Biology subjects

Steele, A. D.

Publications and source records attributed to Steele, A. D..

4 recordsLinked to original sources

bioRxiv2026

Systemic AAV delivery of a calcium indicator in marmosets: functional validation in visual area MT

Functional optical imaging in nonhuman primates provides an important complement to electrophysiological approaches in neuroscience research, but its broader use has been limited by challenges in achieving large-scale, homogeneous expression of genetically encoded reporters, and imaging accessibility in species with gyrencephalic brains with sulci and fissures (e.g., rhesus macaques). Specifically, conventional local intracortical viral injections are invasive and often produce spatially restricted or heterogeneous expression, constraining population-level analyses. Here, we show that systemic intravenous delivery of an adeno-associated virus (AAV) capsid engineered for enhanced blood-brain barrier crossing, AAV.CAP-B10, supports robust and widespread expression of a calcium indicator CAaMP8s in the common marmoset. Intravenous delivery in two marmosets resulted in widespread cortical expression. Using a large cranial window over extrastriate visual area MT (and its satellite areas), we performed widefield single-photon imaging and two-photon cellular-resolution imaging in awake,behaving marmosets to functionally validate activity in this well studied primate visual-motion sensitive cortical area. Population level responses to visual motion and spatial organization measured with widefield imaging, as well as single-cell level motion direction tuning measured with two-photon imaging, were consistent with canonical properties of MT reported in previous electrophysiological studies. Quantitative analyses of lightsheet imaging after whole hemisphere brain clearing further confirmed the broad expression of GCaMP in both cortical and subcortical areas. Together, these results indicate that systemic delivery using AAV.CAP-B10 provides a minimally invasive approach for robust multi-scale functional optical imaging in awake, behaving marmosets.

neuroscience↗

bioRxiv2025

AAVs Targeting Human Carbonic Anhydrase IV Enhance Gene Delivery to the Brain

Clinically approved gene therapies based on natural adeno-associated virus (AAV) serotypes have restricted applications, particularly in the brain, due to their poor targeting, high dose requirements, and resulting safety concerns. Directed evolution of enhanced AAV capsids in mice or non-human primates (NHPs) has resulted in markedly improved performance in those species, but inter-species differences present a serious challenge for translating these vectors into human therapies. Here, we engineer AAVs to target human carbonic anhydrase IV (CA-IV), a recently identified blood-brain barrier (BBB) transcytosis receptor. Among known transcytosis receptors, CA-IV is notable for its relatively specific expression in brain endothelial cells and the potency of AAVs that target it in mice. CA-IVs AAV binding site, and thus the mouse vectors enhanced brain potency, is not conserved across species, so we employed a two-phase engineering strategy to identify AAVs optimized for human CA-IV-dependent gene delivery to the brain. We first used in vitro receptor-based selection of a vast AAV library to exclude capsids that do not bind human CA-IV, followed by in vivo selection in "humanized" mice expressing human CA-IV in brain endothelial cells. Notably, we find that human CA-IV binding capsid variants that were poorly enriched in the pull-down selection outperform strong binders in vivo. The most promising vector, AAV-hCA4-IV77, engages human CA-IV to achieve 100-fold greater brain transduction than AAV9, with robust neuronal and astrocytic coverage throughout multiple brain regions. These results advance our understanding of receptor-targeted capsid design and support the therapeutic potential of human CA-IV-engaging AAVs.

bioengineering↗

bioRxiv2023

The Natural Products Discovery Center: Release of the First 8490 Sequenced Strains for Exploring Actinobacteria Biosynthetic Diversity

Actinobacteria, the bacterial phylum most renowned for natural product discovery, has been established as a valuable source for drug discovery and biotechnology but is underrepresented within accessible genome and strain collections. Herein, we introduce the Natural Products Discovery Center (NPDC), featuring 122,449 strains assembled over eight decades, the genomes of the first 8490 NPDC strains (7142 Actinobacteria), and the online NPDC Portal making both strains and genomes publicly available. A comparative survey of RefSeq and NPDC Actinobacteria highlights the taxonomic and biosynthetic diversity within the NPDC collection, including three new genera, hundreds of new species, and [~]7000 new gene cluster families. Selected examples demonstrate how the NPDC Portals strain metadata, genomes, and biosynthetic gene clusters can be leveraged using genome mining approaches. Our findings underscore the ongoing significance of Actinobacteria in natural product discovery, and the NPDC serves as an unparalleled resource for both Actinobacteria strains and genomes.

microbiology↗

bioRxiv2020

Type 1 dopamine receptor (D1R)-independent circadian food anticipatory activity in mice

Circadian rhythms are entrained by light and influenced by non-photic stimuli, such as feeding. The activity preceding scheduled mealtimes, food anticipatory activity (FAA), is elicited in rodents fed a limited amount at scheduled times. FAA is thought to be the output of an unidentified food entrained oscillator. Previous studies, using gene deletion and receptor pharmacology, implicated dopamine type receptor 1 (D1R) signaling in the dorsal striatum as necessary for FAA in mice. To further understand the role of D1R in promoting FAA, we utilized the Cre-lox system to create cell type-specific deletions of D1R. We were unsuccessful in obtaining conditional deletion of D1R when using transgenically driven D1R-Cre. We then created a conditional deletion of D1R in GABA neurons using Vgat-ires-Cre line, which had attenuated FAA, but the amount was higher than expected based on prior results using a constitutive knockout of D1R, D1R KODrago. This result prompted us to re-test the original D1R KODrago line, which expressed less FAA than controls, but only moderately so. To determine if genetic drift had diminished the effect of D1R deletion on FAA, we re-established the D1R KODrago knockout line from cryopreserved samples. The reestablished D1R KODrago-cryo had a clear impairment of FAA compared to controls, but still developed increased activity preceding mealtime across the 4 weeks of timed feeding. Finally, we tested a different deletion allele of D1R created by the Knockout Mouse Project. This line of D1R KOKOMP mice had a significant impairment in the acquisition of FAA, but eventually reached similar levels of premeal activity compared to controls after 4 weeks of timed feeding. Taken together, our results suggest that D1R signaling promotes FAA, but other dopamine receptors likely contribute to FAA given that mice lacking the D1 receptor still retain some FAA.

neuroscience↗