Project description:Hox genes control domain specific cartilage pattern formation along the proximodistal axis. Hoxa11 and Hoxd11 (Hox11) show overlapping expression in the mesenchyme of the zeugopod and regulate cartilage growth and differentiation. Hoxa13 and Hoxd13 (Hox13) are expressed in the autopodal mesenchyme and are crucial for the autopodal cartilage patterning. Since both Hox11 and Hox13 are involved in the limb cartilage development, they may share the target genes. To identify common target genes among the Hoxa11 and Hoxa13, we performed ChIP-Seq analysis using specific antibody.
Project description:Limb patterning relies in a large part on the function of the Hox family of developmental genes. While the differential expression of Hox genes shifts from the anterior-posterior (A-P) to the proximal-distal (P-D) axis around embryonic day 11 (E11), whether this shift coincides with a more global change of P-D versus A-P patterning program remains unclear. By performing and analyzing the transcriptome of the developing limb bud from E10.5 to E12.5, at single cell resolution, we have uncovered transcriptional trajectories which revealed a general switch from A-P to P-D genetic program between E10.5 and E11.5. Interestingly, the transcriptional trajectories at E10.5 all end with cells expressing either proximal or distal markers suggesting a progressive acquisition of P-D identity. Moreover, we observed that distally expressed Hox genes, namely Hoxa13 and Hoxd13, act as a key determinant for P-D patterning as their transcriptional control results in their distal restricted expression, which in turn restricts Hoxa11 in the proximal limb bud domain, in progenitor cells of the zeugopod. Finally, we identified three categories of genes expressed in the distal limb mesenchyme characterized by distinct temporal expression dynamics. As anticipated from previous results, HOX13 binding was observed within or in the neighborhood of several of these genes consistent with previous evidence suggesting that the transition from the early/proximal to the late/distal transcriptome of the limb mesenchyme largely relies on HOX13 function.
Project description:E11.5 metanephric mesenchyme and ureteric bud were dissected from the E11.5 kidney rudiment using fine manual microdissection (ureteric bud only) or both fine manual microdissection and laser capture microdissection (metanephric mesenchyme) to define the gene expression profiles of these structures. Additionally, HoxA11, HoxD11 compound null E11.5 metanephric mesenchyme was obtained through laser capture microdissection allowing analysis of possible Hox targets in kidney development. Targets from multiple biological replicates of each were generated and the expression profiles were determined using Affymetrix MOE430_v2 arrays. Using microdissection techniques, ureteric bud and metanephric mesenchyme were dissected from E11.5 kidney rudiments allowing the identificated genes specifically regulated in either structure. In addition, Hoxa11, Hoxd11 compound null E11.5 metanephric mesenchyme were normalized to wild type embryonic controls allowing the identification of potential Hox targets in normal kidney development. Each structure/genotype were represented in biological (seperate embryo) replicate.
Project description:Analysis of mouse limb bud (E10.5) lacking the Bhlha9 gene. Bhlha9 knockout mouse shows syndactyly and poliosis in the limb. This microarray results provides insight into the molecular mechanisms underlying Bhlha9 function in the limb development DNA microarray analysis was performed using Affymetrix mouse genome 430 2.0 array. RNA samples were obtained from the whole limb bud of the E10.5 wild-type and Bhlha9 knockout embryos described above. Total RNA (200 ng) was reverse-transcribed and biotinylated using the GeneChip 3â² IVT Express Kit (Affymetrix). The microarray data were summarized using the MAS 5.0 method.
Project description:Comparing gene expression of cells from the E10.5 limb bud ZPA and the rest of the E10.5 limb bud from Shhgfpcre heterozygotes separated by FACS. Experiment Overall Design: 8 samples, 4 ZPA and 4 rest of the limb
Project description:Analysis of mouse limb bud (E10.5) lacking the Bhlha9 gene. Bhlha9 knockout mouse shows syndactyly and poliosis in the limb. This microarray results provides insight into the molecular mechanisms underlying Bhlha9 function in the limb development
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: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:Proper development of limb bud relies on the concordance of various signals, otherwise limb deformities occur. We report that heterogeneous nuclear ribonucleoprotein K (hnRNPK) is essential for limb bud development. here, we knock out Hnrnpk in limb bud and exert the RNA-seq, ATAC-seq, CUT&RUN-seq, and Hi-C assay using primary limb bud cells to explore the function of Hnrnpk in limb bud development.
Project description:We established the differentiation method of a limb bud organoid from mouse embryonic stem cells (mESCs) using SFEBq. mESCs-derived limb bud organoid selectively differentiate into forelimb or hindlimb by adjusting the retinoic acids activity. To evaluate a correlation of gene expression between limb bud organoid and embryonic tissues (limb bud, branchial arch, cardiac, and tail bud), we performed comparative transcriptome analysis using RNA-seq.