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The subcommissural organ regulates brain development via secreted peptides

Abstract

The subcommissural organ (SCO) is a gland located at the entrance of the aqueduct of Sylvius in the brain. It exists in species as distantly related as amphioxus and humans, but its function is largely unknown. Here, to explore its function, we compared transcriptomes of SCO and non-SCO brain regions and found three genes, Sspo, Car3 and Spdef, that are highly expressed in the SCO. Mouse strains expressing Cre recombinase from endogenous promoter/enhancer elements of these genes were used to genetically ablate SCO cells during embryonic development, resulting in severe hydrocephalus and defects in neuronal migration and development of neuronal axons and dendrites. Unbiased peptidomic analysis revealed enrichment of three SCO-derived peptides, namely, thymosin beta 4, thymosin beta 10 and NP24, and their reintroduction into SCO-ablated brain ventricles substantially rescued developmental defects. Together, these data identify a critical role for the SCO in brain development.

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Fig. 1: Identifying genes expressed specifically in SCO cells and developing corresponding Cre/CreER mouse lines.
Fig. 2: Specificity of Sspo-CreER and Sspo-Cre mouse strains for SCO labeling.
Fig. 3: Phenotypes observed in SCO-ablated mice.
Fig. 4: Neuronal properties of the mouse brain after SCO ablation.
Fig. 5: SCO promotes neuronal development.
Fig. 6: Detection of neuropeptides in SCO and their release into CSF.
Fig. 7: SCO-secreted peptides promote neuronal development in vitro.
Fig. 8: SCO-secreted peptides rescue phenotypes in SCO-ablated mice.

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Data availability

All research materials and data are publicly available. New Cre mouse strains are in the process of being donated to the Jackson Laboratory mouse repository for distribution. RNA-seq are publicly available from NCBI GEO (accession numbers GSE214744 and GSE226349) and peptidomic data from IPROX (accession numbers 15E4 and U0Jm). RNA-seq results: these data have been deposited in the publicly available NCBI GEO database and will be released to the public upon publication. The accession information is as follows: SCO bulk RNA-seq (https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE214744), secure token ivatogiazvabpoj; RNA-seq of neurons with peptide incubation (https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE226349), secure token yfkvcygwxbkrvyr; peptideomics data (https://www.iprox.cn/page/PSV023.html;?url=1713509741810ndGU), accession number 15E4; proteomics data (https://www.iprox.cn/page/PSV023.html;?url=1697721703737jgBn), accession number U0Jm.

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Acknowledgements

We thank lab members in Ge laboratories, B. Samuels and T. Taylor for their feedback and critical reading of the manuscript. We thank Q. Guo, X. Gao and M. Jia in the imaging core of CIBR for assistance with imaging and data analysis. We thank W. Li, S. Huang and R. Shen at the animal core facility for assistance with animal care and purchasing. We thank J. Chen and X. Zhang at the Genomics Center for sequencing and fluorescence-activated cell sorting. We thank the MS facility of the Phoenix Center for use of their instrumentation. We thank F. Huang for suggestions. This work was supported in part by grants from the STI2030-Major Projects (2022ZD0204700), the Natural Science Foundation of China and Beijing Scholars to W.G. (32170964); startup funds from CIBR, the National Key R&D Program of China to C.J. (no. 2021YFA1302601); and a grant from the US National Institutes of Health (NIH, R01DK071801) to L.L. The Orbitrap instruments were purchased through support from an NIH shared instrument grant (NIH-NCRR S10RR029531).

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Contributions

W.G. conceived and supervised the project. T.Z., D.A., P.W., L.L., C.J., W.S. and W.G. designed experiments. T.Z. performed most experiments, including mouse genetics, cell ablation, neuronal culture, immunostaining, imaging, RNAscope mFISH, mouse breeding and genotyping, CSF collection and rescue experiments, among others. P.W., Y.X., F.M., Y.Z., T.Z. and C.J. performed peptidomics and proteomics and data analysis. W.G., Z.Z., T.T. and X. Zhang performed SCO bulk RNA-seq; C.X., D.A. and W.G. analyzed the RNA-seq data. W.G. and T.Z. determined the genes for genetic mouse strains. T.Z. and J.L. assisted in MRI data acquisition and analysis, T.T. assisted in MRI imaging, Z.B. and T.Z. performed AAV viral labeling and peptide injection and assisted in slice imaging. X.-J.C., J.-L.L., J.Y. and L. Zheng assisted in brain slice imaging and electrophysiology. F.L. assisted in neuronal culture. M.Y. assisted in RNA-seq. C.L. assisted in electron microscopy. X. Zou and Z.F. assisted in mouse breeding, genotyping and immunostaining. Z.G. assisted in CSF collection. W.G., W.S., L.L., B.L., C.J., T.H., Z.L., L. Zhang, H. Zhang and H. Zeng provided reagents. T.Z. and W.G. wrote the manuscript. All authors discussed, reviewed and edited the manuscript.

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Correspondence to Lingjun Li, Wenzhi Sun or Woo-ping Ge.

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Zhang, T., Ai, D., Wei, P. et al. The subcommissural organ regulates brain development via secreted peptides. Nat Neurosci 27, 1103–1115 (2024). https://doi.org/10.1038/s41593-024-01639-x

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