Kussick Emily, Johansen Nelson, Taskin Naz, Chowdhury Ananya, Quinlan Meagan A, Fraser Alex, Clark Andrew G, Wynalda Brooke, Martinez Refugio, Groce Erin L, Reding Melissa, Liang Elizabeth, Shulga Lyudmila, Huang Cindy, Casper Tamara, Clark Michael, Ho Windy, Gao Yuan, van Velthoven Cindy T J, Sobieski Cassandra, Ferrer Rebecca, Berg Melissa R, Curtis Britni C, English Chris, Day Jesse C, Fortuna Michal G, Donadio Nicholas, Newman Dakota, Yao Shenqin, Chakka Anish Bhaswanth, Goldy Jeff, Torkelson Amy, Guzman Junitta B, Chakrabarty Rushil, Nguy Beagen, Guilford Nathan, Pham Trangthanh H, Wright Vonn, Ronellenfitch Kara, Naidoo Robyn, Kenney Jaimie, Williford Ali, Ramakrishnan Charu, Drinnenberg Antonia, Gudsnuk Kathryn, Thyagarajan Bargavi, Smith Kimberly A, Dee Nick, Deisseroth Karl, Zeng Hongkui, Yao Zizhen, Tasic Bosiljka, Levi Boaz P, Hodge Rebecca, Bakken Trygve E, Lein Ed S, Ting Jonathan T, Daigle Tanya L
Allen Institute for Brain Science, Seattle, WA 98109, USA.
Washington National Primate Research Center, Seattle, WA 98195, USA.
Cell Rep. 2025 May 20:115730. doi: 10.1016/j.celrep.2025.115730.
Experimental access to cell types within the mammalian spinal cord is severely limited by the availability of genetic tools. To enable access to spinal motor neurons (SMNs) and SMN subtypes, we generated single-cell multiome datasets from mouse and macaque spinal cords and discovered putative enhancers for each neuronal population. We cloned these enhancers into adeno-associated viral vectors driving a reporter fluorophore and functionally screened them in the mouse. We extensively characterized the most promising candidate enhancers in rat and macaque and developed an optimized pan-SMN enhancer virus. Additionally, we generated derivative viruses expressing iCre297T recombinase or ChR2-EYFP for labeling and functional studies, and we created a single vector with combined enhancer elements to achieve simultaneous labeling of layer 5 extratelencephalic projecting neurons and SMNs. This unprecedented SMN toolkit will enable future investigations of cell type function across species and potential therapeutic interventions for human neurodegenerative diseases.
由于遗传工具的可用性，对哺乳动物脊髓内细胞类型的实验性研究受到严重限制。为了能够研究脊髓运动神经元（SMN）及其亚型，我们从小鼠和猕猴的脊髓中生成了单细胞多组学数据集，并发现了每个神经元群体的假定增强子。我们将这些增强子克隆到驱动报告荧光团的腺相关病毒载体中，并在小鼠中进行功能筛选。我们对大鼠和猕猴中最有前景的候选增强子进行了广泛表征，并开发了一种优化的泛SMN增强子病毒。此外，我们还生成了表达iCre297T重组酶或ChR2-EYFP的衍生病毒用于标记和功能研究，并创建了一个具有组合增强子元件的单一载体，以实现对第5层脑外投射神经元和SMN的同时标记。这个前所未有的SMN工具包将使未来能够跨物种研究细胞类型功能，并为人类神经退行性疾病开展潜在的治疗干预。
