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TATTOO-seq delineates spatial and cell type-specific regulatory programs in the developing limb

ABSTRACT: The coordinated differentiation of progenitor cells into specialized cell types and their spatial organization into distinct domains is central to embryogenesis. Here, we applied a new unbiased spatially resolved single-cell transcriptomics method to identify the genetic programs underlying the emergence of specialized cell types during limb development and their spatial integration. We identify multiple transcription factors whose expression patterns are predominantly associated with cell type specification or spatial position, suggesting two parallel yet highly interconnected regulatory systems. We demonstrate that the embryonic limb undergoes a complex multi-scale re-organization upon perturbation of one of its spatial organizing centers, including the loss of specific cell populations, specific alterations of pre-existing cell states’ molecular identities and changes in their relative spatial distribution. Our study shows how multi-dimensional single-cell, spatially resolved molecular atlases can allow the deconvolution of spatial identity and cell fate and reveal the interconnected genetic networks that regulate organogenesis and its reorganization upon genetic alterations.

ORGANISM(S): Mus musculus

PROVIDER: GSE202326 | GEO | 2022/11/06

REPOSITORIES: GEO

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Project description:Spatial organization of the transcriptome has emerged as a powerful means for regulating the post-transcriptional fate of RNA in eukaryotes; however, whether prokaryotes use RNA spatial organization as a mechanism for post-transcriptional regulation remains unclear. Here we used super-resolution microscopy to image the E. coli transcriptome and observed a genome-wide spatial organization of RNA: mRNAs encoding inner-membrane proteins are enriched at the membrane, whereas mRNAs encoding outer-membrane, cytoplasmic and periplasmic proteins are distributed throughout the cytoplasm. Membrane enrichment is caused by co-translational insertion of signal peptides recognized by the signal-recognition particle. Our time-resolved RNA-sequencing and live-cell super-resolution imaging experiments revealed a physiological consequence of this spatial organization and the underlying mechanism: membrane localization enhances degradation rates of inner-membrane-protein mRNAs by placing them in proximity to membrane-bound RNA degradation enzymes. Together, these results demonstrate that the bacterial transcriptome is spatially organized and that this organization shapes the posttranscriptional Spatial organization of the transcriptome has emerged as a powerful means for regulating the post-transcriptional fate of RNA in eukaryotes; however, whether prokaryotes use RNA spatial organization as a mechanism for post-transcriptional regulation remains unclear. Here we used super-resolution microscopy to image the E. coli transcriptome and observed a genome-wide spatial organization of RNA: mRNAs encoding inner-membrane proteins are enriched at the membrane, whereas mRNAs encoding outer-membrane, cytoplasmic and periplasmic proteins are distributed throughout the cytoplasm. Membrane enrichment is caused by co-translational insertion of signal peptides recognized by the signal-recognition particle. Our time-resolved RNA-sequencing and live-cell super-resolution imaging experiments revealed a physiological consequence of this spatial organization and the underlying mechanism: membrane localization enhances degradation rates of inner-membrane-protein mRNAs by placing them in proximity to membrane-bound RNA degradation enzymes. Together, these results demonstrate that the bacterial transcriptome is spatially organized and that this organization shapes the post-transcriptional dynamics of mRNAs.

Project description:A fundamental question in biology is how an undifferentiated field of cells acquires spatial pattern and undergoes coordinated differentiation. The development of the vertebrate limb is an important paradigm for understanding these processes. The skeletal and connective tissues of the developing limb all derive from a population of multipotent progenitor cells located in its distal tip. During limb outgrowth, these progenitors segregate into a chondrogenic lineage, located in the center of the limb bud, and soft connective tissue lineages located in its periphery. We report that the interplay of two families of signaling proteins, fibroblast growth factors (FGFs) and Wnts, coordinate the growth of the multipotent progenitor cells with their simultaneous segregation into these lineages. FGF and Wnt signals act together to synergistically promote proliferation while maintaining the cells in an undifferentiated, multipotent state, but act separately to determine cell lineage specification. Withdrawal of both signals results in cell cycle withdrawal and chondrogenic differentiation. Continued exposure to Wnt however, maintains proliferation and re-specifies the cells towards the soft connective tissue lineages. We have identified target genes that are synergistically regulated by Wnts and Fgfs, and show how these factors actively suppress differentiation and promote growth. Finally, we show how the spatial restriction of Wnt and FGF signals to the limb ectoderm and to a specialized region of it, the apical ectodermal ridge, controls the distribution of cell behaviors within the growing limb, and guides the proper spatial organization of the differentiating tissues. Keywords: transcriptional response to growth factor treatment Cells derived from mouse embryonic stage 11.5 limb buds were cultured and treated with purified Wnt3a protein or vehicle controls. The transcriptional response was detected using spotted cDNA microarrays after 2 hrs or 4 hrs of treatment. 4 biological replicates were used per condition.

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TATTOO-seq delineates spatial and cell type-specific regulatory programs in the developing limb

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