Satb2 and Nr4a2 are required for the differentiation of cortical layer 6b.

Cortical layer 6 is divided into two sublayers, and layer 6b is situated above the white matter with distinct architecture from layer 6a. Layer 6b arises from the subplate and contains the earliest born neurons in the development of cerebral cortex. Although great progress has been made in understanding the cortical morphogenesis, there is a dearth of knowledge regarding the molecular mechanisms governing the development of layer 6b neurons. Here we report that transcription factor special AT-rich binding protein 2 (Satb2) and nuclear receptor subfamily 4 group A member 2 (Nr4a2) are required for the normal differentiation layer 6b neurons. Upon conditional deletion of Satb2 in the cortex (Satb2(Emx1) CKO) or selectively inactivation of Satb2 in layer 6b neurons only (Satb2(Nr4a2CreER) CKO), the expressions of layer 6b-specific genes (i.e., Ctgf, Cplx3, Trh and Tnmd) were significantly reduced, whereas that of Nr4a2 was dramatically increased, underscoring that Satb2 is involved in the differentiation of layer 6b neurons in a cell-autonomous manner. On the other hand, when Nr4a2 was deleted in the cortex, the expressions of Trh and Tnmd were upregulated with unchanged expression of Ctgf and Cplx3. Notably, the defective differentiation resulting from the deletion of Satb2 remained in Satb2/Nr4a2 double CKO mice. In summary, our findings indicated that both Satb2 and Nr4a2 are required for the differentiation of layer 6b neurons possibly via different pathways.