Rac signaling impacts a relatively large number of downstream targets; however, few studies have established an association between Rac pathways and pathological conditions. In the present study, we generated mice with double knockout of Rac1 and Rac3 (Atoh1-Cre;Rac1flox/flox;Rac3−/−) in cerebellar granule neurons (CGNs). We observed impaired tangential migration at E16.5, as well as numerous apoptotic CGNs at the deepest layer of the external granule layer (EGL) in the medial cerebellum of Atoh1-Cre;Rac1flox/flox;Rac3−/− mice at P8. Atoh1-Cre;Rac1flox/flox;Rac3−/− CGNs differentiated normally until expression of p27kip1 and NeuN in the deep EGL at P5. Primary CGNs and cerebellar microexplants from Atoh1-Cre;Rac1flox/flox;Rac3−/− mice exhibited impaired neuritogenesis, which was more apparent in Map2-positive dendrites. Such findings suggest that impaired tangential migration and final differentiation of CGNs have resulted in decreased cerebellum size and agenesis of the medial internal granule layer, respectively. Furthermore, Rac depleted/deleted cells exhibited decreased levels of Mid1 and impaired mTORC1 signaling. Mid1 depletion in CGNs produced mild impairments in neuritogenesis and reductions in mTORC1 signaling. Thus, a novel Rac-signaling pathway (Rac1-Mid1-mTORC1) may be involved in medial cerebellar development.
The authors declare no competing or financial interests.
Conceptualization: T.U.; Methodology: T.U.; Formal analysis: T.N., T.U., Y.N.; Investigation: T.N., T.U., Y.N., H.S., N.C., Y. Hishikawa, H.K., M.K., E.K., Y. Hisa; Resources: H.K., M.S., I.d.C., A.A.; Data curation: T.U.; Writing - original draft: T.N., T.U.; Writing - review & editing: T.U.; Visualization: T.U.; Supervision: T.U., H.S., E.K., Y. Hisa, N.S.; Project administration: T.U., N.S.; Funding acquisition: T.U., N.S.
This work was supported by grants from the Japan Society for the Promotion of Science KAKENHI on Innovative Areas ‘Fluorescence Live imaging’ (to N.S.); the Japan Society for the Promotion of Science KAKENHI [17H04042 (to T.U.) and 17024038 (to A.A.)]; the Uehara Memorial Foundation (201320273 to T.U.); and the Hyogo Science and Technology Association (26087 to T.U.).
DNA microarray data are available in the ArrayExpress database (www.ebi.ac.uk/arrayexpress) under accession number E-MTAB-5681.
Supplementary information available online at http://dev.biologists.org/lookup/doi/10.1242/dev.147900.supplemental
- Received December 7, 2016.
- Accepted March 31, 2017.