Background RAC3 encodes a Rho family small GTPase that regulates the behaviour and organisation of actin cytoskeleton and intracellular signal transduction. Variants in RAC3 can cause a phenotypically heterogeneous neurodevelopmental disorder with structural brain anomalies and dysmorphic facies. The pathomechanism of this recently discovered genetic disorder remains unclear.
Methods We investigated an early adolescent female with intellectual disability, drug-responsive epilepsy and white matter abnormalities. Through exome sequencing, we identified the novel de novo variant (NM_005052.3): c.83T>C (p.Phe28Ser) in RAC3. We then examined the pathophysiological significance of the p.F28S variant in comparison with the recently reported disease-causing p.Q61L variant, which results in a constitutively activated version of RAC3.
Results In vitro analyses revealed that the p.F28S variant was spontaneously activated by substantially increased intrinsic GTP/GDP-exchange activity and bound to downstream effectors tested, such as PAK1 and MLK2. The variant suppressed the differentiation of primary cultured hippocampal neurons and caused cell rounding with lamellipodia. In vivo analyses using in utero electroporation showed that acute expression of the p.F28S variant caused migration defects of excitatory neurons and axon growth delay during corticogenesis. Notably, defective migration was rescued by a dominant negative version of PAK1 but not MLK2.
Conclusion Our results indicate that RAC3 is critical for brain development and the p.F28S variant causes morphological and functional defects in cortical neurons, likely due to the hyperactivation of PAK1.
- central nervous system diseases
- mutation, missense
- gain of function mutation
Data availability statement
Data are available on reasonable request.
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MN, MS and MU contributed equally.
Correction notice Since this article was first published, three authors have been listed as equal contributors.
Contributors MN and K-iN designed the study. MN, HI and K-iN performed the experiments and functional data analysis. K-iN was responsible for overseeing data collection and analysis. MS, MU, AW, PS, FZ, GC and VC took care of clinical data collection and analysis. MN and MS conceptualised, drafted and revised the manuscript under the supervision of K-iN. All authors participated in manuscript revision and approval of the final manuscript. MN, MS and K-iN are the guarantors of the manuscript.
Funding This work was supported in part by Japan Society for the Promotion of Science (JSPS) KAKENHI Grant-in-Aid for Scientific Research (B) (Grant Number JP19H03629), Grant-in-Aid for Scientific Research (C) (Grant Number JP19K07059), Grant-in-Aid for Research Activity Start-up (Grant Number JP20K22888), Grant-in-Aid for Early-Career Scientists (Grant Number JP21K15895), and a grant-in-aid of the Practical Research Project for Rare/Intractable Diseases from Japan Agency for Medical Research and Development (AMED) (15ek0109040h0002).
Competing interests None declared.
Provenance and peer review Not commissioned; externally peer reviewed.
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