Biology subjects

Massa, P. A.

Publications and source records attributed to Massa, P. A..

3 recordsLinked to original sources

bioRxiv2025

Herpes simplex virus 1 strain 17+ with R2 mutation in UL37 has residual retrograde transport

Herpes simplex virus 1 (HSV-1) causes lifelong recurrent infections. Following primary infection of the oral or genital mucosa, HSV-1 travels retrogradely through axons and establishes latency in the cell body of ganglionic neurons of the peripheral nervous system. Periodic reactivation in neurons and anterograde transport of virions back to peripheral regions causes oral or genital ulcerations. Many host and viral factors implicated in retrograde and anterograde transport of HSV-1 have been identified. In particular, studies reported that introducing five amino acid substitutions in the R2 region of the viral tegument protein UL37 was sufficient to completely eliminate retrograde transport of HSV-1 strain F. Here, we introduced the same R2 mutations in the highly neurovirulent HSV-1 strain 17+. We report that this R217 virus has residual retrograde travel. We show that R217 can establish latency in mouse models of ocular and vaginal infection and reactivate. These results contradict published evidence and show that the R2 mutation is not sufficient to fully prevent retrograde transport of HSV-1.

microbiology↗

bioRxiv2025

Minimization of gene editing off-target effects by tissue restriction of expression

Therapeutic in vivo gene editing with highly specific nucleases has the potential to revolutionize treatment for a wide range of human diseases, including genetic disorders and latent viral infections like herpes simplex virus (HSV). However, challenges regarding specificity, efficiency, delivery, and safety must be addressed before its clinical application. A key concern is the risk of off-target effects, which can cause unintended and potentially harmful genetic changes. We previously developed a curative in vivo gene editing approach to eliminate latent HSV using HSV-specific meganuclease delivered by an AAV vector. In this study, we investigate off-target effects of meganuclease by identifying potential off-target sites through GUIDE-tag analysis and assessing genetic alterations using amplicon deep sequencing in tissues from meganuclease treated mice. Our results show that meganuclease expression driven by a ubiquitous promoter leads to high off-target gene editing in the mouse liver, a non-relevant target tissue. However, restricting the meganuclease expression with a neuron-specific promoter and/or a liver-specific miRNA target sequence efficiently reduces off-target effects in both liver and trigeminal ganglia. These findings suggest that incorporation of regulatory DNA elements for tissue-specific expression in viral vectors can reduce off-target effects and improve the safety of therapeutic in vivo gene editing.

molecular biology↗

bioRxiv2023

Viral gene drive spread during herpes simplex virus 1 infection in mice

Gene drives are genetic modifications designed to propagate efficiently through a population. Most applications rely on homologous recombination during sexual reproduction in diploid organisms such as insects, but we recently developed a gene drive in herpesviruses that relies on co-infection of cells by wild-type and engineered viruses. Here, we developed a viral gene drive against human herpes simplex virus 1 (HSV-1) and showed that it propagated efficiently in vitro and during HSV-1 infection in mice. We observed high levels of co-infection and gene drive-mediated recombination in neuronal tissues during herpes encephalitis as the infection progressed from the site of inoculation to the peripheral and central nervous systems. In addition, we found evidence that a superinfecting gene drive virus could recombine with wild-type viruses during latent infection. These findings indicated that HSV-1 achieves high rates of co-infection and recombination during viral infection, a phenomenon that is currently underappreciated. Overall, this study showed that a viral gene drive could spread in vivo during HSV-1 infection, paving the way toward therapeutic applications.

microbiology↗