Isolation and characterization of a functional cDNA encoding ICP0 from herpes simplex virus type 1

Area of Science:

• Molecular Virology
• Gene Expression Regulation

Background:

• Herpes simplex virus type 1 (HSV-1) immediate-early gene 0 (IE-0) encodes ICP0, a potent transcriptional activator crucial for viral replication.
• ICP0's function is vital, but its expression in non-eukaryotic systems is challenging due to intron presence in the native gene.

Purpose of the Study:

• To isolate and characterize a functional cDNA clone encoding the HSV-1 ICP0 protein.
• To demonstrate the ICP0 cDNA's ability to direct ICP0 synthesis and transcriptional activation.
• To enable ICP0 overexpression in vector systems lacking eukaryotic splicing machinery.

Main Methods:

• Construction and screening of a lambda gt10 cDNA library from HSV-1 infected HeLa cells.
• DNA sequence analysis to confirm intron/exon boundaries.
• Plasmid transfection and immunofluorescence to assess ICP0 synthesis and localization.
• Transient expression assays to evaluate ICP0's transactivation capabilities on viral promoters.

Main Results:

• A functional cDNA clone encoding ICP0 was successfully isolated.
• Transfection of the cDNA directed ICP0 synthesis, with correct nuclear localization.
• The ICP0 cDNA retained transactivation ability on immediate-early, early, and late HSV-1 promoters.
• ICP0 synergistically activated the thymidine kinase promoter in cooperation with ICP4.

Conclusions:

• The isolated ICP0 cDNA clone is functional and retains biological activity.
• This cDNA provides a tool for expressing ICP0 in diverse expression systems, including those without eukaryotic splicing signals.
• Facilitates further investigation into ICP0's role in HSV-1 gene regulation and viral pathogenesis.