Tbx1 Heterozygosity in the Oligodendrocyte Lineage Shifts Myelinated Axon Composition in the Mouse Fimbria Without Behavioral Impairments

ABSTRACT Constitutive heterozygosity of Tbx1, a T-box transcription factor gene located within the 22q11.2 deletion region, results in behavioral deficits and altered composition of myelinated axons in the fimbria, together with reduced levels of an oligodendrocyte precursor cell (OPC) marker, in mice. However, the cellular origins of these effects and the extent to which axonal changes causally contribute to behavioral impairments remain unclear. We hypothesized that Tbx1 deficiency specifically within the oligodendrocyte lineage contributes to myelin and behavioral phenotypes. To test this hypothesis, we first demonstrated through in vitro siRNA knockdown that Tbx1 regulates both OPCs and mature oligodendrocytes. Subsequently, we assessed the impact of Tbx1 heterozygosity initiated in OPCs on behavioral and myelin phenotypes in male conditional PdgfrαCre;Tbx1+/flox mice. These mice exhibited Cre-mediated recombination in Pdgfrα-expressing brain regions and in the OPC progeny within the fimbria. At one month of age, the mutants displayed a higher rate of spontaneous alternation at the longest inter-trial interval in the T-maze compared to their wild-type littermates—an effect that was dissipated at two months. No significant phenotypic abnormalities were observed in conditional PdgfrαCre;Tbx1+/flox mice regarding neonatal ultrasonic vocalizations, social interaction, novel object approach, anxiety-like behavior (elevated plus maze), or open-field locomotion and thigmotaxis. Electron microscopic analysis revealed a compositional shift in myelinated axons within the fimbria of adult male mutants, characterized by an increased number of myelinated axons in the 300–800 nm diameter range and a decreased number in the ∼1,200 nm and ∼1,400 nm ranges, with myelin thickness remaining unchanged across diameters. These findings indicate that Tbx1 heterozygosity in the oligodendrocyte lineage leads to a selective shift towards smaller myelinated axons in the fimbria and a transiently higher level of capacity for working memory and cognitive flexibility. However, it does not replicate the full spectrum of myelination abnormalities or the broader cognitive and social deficits observed in constitutive Tbx1 heterozygotes, suggesting that Tbx1 deficiency in non-oligodendrocyte lineage cells may lead to altered myelination and neurodevelopmental behavioral impairments. Competing Interest Statement The authors have declared no competing interest. Footnotes AM Wells: wellsa1{at}livemail.uthscsa.edu T Tanifuji: kjh.goodby{at}gmail.com T. Takano: takanot{at}uthscsa.edu A. Endo: endo{at}uthscsa.edu G. Kang: kangg{at}uthscsa.edu M. Esparza: esparzam1{at}uthscsa.edu Q. Shi: shiQ{at}uthscsa.edu M. Bhat: bhatm{at}uthscsa.edu N. Hiroi: hiroi{at}uthscsa.edu The text responded to reviewers' comments. List of abbreviations - CNVs - Copy number variations - ASD - Autism Spectrum disorder - scRNA-seq - single cell RNA-seq - Pdgfra - Platelet-derived growth factor receptor alpha - MAG - Myelin associated glycoprotein - MOG - Myelin oligodendrocyte glycoprotein, - MBP - myelin basic protein