Masters Theses

Donald Henley

Date of Award

5-1990

Degree Type

Thesis

Degree Name

Master of Science

Major

Microbiology

Major Professor

Jerry P. Weir

Committee Members

Jeffrey M. Becker, David A. Brian

Abstract

The relative stabilities of Herpes Simplex Virus Type 1 (HSV-1) mRNAs were investigated by determining the half-lives of representative transcripts of each of the temporal classes of genes. For the immediate-early transcripts of the infected cell proteins (ICP) 0 and 27, tissue cultures of Vero cells were infected with HSV-1 and RNA synthesis inhibited by the addition of actinomycin-D at 3 hours post-infection. Single cultures were harvested at regular intervals, and RNA isolated by the guanidinium isothiocyanate/ cesium chloride method. The half-life of a given transcript was determined by quantitating the level of that transcript by Northern analysis at each successive timepoint after the inhibition of RNA synthesis. Half-lives were determined to be 4.2 ± 0.5 hours for the ICP 0 mRNA and 7.2 ± 1.8 hours for the ICP 27 mRNA. The half-life of the early mRNA for the thymidine kinase (TK) gene was indicated to be 8.2 ± 3.1 hours when RNA synthesis was inhibited at 5 hours post-infection. The stability of these mRNAs contrasts with that found for the three late transcripts for the glycoproteins C, E, and H which were found to have half-lives of 14 ± 2.0, 29 ± 6.4, and 25 ± 9.0 hours, respectively, when RNA synthesis was inhibited at 12 hours postinfection. Additionally, the late glycoprotein H mRNA exhibited reduced stability with a half-life of only 11 ± 4.2 hours when examined at 5 rather than 12 hours hours post-infection. It appears that there is a significant difference in the stability of mRNAs transcribed early in infection versus those transcribed late in infection. The mechanism responsible for this difference could have a role in the overall scheme of HSV-1 genetic regulation.

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