Data_Sheet_2_Smaller Body Size, Early Postnatal Lethality, and Cortical Extracellular Matrix-Related Gene Expression Changes of Cyfip2-Null Embryonic Mice.xlsx

Cytoplasmic FMR1-interacting protein 2 (CYFIP2) is a key component of the WAVE regulatory complex (WRC) which regulates actin polymerization and branching in diverse cellular compartments. Recent whole exome sequencing studies identified de novo hotspot variants in CYFIP2 from patients with early-onset epileptic encephalopathy and microcephaly, suggesting that CYFIP2 may have some functions in embryonic brain development. Although perinatal lethality of Cyfip2-null (Cyfip2−/−) mice was reported, the exact developmental time point and cause of lethality, and whether Cyfip2−/− embryonic mice have brain abnormalities remain unknown. We found that endogenous Cyfip2 is mainly expressed in the brain, spinal cord, and thymus of mice at late embryonic stages. Cyfip2−/− embryos did not show lethality at embryonic day 18.5 (E18.5), but their body size was smaller than that of wild-type (WT) or Cyfip2+/− littermates. Meanwhile, at postnatal day 0, all identified Cyfip2−/− mice were found dead, suggesting early postnatal lethality of the mice. Nevertheless, the brain size and cortical cytoarchitecture were comparable among WT, Cyfip2+/−, and Cyfip2−/− mice at E18.5. Using RNA-sequencing analyses, we identified 98 and 72 differentially expressed genes (DEGs) from the E18.5 cortex of Cyfip2+/− and Cyfip2−/− mice, respectively. Further bioinformatic analyses suggested that extracellular matrix (ECM)-related gene expression changes in Cyfip2−/− embryonic cortex. Together, our results suggest that CYFIP2 is critical for embryonic body growth and for early postnatal survival, and that loss of its expression leads to ECM-related gene expression changes in the embryonic cortex without severe gross morphological defects.细胞质FMR1互作蛋白2（CYFIP2）是WAVE调控复合物（WRC）的关键组成成分，该复合物可调控多种细胞区域内的肌动蛋白聚合与分支过程。近期全外显子组测序研究在早发性癫痫性脑病合并小头畸形患者样本中，鉴定出CYFIP2存在新发热点变异，提示CYFIP2可能在胚胎脑发育中发挥特定功能。尽管已有研究报道Cyfip2基因敲除（Cyfip2−/−）小鼠存在围产期致死现象，但该类小鼠的确切致死发育时间点、致死原因，以及Cyfip2−/−胚胎小鼠是否存在脑异常仍未明确。本研究发现，在小鼠胚胎发育晚期，内源性Cyfip2主要表达于脑、脊髓与胸腺组织。Cyfip2−/−胚胎在胚胎第18.5天（E18.5）时未出现致死现象，但体型较野生型（WT）或Cyfip2杂合子（Cyfip2+/−）同窝小鼠更小。与此同时，在出生后第0天，所有检测到的Cyfip2−/−小鼠均已死亡，提示该类小鼠存在出生早期致死表型。尽管如此，在E18.5阶段，野生型、Cyfip2+/−与Cyfip2−/−小鼠的脑体积与大脑皮层细胞构筑均无明显差异。通过RNA测序分析，本研究分别从Cyfip2+/−与Cyfip2−/−小鼠的E18.5皮层组织中鉴定出98个和72个差异表达基因（DEGs）。进一步的生物信息学分析显示，Cyfip2−/−胚胎皮层中存在与细胞外基质（ECM）相关的基因表达谱改变。综上，本研究结果表明CYFIP2对胚胎躯体生长以及出生早期存活至关重要，其表达缺失会导致胚胎皮层出现与ECM相关的基因表达改变，但不会引发严重的大体形态缺陷。