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Glutaminolysis is involved in the activation of mTORC1 in in vitro-produced porcine embryos.

ABSTRACT: Glutamine supplementation to porcine embryo culture medium improves development, increases leucine consumption, and enhances mitochondrial activity. In cancer cells, glutamine has been implicated in the activation of mechanistic target of rapamycin complex 1 (mTORC1) to support rapid proliferation. The objective of this study was to determine if glutamine metabolism, known as glutaminolysis, was involved in mTORC1 activation in porcine embryos. Culture with 3.75 mM GlutaMAX improved development to the blastocyst stage compared to culture with 1 mM GlutaMAX, and culture with 0 mM GlutaMAX decreased development compared to all groups with GlutaMAX. Ratios of phosphorylated to total MTOR were increased when embryos were cultured with 3.75 or 10 mM GlutaMAX, which was enhanced by the absence of leucine, but ratios for RPS6K were unchanged. As another indicator of mTORC1 activation, colocalization of MTOR and a lysosomal marker was increased in embryos cultured with 3.75 or 10 mM GlutaMAX in the absence of leucine. Culturing embryos with glutaminase inhibitors decreased development and the ratio of phosphorylated to total MTOR, indicating reduced activation of the complex. Therefore, glutaminolysis is involved in the activation of mTORC1 in porcine embryos, but further studies are needed to characterize downstream effects on development.

SUBMITTER: Chen PR

PROVIDER: S-EPMC8361685 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:Fertilization is followed by complex changes in cytoplasmic composition and extensive chromatin reprogramming which results in the abundant activation of totipotent embryonic genome at embryonic genome activation (EGA). While chromatin reprogramming has been widely studied in several species, only a handful of reports characterize changing transcriptome profiles and resulting metabolic changes in cleavage stage embryos. The aims of the current study were to investigate RNA profiles of in vivo developed (ivv) and in vitro produced (ivt) porcine embryos before (2-cell stage) and after (late 4-cell stage) EGA and determine major metabolic changes that regulate totipotency. The period before EGA was dominated by transcripts responsible for cell cycle regulation, mitosis, RNA translation and processing (including ribosomal machinery), protein catabolism, and chromatin remodelling. Following EGA an increase in the abundance of transcripts involved in transcription, translation, DNA metabolism, histone and chromatin modification, as well as protein catabolism was detected. The further analysis of members of overlapping GO terms revealed that despite that comparable cellular processes are taking place before and after EGA (RNA splicing, protein catabolism), different metabolic pathways are involved. This strongly suggests that a complex metabolic switch accompanies EGA. In vitro conditions significantly altered RNA profiles before EGA, and the character of these changes indicates that they originate from oocyte and are imposed either before oocyte aspiration or during in vitro maturation. IVT embryos have altered content of apoptotic factors, cell cycle regulation factors and spindle components, and transcription factors, which all may contribute to reduced developmental competence of embryos produced in vitro. Overall, our data are in good accordance with previously published, genome-wide profiling data in other species. Moreover, comparison with mouse and human embryos showed striking overlap in functional annotation of transcripts during the EGA, suggesting conserved basic mechanisms regulating establishment of totipotency in mammalian development.

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Glutaminolysis is involved in the activation of mTORC1 in in vitro-produced porcine embryos.

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