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Differenetial Gene Expression in Limbs of Wild Type and Lmx1b KO mice during limb dorsalization (e11.5, e12.5 & e13.5)

ABSTRACT: Lmx1b regulates dorsalization of limb fates, but the mechanism of this regulation has not been characterized. To identify candidate genes regulated by Lmx1b we compared the limbs from Lmx1b KO mice to wild type mice during limb dorsalization (e11.5-13.5). Differentially expressed genes that we common to all three stages examined were considered to be likely candidates for Lmx1b regulation and further evaluated.

ORGANISM(S): Mus musculus

PROVIDER: GSE34732 | GEO | 2012/05/22

SECONDARY ACCESSION(S): PRJNA150415

REPOSITORIES: GEO

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Differenetial Gene Expression in Limbs of Wild Type and Lmx1b KO mice during limb dorsalization (e11.5, e12.5 & e13.5)

Project description:Lmx1b regulates dorsalization of limb fates, but the mechanism of this regulation has not been characterized. To identify candidate genes regulated by Lmx1b we compared the limbs from Lmx1b KO mice to wild type mice during limb dorsalization (e11.5-13.5). Differentially expressed genes that we common to all three stages examined were considered to be likely candidates for Lmx1b regulation and further evaluated. At 11.5 and 12.5 dpc, embryos were harvested and the limb buds with the limb girdles were isolated. Embryos at 13.5dpc were also harvested and their distal limb buds (zeugopods and autopods) were isolated. Embryos were genotyped to confirm Lmx1b homozygosity (-/- or +/+). RNA from embryonic forelimbs and hindlimbs of wild type (WT) and Lmx1b KO mice was harvested using the Rneasy Kit (Qiagen). RNA was pooled to decrease genetic variability, i.e., six limbs at 11.5 dpc, three limbs at 12.5 dpc and six limbs at 13.5 dpc. Duplicate samples were generated using different embryos for each stage and then hybridized to the Affymetrix GeneChip® Mouse Genome 430 2.0 Array (UCI, Irvine, CA).

2012-05-21 | E-GEOD-34732 | biostudies-arrayexpress

Lmx1b ChIP-seq dataset in embryonic day 12.5 (e12.5) wild type mouse limb buds

Project description:Lmx1b is a homeodomain transcription factor responsible for limb dorsalization. Despite striking double-ventral (loss-of-function) and double-dorsal (gain-of-function) limb phenotypes, no direct downstream gene targets in the limb have been confirmed. To determine direct targets of Lmx1b during limb dorsalization (E12.5), we performed chromatin immunoprecipitation followed by next generation sequencing (ChIP-seq). Nearly 84% (n=617) of the Lmx1b-bound genomic fragments or intervals (LBIs) identified by two Lmx1b-ChIP-seqs overlap with chromatin regulatory marks indicative of potential cis-regulatory modules (PCRMs). In addition, 73 LBIs mapped to known cis-regulatory modules (CRMs) active during limb development. We compared Lmx1b-bound PCRMs to genes differentially expressed by Lmx1b at E12.5 and found 292 PCRMs within 1 Mb of 254 Lmx1b-regulated genes. Gene ontology analysis of these associated genes suggests that Lmx1b mediates dorsalization through the regulation of extracellular matrix production, bone/joint formation, axonal guidance, vascular development, cell proliferation and cell movement. We validated the functional activity of 2 PCRMs associated to Lmx1b-regulated genes, demonstrating activity and overlap with the associated gene during limb development. This is the first report to describe the genome-wide distribution of Lmx1b binding during limb development, directly linking Lmx1b to targets that accomplish limb dorsalization.

2017-05-06 | GSE84064 | GEO

Identification of genes controlled by LMX1B in the developing mouse hindlimb bud

2008-02-15 | GSE10516 | GEO

Transcription profiling of mouse developing hindlimb bud to identify genes controlled by LMX1B

Project description:A control vs. genetic knockout experiment aimed at determining what RNAs are upregulated or downregulated in e11.5 mouse proximal limb tissue lacking the Lmx1b gene. Because Lmx1b is required for dorsal-ventral patterning of the limb, this screen gives insight into what putative downstream targets of Lmx1b contribute to dorsal-ventral patterning. Experiment Overall Design: The proximal portion of e11.5 hindlimb buds (~500um) was used for RNA extraction. Each array was hybridized with pooled cRNAs from both hindlimb buds of three embryos (6 hindlimbs/sample).

