AUTHOR=Le Nguyet , Appel Haley , Pannullo Nicole , Hoang Thanh , Blackshaw Seth 

TITLE=Ectopic insert-dependent neuronal expression of GFAP promoter-driven AAV constructs in adult mouse retina

JOURNAL=Frontiers in Cell and Developmental Biology

VOLUME=Volume 10 - 2022

YEAR=2022

URL=https://www.frontiersin.org/journals/cell-and-developmental-biology/articles/10.3389/fcell.2022.914386

DOI=10.3389/fcell.2022.914386

ISSN=2296-634X

ABSTRACT=Direct viral-mediated reprogramming of retinal Muller glia is a promising avenue for replacing photoreceptors and retinal ganglion cells lost to retinal dystrophies.  However, questions have recently been raised about the accuracy of claims of efficient glia-to-neuron reprogramming in retina that were conducted using GFAP minipromoter-based adeno-associated virus (AAV) vectors.  In this study, we have addressed this question using GFAP minipromoter-driven AAV constructs to simulaneously overexpress the mCherry reporter and candidate transcription factors predicted to induce glia-to-neuron conversion, in combination with prospective genetic labeling of retinal Muller glia using inducible Cre-dependent GFP reporters.  We find that, while control GFAP-mCherry constructs express faithfully in Muller glia, that 5 out of 7 transcription factor overexpression constructs tested are predominantly expressed in amacrine and retinal ganglion cells, but that genetic cell lineage analysis shows no evidence for glia-to-neuron conversion.  These findings demonstrate major insert-dependent effects on AAV-based GFAP minipromoter specificity that preclude its use in inferring cell lineage relationships in this context.