AUTHOR=Dang Louis T. , Glanowska Katarzyna M. , Iffland II Philip H. , Barnes Allan E. , Baybis Marianna , Liu Yu , Patino Gustavo , Vaid Shivanshi , Streicher Alexandra M. , Parker Whitney E. , Kim Seonhee , Moon Uk Yeol , Henry Frederick E. , Murphy Geoffrey G. , Sutton Michael , Parent Jack M. , Crino Peter B. 

TITLE=Multimodal Analysis of STRADA Function in Brain Development

JOURNAL=Frontiers in Cellular Neuroscience

VOLUME=14

YEAR=2020

URL=https://www.frontiersin.org/journals/cellular-neuroscience/articles/10.3389/fncel.2020.00122

DOI=10.3389/fncel.2020.00122

ISSN=1662-5102

ABSTRACT=<p>mTORopathies are a heterogeneous group of neurological disorders characterized by malformations of cortical development (MCD), enhanced cellular mechanistic target of rapamycin (mTOR) signaling, and epilepsy that results from mutations in mTOR pathway regulatory genes. Homozygous mutations (del exon 9–13) in the pseudokinase STE20-related kinase adaptor alpha (<italic>STRAD-α</italic>; <italic>STRADA</italic>), an mTOR modulator, are associated with Pretzel Syndrome (PS), a neurodevelopmental disorder within the Old Order Mennonite Community characterized by megalencephaly, intellectual disability, and intractable epilepsy. To study the cellular mechanisms of STRADA loss, we generated CRISPR-edited <italic>Strada</italic> mouse N2a cells, a germline mouse <italic>Strada</italic> knockout (KO−/−) strain, and induced pluripotent stem cell (iPSC)-derived neurons from PS individuals harboring the <italic>STRADA</italic> founder mutation. <italic>Strada</italic> KO <italic>in vitro</italic> leads to enhanced mTOR signaling and iPSC-derived neurons from PS individuals exhibit enhanced cell size and mTOR signaling activation, as well as subtle alterations in electrical firing properties e.g., increased input resistance, a more depolarized resting membrane potential, and decreased threshold for action potential (AP) generation. <italic>Strada</italic>−/− mice exhibit high rates of perinatal mortality and out of more than 100 litters yielding both WT and heterozygous pups, only eight <italic>Strada</italic>−/− animals survived past P5. <italic>Strada</italic>−/− mice are hypotonic and tremulous. Histopathological examination (<italic>n</italic> = 5 mice) revealed normal gross brain organization and lamination but all had ventriculomegaly. Ectopic neurons were seen in all five <italic>Strada</italic>−/− brains within the subcortical white matter mirroring what is observed in human PS brain tissue. These distinct experimental platforms demonstrate that STRADA modulates mTOR signaling and is a key regulator of cell size, neuronal excitability, and cortical lamination.</p>