# CYTOMEGALOVIRUS DNA REPLICATION AND RECOMBINATION

> **NIH NIH R01** · STANFORD UNIVERSITY · 1993 · $184,578

## Abstract

This proposal aims to understand the process and functions involved in
cytomegalovirus DNA replication.  Cytomegalovirus remains an important
viral pathogen in immunocompromized individuals such as organ transplant
recipients and ADDS patients and the only licenced anti-CMV antiviral,
gancyclovir, is directed at the DNA replication machinery.  The first aim
in the next grant period, is to study a cisacting replication initiation
sequence (oriLYT) located near the ICP8 gene and to localize the minimal
sequence elements necessary for origin function in transient
electroporation/infection assays. The oriLYT sequence from both the human
and murine cytomegalovirus genomes will be mapped and studied.  After
functionally important region(s) are defined, viral and cellular proteins
that interact with oriLYT sequence elements will be identified and cloned.
We will use direct functional assays and mutagenesis to investigate the
activity of the known CMV origin within the viral genome and to search for
other replication origins. The second aim is to identify the set of
trans-acting viral genes necessary to replicate oriLYT plasmids carrying
the human or murine CMV DNA replication origins by a transient replication
assay.  Genomic DNA clones as well as existing murine and human CMV
full-length cDNA libraries made in an SV40-based eukaryotic expression
vector will be screened to isolate and study replication functions believed
to encode the DNA polymerase, processivity factor, single stranded DNA
binding protein, helicase-primase and origin binding protein based on
similarity to HSV-1 gene products.  The candidate cDNA expression clones
encoding the necessary functions will be assayed by transfection of
CMV-permissive human fibroblast and U373-MG cells as well as nonpermissive
Vero cells and COS-7 cells, a cell line which will produce high levels of
each of the products.  All of the replication functions will also be
expressed as recombinant proteins in baculovirus vectors for study of
enzymatic function.  The final aim will be to develop genetic manipulation
strategies to construct defined mutants disrupting replication gene
function.  Viral mutants will be generated in each of the known or putative
replication functions to determine their role during viral growth.
Approaches that rely on cell lines that complement essential replication
functions and approaches that make use novel methods to disrupt gene
expression are proposed.  These investigations will generate new
information on an important pathogen as well as provide basic information
on how these large enveloped DNA viruses replicate their genomes.

## Key facts

- **NIH application ID:** 3129726
- **Project number:** 5R01AI020211-10
- **Recipient organization:** STANFORD UNIVERSITY
- **SAM.gov UEI:** HJD6G4D6TJY5
- **Principal Investigator:** EDWARD S. Edward S Mocarski
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 1993
- **Award amount:** $184,578
- **Award type:** 5
- **Project period:** 1984-03-01 → 1997-02-28

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/3129726

## Citation

> US National Institutes of Health, RePORTER application 3129726, CYTOMEGALOVIRUS DNA REPLICATION AND RECOMBINATION (5R01AI020211-10). Retrieved via MORVS 2026-10-25 from https://api.morvs.ai/grant/nih/3129726. Licensed CC0.

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