Unknown,Transcriptomics,Genomics,Proteomics

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Effects of DKK1 on global gene expression in female and male bovine embryos at the morula stage

ABSTRACT: Successful embryonic development depends upon molecules secreted by the reproduc-tive tract. Among such molecules is dickkopf 1 (DKK1), an canonical WNT antagonist that can also activate the planar cell polarity (PCP) pathway. Objectives were to determine whether DKK1 regulates expression of genes that promote differentiation, possibly through the PCP pathway, and to determine if actions of DKK1 on the embryonic transcriptome were dependent on embryo sex. Bovine oocytes were fertilized with pools of X- or Y-sorted sperm. Embryos were treated with 100 ng/ml DKK1 or vehicle at Day 5 of development and harvested 24 h later for microarray analysis of global gene expression. A total of 9,931 transcripts were identified as being expressed. with124 being differentially expressed genes between females and males. A total of 68% of the genes upregulated in females were located on the X chromosome. DKK1 changed expression of 132 genes in females and 136 in males. Of these, 34 genes were regulated by DKK1 in both sexes - 14 in the same direction and 20 in opposite directions. Evidence for regulation of the PCP pathway by DKK1 was the finding that DKK1 regulated expression of genes involved in cell polarization and differentiation in both females and males. In both sexes, DKK1 regulated expression of many genes associated with HNF4A, a marker of hypoblast cells that promotes formation of cell junctions. In conclusion, DKK1 regulates cell differentiation and embryonic development in a sex-dependent manner and effects may be mediated, at least in part, by activation of the PCP pathway. Bovine embryos produced with X- or Y-sorted sperm were cultured in vitro and treated with vehicle or DKK1 at day 5 of development and harvested at day 6 for RNA isolation, amplification and hibridization to Affymetrix Bovine Gene 1.0 ST arrays. The study was done in 5 replicates using pools of 20 morulae for each treatment.

ORGANISM(S): Bos taurus

SUBMITTER: Anna Denicol

PROVIDER: E-GEOD-60098 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:Sex affects function of the developing mammalian embryo as early as the preimplantation period. There were two goals of the current objective. The first was to determine the degree and nature of differences in gene expression between female and male embryos in the cow at the morula stage of development. The second objective was to determine whether DKK1, a molecule known to alter differentiation of the blastocyst, would affect gene expression differently for female and male morulae. In Experiment 1, female and male embryos were treated with DKK1 at Day 5 after insemination. Morulae were harvested 24 h after treatment, pooled in groups of 20 for microarray analysis and RNA subjected to analysis of gene expression by microarray hybridization. There were 662 differentially expressed genes between females and males and 128 of these genes had a fold change ≥ 1.5 between the two sexes. Of the genes upregulated in females, 49.5% were located in the X chromosome. Functional analysis predicted that cell survival was greater in female embryos. Experiment 2 involved a similar design except that transcripts for 12 genes previously reported to be affected by sex, DKK1 or the interaction were quantified by quantitative polymerase chain reaction. Expression of all genes tested that were affected by sex in experiment 1 was affected in a similar manner in Experiment 2. In contrast, effects of DKK1 on gene expression were largely not repeatable in Experiment 2. The exception was for the Hippo signaling gene AMOT, which was inhibited by DKK1. In Experiment 3, embryos produced by fertilization with unsorted sperm were treated with DKK1 at Day 5 and abundance of transcripts for CDX2, GATA6, and NANOG determined at Days 5, 6 and 7 after insemination. There was no effect of DKK1 on expression of any of the three genes. In conclusion, female and male bovine embryos have a different pattern of gene expression as early as the morula stage, and this is due to a large extent to expression of genes in the X chromosomes in females. Differential gene expression between female and male embryos is likely the basis for increased resistance to cell death signals in female embryos and disparity in responses of female and male embryos to changes in the maternal environment.