Proteomic analysis of parthenogenetic and <i>in vitro</i> fertilized porcine embryos

Gupta, Mukesh Kumar; Jang, JinHyeok; Jung, Jin Woo; Uhm, Sang Jun; Kim, Kwang Pyo; Lee, Hoon Taek

doi:10.1002/pmic.200800700

Cited by 33 publications

(28 citation statements)

References 79 publications

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“…It is also functionally linked to cell growth and its expression decreases concomitantly with the growth arrest during differentiation [45]. Most interestingly, it has been reported that TCP-1 is related to the growth and survival during pig embryo development, and it is more drastically upregulated in pig parthenogenetic embryos than in fertilized embryos [46]. In this study, TCP-1α was found expressed in all the three cell types (Fig.…”

Section: Discussionsupporting

confidence: 58%

Proteomic Profiling of Rabbit Embryonic Stem Cells Derived from Parthenotes and Fertilized Embryos

et al. 2013

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Rabbit embryonic stem (rES) cells can be derived from various sources of embryos. However, understanding of the gene expression profile, which distincts embryonic stem (ES) cells from other cell types, is still extremely limited. In this study, we compared the protein profiles of three independent lines of rabbit cells, i.e., fibroblasts, fertilized embryo-derived stem (f-rES) cells, and parthenote-derived ES (p-rES) cells. Proteomic analyses were performed using two-dimensional gel electrophoresis (2-DE) and mass spectrometry. Collectively, the expression levels of 100 out of 284 protein spots differed significantly among these three cell types (p<0.05). Of those differentially expressed spots, 91% were identified in the protein database and represented 63 distinct proteins. Proteins with known identities are mainly localized in the cytoplasmic compartments (48%), nucleus (14%), and cytoskeletal machineries (13%). These proteins were majorly involved in biological functions of energy and metabolic pathways (25%), cell growth and maintenance (25%), signal transduction (14%), and protein metabolisms (10%). When protein expression levels among cell types were compared, six proteins associated with a variety of cellular activities, including structural constituents of the cytoskeleton (tubulins), structural molecule (KRT8), catalytic molecules (α-enolase), receptor complex scaffold (14-3-3 protein sigma), microfilament motor proteins (Myosin-9), and heat shock protein (HSP60), were found highly expressed in p-rES cells. Two proteins related to HSP activity and structural constituent of cytoskeleton in f-rES cells, and one structural molecule activity protein in fibroblasts showed significantly higher expression levels (p<0.05). Marker protein expressions in f-rES and p-rES cells were further confirmed by Western blotting and immunocytochemical staining. This study demonstrated unique proteomic profiles of the three rabbit cell types and revealed some novel proteins differentially expressed between f-rES and p-rES cells. These analyses provide insights into rES cell biology and would invite more in-depth studies toward rES cell applications.

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Section: Discussionsupporting

confidence: 58%

Proteomic Profiling of Rabbit Embryonic Stem Cells Derived from Parthenotes and Fertilized Embryos

et al. 2013

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“…Peptides corresponding to CAR protein (NP_001070948) were identified using TurboSEQUEST (Thermo Finnigan) and SEQUEST algorithm [20, 21], which was further validated using Scaffold software (Proteome Software, Portland, OR; version 3_00_02) [22]. Probabilities that the identified peptide and protein are correctly from CAR were >95% and >99%, respectively.…”

Section: Methodsmentioning

confidence: 99%

Estradiol induces cytochrome P450 2B6 expression at high concentrations: Implication in estrogen-mediated gene regulation in pregnancy

Koh

Jurkovic

Yang

et al. 2012

Biochemical Pharmacology

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Pregnancy alters the rate and extent of drug metabolism, but little is known about the underlying molecular mechanism. We have found that 17β-estradiol (E2) upregulates expression of the major drug-metabolizing enzyme CYP2B6 in primary human hepatocytes. Results from promoter reporter assays in HepG2 cells revealed that E2 activates constitutive androstane receptor (CAR) and enhances promoter activity of CYP2B6, for which high concentrations of E2 reached during pregnancy were required. E2 triggered nuclear translocation of CAR in primary rat hepatocytes that were transiently transfected with human CAR as well as in primary human hepatocytes, further confirming transactivation of CAR by E2. E2-activated estrogen receptor (ER) also enhanced CYP2B6 promoter activity. The DNA-binding domain of ER was not required for the induction of CYP2B6 promoter activity by E2, suggesting involvement of a non-classical mechanism of ER action. Results from deletion and mutation assays as well as electrophorectic mobility shift and supershift assays revealed that two AP-1 binding sites (−1782/−1776 and −1664/−1658 of CYP2B6) are critical for ER-mediated activation of the CYP2B6 promoter by E2. Concurrent activation of both ER and CAR by E2 enhanced CYP2B6 expression in a synergistic manner. Our data demonstrate that at high concentrations reached during pregnancy, E2 activates both CAR and ER that synergistically induce CYP2B6 expression. These results illustrate pharmacological activity of E2 that would likely become prominent during pregnancy.

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“…State-of-the-art LC-MS/MS has been applied for a proteomic characterisation of both in vitro-fertilised (IVF) as well as parthenogenetic (PA) porcine embryos (Gupta et al 2009) 24 h after fertilisation or activation, respectively. As shown in mice, PA embryos are unable to develop to term except after genetic modification to correct for imprinted genes such as IGF2/H19 (Kono et al 2004).…”

Section: Proteomics From In Vitro-fertilised and Parthenogenetic Embryosmentioning

confidence: 99%

Dynamic proteome signatures in gametes, embryos and their maternal environment

Arnold

Fröhlich

2011

Reprod. Fertil. Dev.

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Comprehensive molecular analysis at the level of proteins represents a technically demanding, but indispensable, task since several post-transcriptional regulation mechanisms disable a valid prediction of quantitative protein expression profiles from transcriptome analysis. In crucial steps of gamete and early embryo development, de novo transcription is silenced, meaning that almost all macromolecular events take place at the level of proteins. In this review, we describe selected examples of dynamic proteome signatures addressing capacitation of spermatozoa, in vitro maturation of oocytes, effect of oestrous cycle on oviduct epithelial cells and embryo-induced alterations to the maternal environment. We also present details of the experimental strategies applied and the experiments performed to verify quantitative proteomic data. Far from being comprehensive, examples were selected to cover several mammalian species as well as the most powerful proteomic techniques currently applied. To enable non-experts in the field of proteomics to appraise published proteomic data, our examples are preceded by a customised description of quantitative proteomic methods, covering 2D difference gel electrophoresis (2D-DIGE), nano-liquid chromatography combined with tandem mass spectrometry, and label-free as well as stable-isotope labelling strategies for mass spectrometry-based quantifications.

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Proteomic analysis of parthenogenetic and in vitro fertilized porcine embryos

Cited by 33 publications

References 79 publications

Proteomic Profiling of Rabbit Embryonic Stem Cells Derived from Parthenotes and Fertilized Embryos

Proteomic Profiling of Rabbit Embryonic Stem Cells Derived from Parthenotes and Fertilized Embryos

Estradiol induces cytochrome P450 2B6 expression at high concentrations: Implication in estrogen-mediated gene regulation in pregnancy

Dynamic proteome signatures in gametes, embryos and their maternal environment

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