Dataset Information

Identification and Characterization of the OCT4 Upstream Regulatory Region in Sus scrofa.

ABSTRACT: OCT4 plays pivotal roles in maintaining pluripotency during early mammalian embryonic development and in embryonic stem cells. It is essential to establish a reporter system based on the OCT4 promoter region to study pluripotency. However, there is still a lack of information about the porcine OCT4 upstream reporter system. To improve our understanding of the porcine OCT4 regulatory region, we identified conserved regions in the porcine OCT4 promoter upstream region by sequence-based comparative analysis using various mammalian genome sequences. The similarity of nucleotide sequences in the 5' upstream region was low among mammalian species. However, the OCT4 promoter and four regulatory regions, including distal and proximal enhancer elements, had high similarity. Next, a functional analysis of the porcine OCT4 promoter region was conducted. Luciferase reporter assay results indicated that the porcine OCT4 distal enhancer and proximal enhancer were highly activated in mouse embryonic stem cells and embryonic carcinoma cells, respectively. A comparison analysis of naïve and primed state marker gene expression in a dual-reporter assay showed that the expression levels of naïve and primed markers differed in fluorescence signal between high-expressing cells and low-expressing cells. Similar to OCT4 upstream-based reporter systems derived from other species, the porcine OCT4 upstream region-based reporter constructs showed exclusive expression patterns depending on the state of pluripotency. This work provides basic information about the porcine OCT4 upstream region and various porcine OCT4 fluorescence reporter constructs, which can be applied to study species-specific pluripotency in early embryo development and the establishment of embryonic stem cells in pigs.

SUBMITTER: Kim SH

PROVIDER: S-EPMC6434273 | biostudies-literature | 2019

REPOSITORIES: biostudies-literature

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Identification and Characterization of the OCT4 Upstream Regulatory Region in Sus scrofa.

Kim Seung-Hun SH Choi Kwang-Hwan KH Lee Dong-Kyung DK Lee Mingyun M Hwang Jae Yeon JY Lee Chang-Kyu CK

Stem cells international 20190312

OCT4 plays pivotal roles in maintaining pluripotency during early mammalian embryonic development and in embryonic stem cells. It is essential to establish a reporter system based on the OCT4 promoter region to study pluripotency. However, there is still a lack of information about the porcine OCT4 upstream reporter system. To improve our understanding of the porcine OCT4 regulatory region, we identified conserved regions in the porcine OCT4 promoter upstream r ...[more]

PMID: 30992705

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Project description:BACKGROUND:In animal breeding, identification of causative genetic variants is of major importance and high economical value. Usually, the number of candidate variants exceeds the number of variants that can be validated. One way of prioritizing probable candidates is by evaluating their potential to have a deleterious effect, e.g. by predicting their consequence. Due to experimental difficulties to evaluate variants that do not cause an amino-acid substitution, other prioritization methods are needed. For human genomes, the prediction of deleterious genomic variants has taken a step forward with the introduction of the combined annotation dependent depletion (CADD) method. In theory, this approach can be applied to any species. Here, we present pCADD (p for pig), a model to score single nucleotide variants (SNVs) in pig genomes. RESULTS:To evaluate whether pCADD captures sites with biological meaning, we used transcripts from miRNAs and introns, sequences from genes that are specific for a particular tissue, and the different sites of codons, to test how well pCADD scores differentiate between functional and non-functional elements. Furthermore, we conducted an assessment of examples of non-coding and coding SNVs, which are causal for changes in phenotypes. Our results show that pCADD scores discriminate between functional and non-functional sequences and prioritize functional SNVs, and that pCADD is able to score the different positions in a codon relative to their redundancy. Taken together, these results indicate that based on pCADD scores, regions with biological relevance can be identified and distinguished according to their rate of adaptation. CONCLUSIONS:We present the ability of pCADD to prioritize SNVs in the pig genome with respect to their putative deleteriousness, in accordance to the biological significance of the region in which they are located. We created scores for all possible SNVs, coding and non-coding, for all autosomes and the X chromosome of the pig reference sequence Sscrofa11.1, proposing a toolbox to prioritize variants and evaluate sequences to highlight new sites of interest to explain biological functions that are relevant to animal breeding.

Dataset Information

Identification and Characterization of the OCT4 Upstream Regulatory Region in Sus scrofa.

Publications

Identification and Characterization of the <i>OCT4</i> Upstream Regulatory Region in <i>Sus scrofa</i>.

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