Precise CRISPR-Cas9-mediated mutation of a membrane trafficking domain in the Drosophila vesicular monoamine transporter gene

• DOI: 10.1016/j.crphys.2023.100101
• Publication Date: 2023-06-20T17:02:01Z
• Authors: James D. Asuncion; Aditya Eamani; Ethan W. Rohrbach; Elizabeth M. Knapp; Sonali A. Deshpande; Shivan L. Bonanno; Jeremy E. Murphy; Hakeem O. Lawal; David E. Krantz
• Keywords: Vesicular Transport Proteins; Vesicular monoamine transporter

Monoamine neurotransmitters such as noradrenalin are released from both synaptic vesicles (SVs) and large dense-core (LDCVs), the latter mediating extrasynaptic signaling. The contribution of versus signaling to circuit function behavior remains poorly understood. To address this question, we have previously used transgenes encoding a mutation in Drosophila Vesicular Transporter (dVMAT) that shifts amine release SVs LDCVs. circumvent use with non-endogenous patterns expression, now CRISPR-Cas9 generate trafficking mutant endogenous dVMAT gene. minimize disruption coding sequence nearby RNA splice site, precisely introduced point using single-stranded oligonucleotide repair. A predicted decrease fertility was phenotypic screen identify founders lieu visible marker. Phenotypic analysis revealed defect ovulation mature follicles egg retention ovaries. We did not detect defects contraction lateral oviducts following optogenetic stimulation octopaminergic neurons. Our findings suggest eggs ovary is disrupted by changing balance VMAT between Further experiments model will help determine mechanisms sensitize specific circuits changes