INDUCTION AND SHUT OFF OF DIHYDROFOLATE REDUCTASE GENE EXPRESSION DURING PRODUCTIVE INFECTION BY ADENOVIRUS TYPE 2
ROTH, SHARON YODER
Doctor of Philosophy
The effects of productive adenovirus infection on host gene expression were studied using a line of methotrexate-resistant HeLa cells containing amplified amounts of the gene coding for dihydrofolate reductase (DHFR). These cells contain several hundred copies of the DHFR gene and overproduce DHFR RNA and protein. Synthesis of DHFR is induced early in adenovirus infection, before the onset of viral replication, and is suppressed late in infection along with general host protein synthesis. These changes in DHFR protein synthesis are accompanied by changes in both the steady state cytoplasmic levels of DHFR mRNA and the rate of appearance of DHFR mRNA into the cytoplasm. These data indicate that nuclear events are important in the control of DHFR gene expression during productive infection. Transcription of DHFR-specific sequences continues at a constant rate throughout infection. In contrast, steady state nuclear levels of DHFR-specific sequences change in correspondence to the changing rate of appearance of DHFR mRNA into the cytoplasm through the course of infection. Thus, constitutive transcription of DHFR sequences is followed by a differential accumultion of DHFR nuclear RNA at different times during infection, indicating that post-transcriptional events are important in the regulation of DHFR gene expression by adenovirus. Sepcifically, the regulation of DHFR gene expression during infection is accomplished by a modulation of the relative stability of DHFR-specific sequences through the course of infection. During induction, DHFR sequences are more stable than in either uninfected cells or infected cells late in infection. The induction of DHFR is temporally linked to viral replication during both wild type infection and infection with defined early adenoviral mutants which delay replication. Studies with mutant defective in the production of E4 or E1b gene products indicate that E4 gene products are involved in the switch from early to late viral gene expression during the time of DHFR induction, and that both E4 and E1b gene products are necessary for the efficient shut off of DHFR and other host genes late in infection.