Sara Gharib

Canada

Determination of the Neurobiological Origins of the CHD8 Phenotype Using Genetic Cre Drivers

Sara Gharib¹, Ritah Sansa², Esther M. Garfield², Samantha Dhanani², Ariine Hoque², Muhammad O. Chohan², Jeremy Veenstra-VanderWeele², Rebecca Muhle²

1. Department of Neuroscience and Behavior, Barnard College, Columbia University, New York, NY, USA
2. Department of Adolescent Psychiatry, New York State Psychiatric Institute, Columbia University Medical Center, New York, NY, USA

Abstract

Background

Background: CHD8 (chromodomain helicase DNA-binding protein 8) is a high-confidence gene associated with autism spectrum disorders (ASD). Its neurobiological role is studied through Cre-driver systems in mice, aiming to understand the genetic and molecular mechanisms underlying ASD phenotypes.

Methods

Methods: Using Th-Cre genetic drivers, a mouse model was designed to induce targeted CHD8 deletions in neural tissues. Mating schemes were implemented to produce desired genotypes. Neural tissues were collected, and DNA and RNA were extracted for analysis. Deletion PCR and qPCR were performed to confirm CHD8 deletions and evaluate transcriptional changes.

Results

Results: Although Th-Cre expression was confirmed, no deletion of CHD8 was observed in brain tissues, suggesting potential technical or biological challenges. Issues such as inaccurate brain dissections, interference by Th occupancy, or ineffective Cre recombinase activity were identified as possible explanations. These findings refute the hypothesis that Th-Cre would successfully mediate CHD8 deletion in neural tissues using the current breeding strategy.

Conclusions

Conclusion: The study highlights significant challenges in relying on Th-Cre drivers for CHD8 deletion in the brain. Future research will refine experimental protocols, including improved dissection techniques, alternative breeding strategies, and testing other Cre drivers (e.g., Emx1-Cre). Additional experiments, such as crossing CHD8fl/fl mice with floxed GFP mice for fluorescence-based validation, and incorporating immunohistochemistry, will enhance the reliability of Cre-mediated recombination. These steps aim to further explore CHD8’s role in ASD and uncover the genetic mechanisms involved.