Biology subjects

tenOever, B.

Publications and source records attributed to tenOever, B..

2 recordsLinked to original sources

bioRxiv2020

An inability to maintain the ribonucleoprotein genomic structure is responsible for host detection of negative-sense RNA viruses

Cellular biology has a uniformity not shared amongst viruses. This is perhaps best exemplified by negative-sense RNA viruses that encode their genetic material as a ribonucleoprotein complex composed of genome, RNA-dependent RNA polymerase, and the nucleoprotein. Here we demonstrate that limiting nucleoprotein availability not only universally culminates in a replicative catastrophe for negative-sense RNA viruses, but it results in the production of aberrant genomic material and induction of the interferon-based host defenses. This dynamic illustrates the tremendous stress imposed on negative-sense RNA viruses during replication as genomic products accumulate in an environment that requires an increasing demand on nucleoprotein availability. We show that limiting NP by RNA interference or drug targeting blocks replication and primes neighboring cells through the production of interferon. Together, these results demonstrate that the nucleoprotein represents the Achilles heel of the entire phylum of negative-sense RNA viruses. Here we establish this principle for a diverse collection of human pathogens and propose that the nucleoprotein should be a primary target for the development of future antiviral drugs. HIGHLIGHTSO_LILimited levels of NP result in production of defective viral genomes C_LIO_LIDefective viral genomes and viral antagonists are key determinants of the host antiviral response C_LIO_LIThe host response and defective viral genome generation further exasperate NP availability C_LIO_LINP is an optimal drug target for the whole phylum of negative-sense RNA viruses C_LI

microbiology↗

bioRxiv2020

Engineering an RNA-based tissue-specific platform for genetic editing through use of a miRNA-enabled Cas12a

The capacity to edit genomes using CRISPR-Cas systems holds immense potential for countless genetic-based diseases. However, one significant impediment preventing broad therapeutic utilization is in vivo delivery. While genetic editing at a single cell level in vitro can be achieved with high efficiency, the capacity to utilize these same biologic tools in a desired tissue in vivo remains challenging. Non-integrating RNA virus-based vectors constitute efficient platforms for transgene expression and surpass several barriers to in vivo delivery. However, the broad tissue tropism of viral vectors raises the concern for off-target effects. Moreover, prolonged expression of the Cas proteins, regardless of delivery method, can accumulate aberrant RNAs leading to unwanted immunological responses. In an effort to circumvent these shortcomings, here we describe a versatile RNA virus-based technology that can achieve cell-specific activity and self-inactivation by combining host microRNA (miRNA) biology with the CRISPR-Cas12a RNA-guided nuclease. Exploiting the RNase activity of Cas12a, we generated a vector that self-inactivates upon delivery of Cas12a and an accompanying CRISPR RNA (crRNA). Furthermore, we show that maturation of the crRNA can be made dependent on cell-specific miRNAs, which confers cell-specificity. We demonstrate that this genetic editing circuit delivers diminished yet sufficient levels of Cas12a to achieve effective genome editing whilst inducing a minimal immunological response. It can also function in a cell-specific manner thereby facilitating in vivo editing and mitigating the risk of unwanted, off-target effects.

bioengineering↗