2008-04-08 | E-GEOD-10516 | biostudies-arrayexpress

Identification of genes controlled by LMX1B in the developing limb, embryonic day (E) 13.5

Project description:A control vs. genetic knockout experiment aimed at determining what RNAs are upregulated or downregulated in E13.5 mouse limb tissue lacking the Lmx1b gene. Because LMX1B is required for dorsal-ventral patterning of the limb, this screen gives insight into what putative downstream targets of Lmx1b contribute to dorsal-ventral patterning. The forelimb or hindlimb of wild-type controls or Lmx1b loss-of-function mutants was used for RNA extraction. Each array was hybridized with cDNAs of an individual limb.

2010-07-30 | E-GEOD-21446 | biostudies-arrayexpress

Identification of genes controlled by LMX1B in the developing limb, embryonic day (E) 13.5

2010-07-06 | GSE21446 | GEO

Chromatin conformation of the Shh locus in limbs.

Project description:Expression of Shh in the limb is dependent on a distant enhancer located 850kb away from the gene promoter. We performed chromatin conformation capture (4C-Seq) analysis of the Shh locus in E11.5 mouse limbs so as to better understand the relationship between this regulatory interaction and the structural organisation of the locus. The experiments were performed on wild-type animals as well as on animals carrying an engineered chromosomal inversion (INV(6-C2) (INV6)) . The different viewpoints used (Shh, ZRS, Nom1, Rbm33, Rnf32) enable to characterize the 3D organisation of the locus in the different contexts. The inversion INV6 disrupts the normal topological organisation of the Shh locus in a compact 3D-domain and prevents the physical and regulatory interaction between Shh and its limb enhancer, even though the genomic distance separating the two elements is reduced compared to WT.

2017-01-12 | E-MTAB-4980 | biostudies-arrayexpress

Failure of digit tip regeneration in the absence of Lmx1b suggests Lmx1b functions disparate from dorsoventral polarity

Project description:Mammalian digit tip regeneration is linked to the presence of nail tissue, but a nail-explicit model is missing. Here, we report that nail-less double-ventral digits of ΔLARM1/2 mutants that lack limb-specific Lmx1b enhancers, fail to regenerate. To separate the nail’s effect from the lack of DV polarity, we also interrogate double-dorsal double-nail digits and show that they regenerate. Thus, DV polarity is not a prerequisite for regeneration and the nail requirement is supported. Transcriptomic comparison between wild-type and non-regenerative ΔLARM1/2 mutant blastemas reveals differential up-regulation of vascularization and connective tissue functional signatures in wild-type versus upregulation of inflammation in the mutant. These results, together with the finding of Lmx1b expression in the postnatal dorsal dermis underneath the nail and uniformly in the regenerative blastema opens the possibility of additional Lmx1b roles in the progression of digit tip regeneration, in addition to the indirect effect of mediating the formation of the nail.

2022-12-02 | GSE202223 | GEO

Expression profiles of E11.5 wildtype and Shox2 knockout embryonic forelimbs

Project description:The development of vertebrate extremities is a complex process which requires a highly coordinated network of different transcriptional activities. The homeodomain transcription factor Shox2 is a key player in limb formation controlling neural, muscular and skeletal development. Here, we compared gene expression profiles of wildtype and Shox2 knockout limbs using microarray experiments to identify Shox2 target genes. Forelimbs of E11.5 mouse embryos were dissected and genotyped for RNA extraction. RNA from 3-4 embryos of 2 different pregnancies was used for hybridisation to 2 arrays per genotype (wildtype and Shox2 knockout) and compared.

2014-01-16 | E-GEOD-51523 | biostudies-arrayexpress

Sequencing of iMDK-treated regenerating limbs

Project description:Treatment with midkine inhibitor iMDK inhibits limb regeneration. To elucidate the transcriptional mechanisms of iMDK treatment, we sequenced regenerating limbs treated with either DMSO or iMDK (bulk) in biological triplicate at 11 days post-amputation (dpa).

2020-01-15 | GSE132325 | GEO

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