Kirk Ryan W, Sun Liwei, Xiao Ruixuan, Clark Erin A, Nelson Sacha
Department of Biology, Brandeis University, Waltham, MA 02453, USA.
bioRxiv. 2025 Feb 15:2025.02.07.636951. doi: 10.1101/2025.02.07.636951.
A critical phase of mammalian brain development takes place after birth. Neurons of the mouse neocortex undergo dramatic changes in their morphology, physiology, and synaptic connections during the first postnatal month, while properties of immature neurons, such as the capacity for robust axon outgrowth, are lost. The genetic and epigenetic programs controlling prenatal development are well studied, but our understanding of the transcriptional mechanisms that regulate postnatal neuronal maturation is comparatively lacking. By integrating chromatin accessibility and gene expression data from two subtypes of neocortical pyramidal neurons in the neonatal and maturing brain, we predicted a role for the Krüppel-Like Factor (KLF) family of Transcription Factors in the developmental regulation of neonatally expressed genes. Using a multiplexed CRISPR Interference (CRISPRi) knockdown strategy, we found that a shift in expression from KLF activators (Klf6, Klf7) to repressors (Klf9, Klf13) during early postnatal development functions as a transcriptional 'switch' to first activate, then repress a set of shared targets with cytoskeletal functions including and . We demonstrate that this switch is buffered by redundancy between KLF paralogs, which our multiplexed CRISPRi strategy is equipped to overcome and study. Our results indicate that competition between activators and repressors within the KLF family regulates a conserved component of the postnatal maturation program that may underlie the loss of intrinsic axon growth in maturing neurons. This could facilitate the transition from axon growth to synaptic refinement required to stabilize mature circuits.
哺乳动物大脑发育的一个关键阶段发生在出生后。小鼠新皮质的神经元在出生后的第一个月内，其形态、生理和突触连接会发生巨大变化，而未成熟神经元的特性，如强大的轴突生长能力，则会丧失。控制产前发育的遗传和表观遗传程序已得到充分研究，但我们对调节出生后神经元成熟的转录机制的了解相对较少。通过整合来自新生和成熟大脑中两种新皮质锥体神经元亚型的染色质可及性和基因表达数据，我们预测了转录因子Krüppel样因子（KLF）家族在新生儿表达基因的发育调控中的作用。使用多重CRISPR干扰（CRISPRi）敲低策略，我们发现出生后早期发育过程中从KLF激活因子（Klf6、Klf7）到抑制因子（Klf9、Klf13）的表达转变，起到了转录“开关”的作用，先激活然后抑制一组具有细胞骨架功能的共同靶点，包括 和 。我们证明，这种开关由KLF旁系同源物之间的冗余所缓冲，而我们的多重CRISPRi策略能够克服并研究这种冗余。我们的结果表明，KLF家族内激活因子和抑制因子之间的竞争调节了出生后成熟程序的一个保守组成部分，这可能是成熟神经元内在轴突生长丧失的基础。这可能有助于从轴突生长向稳定成熟回路所需的突触细化的转变。
