Metabolomics,Unknown,Transcriptomics,Genomics,Proteomics

Dataset Information

Brain BRCA1 conditional knockout

ABSTRACT: BRCA1 nestin CRE conditional knockout cortrices of P7 animals were compared to wildtype littermates to characterize the mutant phenotype. Keywords: expression BRCA1 conditional knockouts using nestin CRE and a null allele with an inverted neo cassette at the 5' end of the exon 11 of BRCA1 on one floxed allele flanking exons 5-13. Cortices of 3 wildtype animals were compred to 3 BRCA cKO at postnatal day 7.

ORGANISM(S): Mus musculus

SUBMITTER: Gerald Pao

PROVIDER: E-GEOD-6310 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Publications

BRCA1 tumour suppression occurs via heterochromatin-mediated silencing.

Zhu Quan Q Pao Gerald M GM Huynh Alexis M AM Suh Hoonkyo H Tonnu Nina N Nederlof Petra M PM Gage Fred H FH Verma Inder M IM

Nature 20110907 7363

Mutations in the tumour suppressor gene BRCA1 lead to breast and/or ovarian cancer. Here we show that loss of Brca1 in mice results in transcriptional de-repression of the tandemly repeated satellite DNA. Brca1 deficiency is accompanied by a reduction of condensed DNA regions in the genome and loss of ubiquitylation of histone H2A at satellite repeats. BRCA1 binds to satellite DNA regions and ubiquitylates H2A in vivo. Ectopic expression of H2A fused to ubiquitin reverses the effects of BRCA1 lo ...[more]

PMID: 21901007

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Project description:The splicing regulator PTBP2 controls a program of embryonic splicing required for neuronal maturation. The splicing regulatory proteins PTBP1 and PTBP2 show distinct temporal expression profiles in the developing brain. Neuronal progenitor cells predominantly express PTBP1, whereas developing neurons express high levels of PTBP2, which are subsequently reduced late in neuronal maturation. We show here that PTBP2 and the program of splicing it controls are essential to proper neuronal maturation and survival. To investigate its in vivo function, we generated conditional PTBP2 null alleles in mice. Loss of PTBP2 in neuronal progenitor cells leads to neonatal death without gross defects in brain architecture. Mice with specific depletion of PTBP2 in the cortex and forebrain are viable. However over the first three postnatal weeks, when the normal cortex expands and develops mature circuits, the PTBP2 null cortices degenerate. We find that PTBP2-/- neurons cultured from embryonic brain show the same initial viability as wild type cells with proper early marker expression and neurite outgrowth. Strikingly, between 10 and 20 days in culture PTBP2 null neurons undergo a catastrophic failure to mature and die. To assess the target transcripts leading to these phenotypes, we examined the genomewide splicing changes in the PTBP2 null brains. This identified a large number of mis-regulated exons that share a temporal pattern of regulation; in the absence of PTBP2 many isoforms normally found in adults are precociously expressed in the developing brain. Transcripts following this pattern encode essential neuronal proteins affecting neurite growth, pre- and post-synaptic assembly, and synaptic transmission. Our results define a new genetic regulatory program essential for neuronal survival and maturation, where PTBP2 acts to temporarily repress expression of protein isoforms until the final maturation of the neuron. Mice carrying a conditional "floxed" PTBP2 allele of PTBP2 were crossed to mice carrying Cre recombinase driven by either the nestin or Emx1 promoters. The resulting knockout mutant mouse brains were analyzed for changes in gene expression and alternative splicing. Knockout mice were compared to wildtype littermates. Whole mouse brain polyA plus RNA was isolated from two Nestin-cre knockout embryos at embryonic day 18 and compared to two wildtype littermates. Total polyA plus RNA was isolated from the cortices of an Emx-cre knockout and its wildtype littermate at postnatal day 1. This RNA was converted to standard Illumina paired end libraries using the Truseq kit. The Emx sample and control libraries were strand specific through elimination of the second strand of cDNA using the USER enzyme. Libaries were sequenced on an Illumina HiSeq using the standard paired end protocol. Data was analyzed using the Cufflinks pipeline. Splicing analysis was carried out using the SpliceTrap program.

Dataset Information

Brain BRCA1 conditional knockout

Publications

BRCA1 tumour suppression occurs via heterochromatin-mediated silencing.

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