Human Cytomegalovirus (HCMV) is a leading cause of birth defects. Ramifications of HCMV infection are primarily observed in the central nervous system (CNS) and include hearing loss, vision loss, microcephaly and mental retardation. Despite considerable effort, the underlying mechanisms causing these CNS defects remain unknown. Our previously funded studies of HCMV interaction with the host cell DNA and its DNA repair machinery have identified three areas of particular interest to pursue. First, HCMV is one of only two viruses known to inflict site-specific chromosomal damage to the host DNA. Fine mapping of the chromosome 1q breaksites has revealed two genes, nidogen 1 (NID1) and myelin protein zero (MPZ), linked to the development of hearing loss in infected infants. Second, studies on DNA repair in HCMV infected permissive fibroblasts found that, although DNA damage responses were activated during infection, they were not completed. We suspect that compromised repair of specific and nonspecific DNA damage may play a role in the development of HCMV-induced birth defects. Third, we have recently begun working with a promising new in vitro model, the Neural Progenitor Cell (NPCs), and its derivatives. These cells, derived from post mortem neonatal brain tissue, provide a unique opportunity to investigate HCMV infection in a model system directly relevant to the human fetal CNS. NPCs, their glial derivatives, and the large majority of their neuronal derivatives, are fully permissive and suffer a lytic infection. However, a subpopulation of differentiated neurons, although permissive, exhibit extended viral antigen expression and release of virions. The long term goal of our work is to translate the information gained from studying infection in vitro, into understanding the development of CNS defects in congenitally infected infants. We propose examining clinical specimens for confirmatory evidence of the results found in our in vitro experiments. We have procured sample archival brain and auditory system tissues from neonates that have succumbed to HCMV infection, which will feature prominently in our proposed experiments. We will advance our long term goal with the testing of four hypotheses: 1) that a viral protein (or proteins) induces the site-specific breaks on Chromosome 1q;2) that the compromised repair of HCMV-induced breaks causes downregulation of breaksite-encoded genes;3) that HCMV disrupts the cellular DNA repair machinery's ability to repair non-specific damage in neural cells;and 4) that HCMV infection within the CNS affects expression of specific genes involved in differentiation, migration and cell function in NPCs and long-term neurons. The experiments described in this proposal will provide a detailed understanding of the molecular mechanisms underlying the genesis of HCMV-induced birth defects and contribute to the development of strategies to interrupt these mechanisms and, hopefully, prevent their frequently devastating consequences.

Public Health Relevance

Human Cytomegalovirus is a leading cause of birth defects. Those defects can dramatically impact the lives of the children (and their parents) that suffer from them. My research is aimed at understanding the underlying mechanism for the development of those defects in order to someday prevent their occurrence.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Research Project (R01)
Project #
5R01AI051463-08
Application #
8197095
Study Section
Virology - B Study Section (VIRB)
Program Officer
Beisel, Christopher E
Project Start
2002-06-01
Project End
2014-11-30
Budget Start
2011-12-01
Budget End
2012-11-30
Support Year
8
Fiscal Year
2012
Total Cost
$319,424
Indirect Cost
$96,674
Name
University of Idaho
Department
Microbiology/Immun/Virology
Type
Schools of Earth Sciences/Natur
DUNS #
075746271
City
Moscow
State
ID
Country
United States
Zip Code
83844
Zavala, Anamaria G; Kulkarni, Amit S; Fortunato, Elizabeth A (2014) A dual color Southern blot to visualize two genomes or genic regions simultaneously. J Virol Methods 198:64-8
Duan, Ying-Liang; Ye, Han-Qing; Zavala, Anamaria G et al. (2014) Maintenance of large numbers of virus genomes in human cytomegalovirus-infected T98G glioblastoma cells. J Virol 88:3861-73
Kulkarni, Amit S; Fortunato, Elizabeth A (2014) Modulation of homology-directed repair in T98G glioblastoma cells due to interactions between wildtype p53, Rad51 and HCMV IE1-72. Viruses 6:968-85
Kulkarni, A S; Fortunato, E A (2011) Stimulation of homology-directed repair at I-SceI-induced DNA breaks during the permissive life cycle of human cytomegalovirus. J Virol 85:6049-54
Luo, Min Hua; Hannemann, Holger; Kulkarni, Amit S et al. (2010) Human cytomegalovirus infection causes premature and abnormal differentiation of human neural progenitor cells. J Virol 84:3528-41
Hannemann, Holger; Rosenke, Kyle; O'Dowd, John M et al. (2009) The presence of p53 influences the expression of multiple human cytomegalovirus genes at early times postinfection. J Virol 83:4316-25
Luo, Min Hua; Schwartz, Philip H; Fortunato, Elizabeth A (2008) Neonatal neural progenitor cells and their neuronal and glial cell derivatives are fully permissive for human cytomegalovirus infection. J Virol 82:9994-10007
Nystad, Mona; Fagerheim, Toril; Brox, Vigdis et al. (2008) Human cytomegalovirus (HCMV) and hearing impairment: infection of fibroblast cells with HCMV induces chromosome breaks at 1q23.3, between loci DFNA7 and DFNA49 -- both involved in dominantly inherited, sensorineural, hearing impairment. Mutat Res 637:56-65
Luo, Min Hua; Fortunato, Elizabeth A (2007) Long-term infection and shedding of human cytomegalovirus in T98G glioblastoma cells. J Virol 81:10424-36
Luo, Min Hua; Rosenke, Kyle; Czornak, Kamila et al. (2007) Human cytomegalovirus disrupts both ataxia telangiectasia mutated protein (ATM)- and ATM-Rad3-related kinase-mediated DNA damage responses during lytic infection. J Virol 81:1934-50

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