title: Light-Activated Nuclear Translocation of Adeno-Associated Virus Nanoparticles Using Phytochrome B for Enhanced, Tunable, and Spatially Programmable Gene Delivery.
abstract: Gene delivery vectors that are activated by external stimuli may allow improved control over the location and the degree of gene expression in target populations of cells. Light is an attractive stimulus because it does not cross-react with cellular signaling networks, has negligible toxicity, is noninvasive, and can be applied in space and time with unparalleled precision. We used the previously engineered red (R)/far-red (FR) light-switchable protein phytochrome B (PhyB) and its R light dependent interaction partner phytochrome interacting factor 6 (PIF6) from Arabidopsis thaliana to engineer an adeno-associated virus (AAV) platform whose gene delivery efficiency is controlled by light. Upon exposure to R light, AAV engineered to display PIF6 motifs on the capsid bind to PhyB tagged with a nuclear localization sequence (NLS), resulting in significantly increased translocation of viruses into the host cell nucleus and overall gene delivery efficiency. By modulating the ratio of R to FR light, the gene delivery efficiency can be tuned to as little as 35% or over 600% of the unengineered AAV. We also demonstrate spatial control of gene delivery using projected patterns of codelivered R and FR light. Overall, our successful use of light-switchable proteins in virus capsid engineering extends these important optogenetic tools into the adjacent realm of nucleic acid delivery and enables enhanced, tunable, and spatially controllable regulation of viral gene delivery. Our current light-triggered viral gene delivery prototype may be broadly useful for genetic manipulation of cells ex vivo or in vivo in transgenic model organisms, with the ultimate prospect of achieving dose- and site-specific gene expression profiles for either therapeutic (e.g., regenerative medicine) or fundamental discovery research efforts.
authors.affiliation: Department of Bioengineering, ‡Systems, Synthetic, and Physical Biology Program, and §Department of Biochemistry and Cell Biology, Rice University , Houston, Texas 77005, United States.
authors.forename: Eric J
authors.initials: EJ
authors.lastname: Gomez
authors.affiliation: Department of Bioengineering, ‡Systems, Synthetic, and Physical Biology Program, and §Department of Biochemistry and Cell Biology, Rice University , Houston, Texas 77005, United States.
authors.forename: Karl
authors.initials: K
authors.lastname: Gerhardt
authors.affiliation: Department of Bioengineering, ‡Systems, Synthetic, and Physical Biology Program, and §Department of Biochemistry and Cell Biology, Rice University , Houston, Texas 77005, United States.
authors.forename: Justin
authors.initials: J
authors.lastname: Judd
authors.affiliation: Department of Bioengineering, ‡Systems, Synthetic, and Physical Biology Program, and §Department of Biochemistry and Cell Biology, Rice University , Houston, Texas 77005, United States.
authors.forename: Jeffrey J
authors.initials: JJ
authors.lastname: Tabor
authors.affiliation: Department of Bioengineering, ‡Systems, Synthetic, and Physical Biology Program, and §Department of Biochemistry and Cell Biology, Rice University , Houston, Texas 77005, United States.
authors.forename: Junghae
authors.initials: J
authors.lastname: Suh
mesh_terms: D017360:Arabidopsis / Q000737:chemistry; D029681:Arabidopsis Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D051792:Basic Helix-Loop-Helix Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D002213:Capsid / Q000737:chemistry / Q000378:metabolism; D002467:Cell Nucleus / Q000235:genetics* / Q000378:metabolism / Q000528:radiation effects / Q000648:ultrastructure; D000229:Dependovirus / Q000235:genetics / Q000378:metabolism / Q000528:radiation effects; D004023:Dictyostelium / Q000235:genetics / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D015870:Gene Expression; D018014:Gene Transfer Techniques; D005822:Genetic Vectors / Q000737:chemistry / Q000378:metabolism; D057809:HEK293 Cells; D006801:Humans; D008027:Light; D053758:Nanoparticles / Q000737:chemistry / Q000528:radiation effects; D019913:Nuclear Localization Signals; D062308:Optogenetics / Q000379:methods*; D051939:Phytochrome B / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D011485:Protein Binding; D015202:Protein Engineering; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism
chemical_list: D029681:Arabidopsis Proteins; D051792:Basic Helix-Loop-Helix Proteins; D019913:Nuclear Localization Signals; D051939:Phytochrome B; D011994:Recombinant Proteins; C497730:PIF6 protein, Arabidopsis
keywords: light-activatable; light-switchable; optogenetics; phytochrome; stimulus-responsive; synthetic biology; synthetic virology; viral gene delivery