Precise formation of complex neural circuits in the spinal cord,achieved through the integration of diverse neuronal populations,is essential for central nervous system function.However,the specialization and migratio...Precise formation of complex neural circuits in the spinal cord,achieved through the integration of diverse neuronal populations,is essential for central nervous system function.However,the specialization and migration of human spinal cord neurons remain poorly understood.In this study,we perform single-cell transcriptome sequencing of human spinal cord from Carnegie Stages(CS)16–21 and mouse spinal cord from embryonic day(E)8.0–11.5,complemented by in situ sequencing of human spinal cord(CS 16–20).Our results reveal the critical role of the precursor state in neuronal differentiation and migration,identifying key transcription factors that regulate these processes across species.Notably,each neuronal lineage expresses unique markers as early as the progenitor stage at the spinal cord midline,and subsequently undergoes a shared transcriptional program during precursor commitment that guides migration.This synchronized migration,validated by spatial transcriptomics,occurs in both dorsal and ventral regions.Our findings offer important insights into the migration patterns and regulatory factors that guide spinal cord neuron subtype specification during embryogenesis.展开更多
Spatial transcriptomics enables the study of localization-indexed gene expression activity in tissues,providing the transcriptional landscape that in turn indicates the potential regulatory networks of gene expression...Spatial transcriptomics enables the study of localization-indexed gene expression activity in tissues,providing the transcriptional landscape that in turn indicates the potential regulatory networks of gene expression.In situ sequencing(ISS)is a targeted spatial transcriptomic technique,based on padlock probe and rolling circle amplification combined with next-generation sequencing chemistry,for highly multiplexed in situ gene expression profiling.Here,we present improved in situ sequencing(IISS)that exploits a new probing and barcoding approach,combined with advanced image analysis pipelines for high-resolution targeted spatial gene expression profiling.We develop an improved combinatorial probe anchor ligation chemistry using a 2-base encoding strategy for barcode interrogation.The new encoding strategy results in higher signal intensity as well as improved specificity for in situ sequencing,while maintaining a streamlined analysis pipeline for targeted spatial transcriptomics.We show that IISS can be applied to both fresh frozen tissue and formalin-fixed paraffin-embedded tissue sections for single-cell level spatial gene expression analysis,based on which the developmental trajectory and cell-cell communication networks can also be constructed.展开更多
基金supported by the National Key R&D Program of China (2024YFA1802300 and 2023YFF1204701)the National Natural Science Foundation of China (32225012 and 32200662)+3 种基金Major Project of Guangzhou National Laboratory (GZNL2023A02005)Pearl River Talent Recruitment Program (2021ZT09Y233)Science and Technology Planning Project of Guangdong Province,China(2023B1212060050 and 2023B1212120009)Health@Inno HK Program launched by Innovation Technology Commission of the Hong Kong SAR,P.R. China.
文摘Precise formation of complex neural circuits in the spinal cord,achieved through the integration of diverse neuronal populations,is essential for central nervous system function.However,the specialization and migration of human spinal cord neurons remain poorly understood.In this study,we perform single-cell transcriptome sequencing of human spinal cord from Carnegie Stages(CS)16–21 and mouse spinal cord from embryonic day(E)8.0–11.5,complemented by in situ sequencing of human spinal cord(CS 16–20).Our results reveal the critical role of the precursor state in neuronal differentiation and migration,identifying key transcription factors that regulate these processes across species.Notably,each neuronal lineage expresses unique markers as early as the progenitor stage at the spinal cord midline,and subsequently undergoes a shared transcriptional program during precursor commitment that guides migration.This synchronized migration,validated by spatial transcriptomics,occurs in both dorsal and ventral regions.Our findings offer important insights into the migration patterns and regulatory factors that guide spinal cord neuron subtype specification during embryogenesis.
基金supported by the Natural Science Foundation of Fujian Province(2022J06022)the Quanzhou Science and Technology Plan Project(2021C040R)the Scientific Research Funds of Huaqiao University.
文摘Spatial transcriptomics enables the study of localization-indexed gene expression activity in tissues,providing the transcriptional landscape that in turn indicates the potential regulatory networks of gene expression.In situ sequencing(ISS)is a targeted spatial transcriptomic technique,based on padlock probe and rolling circle amplification combined with next-generation sequencing chemistry,for highly multiplexed in situ gene expression profiling.Here,we present improved in situ sequencing(IISS)that exploits a new probing and barcoding approach,combined with advanced image analysis pipelines for high-resolution targeted spatial gene expression profiling.We develop an improved combinatorial probe anchor ligation chemistry using a 2-base encoding strategy for barcode interrogation.The new encoding strategy results in higher signal intensity as well as improved specificity for in situ sequencing,while maintaining a streamlined analysis pipeline for targeted spatial transcriptomics.We show that IISS can be applied to both fresh frozen tissue and formalin-fixed paraffin-embedded tissue sections for single-cell level spatial gene expression analysis,based on which the developmental trajectory and cell-cell communication networks can also be constructed.
