The ERK signaling drives evolutionary expansion of the mammalian cerebral cortex

The molecular basis for cortical expansion during evolution remains largely unknown. Here, we found that FGF-ERK signaling promotes the self-renewal and expansion of cortical radial glial (RG) cells. Furthermore, FGF-ERK signaling induces Bmp7 expression in cortical RG cells, which increases the length of the neurogenic period. We demonstrate that ERK signaling and SHH signaling mutually inhibit each other in cortical RG cells. We also provide evidence that ERK signaling is elevated in cortical RG cells during development and evolution. We conclude that the evolutionary expansion of the human cortex is driven by ERK-BMP7-GLI3R signaling pathway in cortical RG cells, which participates in a positive feedback loop through antagonizing SHH signaling. We also propose that the molecular basis for cortical evolutionary dwarfism, exemplified by the lissencephalic mouse which originated from a larger and gyrencephalic ancestor, is due to mouse cortical RG cells receiving higher SHH signaling that antagonizes ERK signaling. The whole mouse cortex at E14.5 was dissected out under a microscope for scRNA-Seq analysis and bulk RNA-Seq analysis. Briefly, mouse embryos were dissected out and immediately submerged in fresh ice-cold Hanks balanced salt solution (Gibco 12175-095). The cortex was then cut into pieces and dissociated into a single-cell suspension using a Papain Cell Dissociation Kit (Miltenyi Biotec, catalog no. 130-092-628) according to the manufacturer's instructions.进化过程中大脑皮层扩张的分子基础在很大程度上仍未明确。本研究发现，成纤维细胞生长因子-细胞外调节蛋白激酶（FGF-ERK）信号通路可促进皮层放射状胶质细胞（radial glial, RG）的自我更新与扩增。进一步研究表明，该信号通路可诱导皮层RG细胞表达骨形态发生蛋白7（Bmp7），进而延长神经发生期时长。本研究证实，ERK信号通路与音猬因子（SHH）信号通路在皮层RG细胞中存在相互抑制作用。同时，我们提供证据显示，在发育与进化过程中，皮层RG细胞内的ERK信号通路活性均有所升高。本研究得出结论：人类大脑皮层的进化扩张由皮层RG细胞内的ERK-BMP7-GLI3R信号通路驱动，该通路通过拮抗SHH信号通路参与形成正反馈环路。此外，我们提出皮层进化性侏儒症的分子基础——以源自更大体积且具有脑沟回的祖先的无脑回小鼠为例——源于小鼠皮层RG细胞接收到更高水平的SHH信号，进而拮抗ERK信号通路。我们在显微镜下剥离胚胎发育第14.5天（E14.5）的完整小鼠皮层，用于单细胞RNA测序（single-cell RNA sequencing, scRNA-Seq）与批量RNA测序（bulk RNA-Seq）分析。简言之，我们剥离小鼠胚胎后立即将其浸没于新鲜预冷的汉克斯平衡盐溶液（Gibco 12175-095）中，随后将皮层剪碎，按照制造商说明书使用木瓜蛋白酶细胞解离试剂盒（Miltenyi Biotec，货号130-092-628）制备单细胞悬液。