Abstract：
GREMLIN1 (GREM1) is a secreted protein that antagonizes bone morphogenetic proteins (BMPs). While abnormal GREM1 expression has been reported to cause behavioral defects in postpartum mice, the spatial and cellular distribution of GREM1 in the brain and the influence of the GREM1-secreting cells on brain function and behavior remain unclear. To address this, we designed a genetic cassette incorporating a 3×Flag-TeV-HA-T2A-tdTomato sequence, resulting in the creation of a novel Grem1Tag mouse model, expressing an epitope tag (3×Flag-TeV-HA-T2A) followed by a fluorescent reporter (tdTomato) under the control of the endogenous Grem1 promoter. This design facilitated precise tracking of the cell origin and distribution of GREM1 in the brain using tdTomato and Flag (or HA) markers, respectively. We confirmed that the Grem1Tag mouse exhibited normal motor, cognitive, and social behaviors at postnatal 60 days (P60), compared with C57BL/6J controls. Through immunofluorescence staining, we comprehensively mapped the distribution of GREM1-secreting cells across the central nervous system. Pervasive GREM1 expression was observed in the cerebral cortex (Cx), medulla, pons, and cerebellum, with the highest levels in the Cx region. Notably, within the Cx, GREM1 was predominantly secreted by excitatory neurons, particularly those expressing calcium/calmodulin-dependent protein kinase II alpha (Camk2a), while inhibitory neurons (parvalbumin-positive, PV+) and glial cells (oligodendrocytes, astrocytes, and microglia) showed little or no GREM1 expression. To delineate the functional significance of GREM1-secreting cells, a selective ablation at P42 using a diphtheria toxin A (DTA) system resulted in increased anxiety-like behavior and impaired memory in mice. Altogether, our study harnessing the Grem1Tag mouse model reveals the spatial and cellular localization of GREM1 in the mouse brain, shedding light on the involvement of GREM1-secreting cells in modulating brain function and behavior. Our Grem1Tag mouse serves as a valuable tool for further exploring the precise role of GREM1 in brain development and disease.
摘要：
Gremlin1（Grem1）是一种分泌蛋白，可拮抗骨形态发生蛋白（BMP）。据报道，异常的Grem1表达会导致产后小鼠的行为缺陷，大脑中GREM1的空间和细胞分布以及Grem1分泌细胞对脑功能和行为的影响尚不清楚。为了解决这个问题，我们设计了一个包含3×Flag-TeV-HA-T2A-td番茄序列的基因盒，从而创造了一个新的Grem1Tag小鼠模型，表达表位标签（3×Flag-TeV-HA-T2A），然后在内源Grem1启动子的控制下表达荧光报告分子（tdTomato）。这种设计有助于使用tdTomato和Flag（或HA）标记物精确跟踪GREM1在大脑中的细胞起源和分布。分别。我们确认Grem1Tag鼠标表现出正常的运动，认知，和产后60天的社会行为（P60），与C57BL/6J对照相比。通过免疫荧光染色，我们全面绘制了中枢神经系统中Grem1分泌细胞的分布图。在大脑皮层（Cx）中观察到普遍的Grem1表达，髓质,pons,还有小脑,Cx地区的最高水平。值得注意的是,在Cx内，GREM1主要由兴奋性神经元分泌，特别是那些表达钙/钙调蛋白依赖性蛋白激酶IIα（Camk2a），而抑制性神经元（小白蛋白阳性，PV+）和神经胶质细胞（少突胶质细胞，星形胶质细胞,和小胶质细胞）显示很少或没有Grem1表达。为了描述Grem1分泌细胞的功能意义，使用白喉毒素A(DTA)系统选择性消融P42导致小鼠焦虑样行为增加和记忆受损.总之,我们利用Grem1Tag小鼠模型的研究揭示了GREM1在小鼠大脑中的空间和细胞定位，揭示Grem1分泌细胞参与调节脑功能和行为的作用。我们的Grem1Tag鼠标是进一步探索Grem1在大脑发育和疾病中的精确作用的宝贵工具。
