplasm.
Insights
The synthesis of early adenovirus 75K DNA binding protein is regulated by viral DNA replication. Blocking replication with arabinosyl cytosine extends 75K protein synthesis, unlike cycloheximide treatment.
Area of Science:
- Molecular Biology
- Virology
- Cell Biology
Background:
- Adenovirus 2 infection involves the synthesis of early and late viral proteins.
- A specific 75K DNA binding protein is identified as an early virus-coded protein.
Purpose of the Study:
- To investigate the synthesis and regulation of the early adenovirus 75K DNA binding protein.
- To understand the relationship between 75K protein synthesis, viral DNA replication, and late protein production.
Main Methods:
- Radioimmune precipitation inhibition assay was used to detect and quantify the 75K protein.
- Experiments involved arabinosyl cytosine treatment to block viral DNA replication.
- Cycloheximide treatment was employed to study protein synthesis regulation.
Main Results:
- 75K protein synthesis began at 4 h postinfection and accumulated exponentially until 10-11 h, coinciding with late protein synthesis.
- Blocking viral DNA replication with arabinosyl cytosine allowed 75K protein synthesis to continue for up to 36 h.
- Cycloheximide treatment did not increase 75K protein levels and synthesis still arrested at the onset of late protein production.
- The 75K protein was detected first in the cytoplasm and later in the nucleus, with higher accumulation in the cytoplasm.
Conclusions:
- The synthesis of the early adenovirus 75K DNA binding protein is regulated by viral DNA replication.
- Viral DNA replication appears to be a trigger for the arrest of 75K protein synthesis and the onset of late protein production.