ABSTRACT: Chip-chip from Mouse E10.5 anterior and posterior immortalised limb cell lines and limb tissue for H3K27me3 antibody and from Mouse E10.5 anterior and posterior limb tissue for Ring1B antibody.
Project description:Anterior-posterior differences in H3K27me3 and Ring1B enrichment over the 5 prime Hoxd genes in E10.5 murine distal forelimbs. Chromatin immunoprecipitation (ChIP) of H3K27me3 together with Ring1B and by ChIP-on-chip analysis demonstrated that over the 5 prime HoxD locus H3K27me3 enrichment is decreased and Ring1B enrichment is sparse in limb cells derived from the distal posterior forelimb bud of E10.5 mouse embryos. Array design includes 2 biological replicates for H3K27me3 in the cell lines and Ring1B in the limb tissue, and 2 biological replicates and 2 dye swap replicates for H3K27me3 in the limb tissue.
Project description:Anterior-posterior differences in H3K27me3 and Ring1B enrichment over the 5 prime Hoxd genes in E10.5 murine distal forelimbs. Chromatin immunoprecipitation (ChIP) of H3K27me3 together with Ring1B and by ChIP-on-chip analysis demonstrated that over the 5 prime HoxD locus H3K27me3 enrichment is decreased and Ring1B enrichment is sparse in limb cells derived from the distal posterior forelimb bud of E10.5 mouse embryos.
Project description:The transcriptomes of immortalized anterior and posterior limb cell lines derived from E10.5 embryos were compared. Total RNA extracted from three E10.5 anterior and posterior limb cell line samples were compared.
Project description:Shh signal mediated by Gli family of transcription factors regulates digit growth and patterning in early limb development. Shh expression in the posterior margin of the limb bud defines the zone of polarizing activity. However, much less is know about downstream targets that mediate Shh signal functions. In this dataset, we include the expression data obtained from dissected anterior and posterior halves of mouse limb bud respectively. These data are used to obtain 889 transcripts that were upregulated 1.3 fold or more in the posterior limb bud, and 1189 transcripts that were enriched in the anterior limb bud at 1.3 fold or more. Two samples were analyzed. We generate pairwise comparisons between anterior and posterior limb tissues. Genes with a fold-change ≥1.3 were selected.
Project description:Shh signal mediated by Gli family of transcription factors regulates digit growth and patterning in early limb development. Shh expression in the posterior margin of the limb bud defines the zone of polarizing activity. However, much less is know about downstream targets that mediate Shh signal functions. In this dataset, we include the expression data obtained from dissected anterior and posterior halves of mouse limb bud respectively. These data are used to obtain 889 transcripts that were upregulated 1.3 fold or more in the posterior limb bud, and 1189 transcripts that were enriched in the anterior limb bud at 1.3 fold or more.
Project description:Amputation of a salamander limb triggers anterior and posterior connective tissue cells to form distinct signalling centres that together fuel regeneration. Anterior and posterior identities are established during development and are thought to persist lifelong in the form of positional memory. However, neither the molecular basis of positional memory nor whether positional memory can be altered is known. Here, we identify a positive feedback loop responsible for posterior identity in the axolotl (Ambystoma mexicanum) limb. Posterior cells express residual Hand2 transcription factor from development, which primes them to form a Shh signalling centre after limb amputation. During regeneration, Shh signalling is also upstream of Hand2 expression. After regeneration, Shh is shut down but Hand2 is sustained, safeguarding posterior memory. We exploited this regeneration circuitry to convert anterior cells to a posterior cell memory state. Transient exposure of anterior cells to Shh during regeneration kick-started an ectopic Hand2-Shh loop, leading to stable Hand2 expression and a lasting competence to express Shh. Our results implicate positive feedback in the stability of positional memory and reveal that positional memory is more easily reprogrammed in one direction (anterior to posterior) than the other. Modifying positional memory in regenerative cells changes their signalling outputs, which has implications for tissue engineering.
Project description:Amputation of a salamander limb triggers anterior and posterior connective tissue cells to form distinct signalling centres that together fuel regeneration. Anterior and posterior identities are established during development and are thought to persist lifelong in the form of positional memory. However, neither the molecular basis of positional memory nor whether positional memory can be altered is known. Here, we identify a positive feedback loop responsible for posterior identity in the axolotl (Ambystoma mexicanum) limb. Posterior cells express residual Hand2 transcription factor from development, which primes them to form a Shh signalling centre after limb amputation. During regeneration, Shh signalling is also upstream of Hand2 expression. After regeneration, Shh is shut down but Hand2 is sustained, safeguarding posterior memory. We exploited this regeneration circuitry to convert anterior cells to a posterior cell memory state. Transient exposure of anterior cells to Shh during regeneration kick-started an ectopic Hand2-Shh loop, leading to stable Hand2 expression and a lasting competence to express Shh. Our results implicate positive feedback in the stability of positional memory and reveal that positional memory is more easily reprogrammed in one direction (anterior to posterior) than the other. Modifying positional memory in regenerative cells changes their signalling outputs, which has implications for tissue engineering.
Project description:Mammalian primordial germ cells (PGCs) migrate asynchronously through the embryonic hindgut and dorsal mesentery to reach the gonads. To characterize transcriptional heterogeneity of migrating PGCs and their niches, we performed single-cell RNA sequencing of 13,262 mouse PGCs and 7,868 surrounding somatic cells during migration (E9.5, E10.5, E11.5) and in anterior versus posterior locations to enrich for leading and lagging migrants. Analysis of PGCs by position revealed dynamic gene expression changes between faster or earlier migrants in the anterior and slower or later migrants in the posterior at E9.5. At E10.5, we found that anterior PGCs upregulate Nodal transcriptional targets including Lefty1/2 and validated this LEFTY1/2 upregulation via whole-mount immunofluorescence staining. This positional and temporal atlas of mouse PGCs supports the idea that niche interactions along the migratory route elicit changes in proliferation, actin dynamics, pluripotency and epigenetic reprogramming.
