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Curr Dev Nutr. 2019 Jun 13;3(Suppl 1). pii: nzz031.P06-030-19. doi: 10.1093/cdn/nzz031.P06-030-19. eCollection 2019 Jun.

Bovine mRNAs in Cow's Milk Exosomes Are Bioavailable and Translated into Peptides in Non-bovine Systems (P06-030-19).

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1
University of Nebraska Lincoln.

Abstract

Objectives:

Exosomes are natural nanoparticles that participate in cell-to-cell communication by transferring regulatory molecules such as microRNAs from donor cells to recipient cells. Previously, we have demonstrated that exosomes and microRNAs are not exclusively obtained from endogenous synthesis but may also be absorbed from milk.We assessed whether 1) exosomes contain mRNAs (i.e., RNAs other than microRNAs), 2) mRNAs are bioavailable in mice, and 3) bovine mRNAs are translated into peptides.

Methods:

For mRNA analysis, exosomes were isolated from bovine milk using differential ultracentrifugation, and RNA cargos were analyzed by RNA-sequencing. For bioavailability analysis, a synthetic fragment of fluorophore (IRDye)-labeled bovine CSN3 mRNA was transfected into bovine milk exosomes, which were administered to Balb/c mice by oral gavage. Tissues distribution of CSN3 mRNA was assessed 24 h after gavage by using a LiCor Odyssey CLx imager. For analysis of mRNA translation, mRNA from bovine milk exosomes were translated using the rabbit reticulocyte lysate system and BODIPY-labeled lysine. Peptides were separated by two-dimensional gel electrophoresis and fluorescence was visualized using a Typhoon FLA 7000 scanner. Statistical analysis, NA.

Results:

We detected > 3600 bovine mRNAs in exosomes. Most mRNAs were truncated; 107 mRNAs contained their natural ATG start codons. Thirteen of these mRNAs, including CSN3, encoded bovine proteins and peptides with amino acid sequences distinct from those in human and murine orthologs (Table 1). Mice absorbed CSN3 mRNA encapsulated in milk exosomes, which accumulated primarily in the liver (Fig. 1). Nine bovine peptide spots were detected by two-dimensional electrophoresis (Fig. 2).

Conclusions:

Bovine milk exosomes contain mRNAs which are bioavailable and translated into peptides in non-bovine systems. We speculate that food mRNAs might play a role in food allergies and immune tolerance. Future studies: Identification of peptides by LC/MS-MS is in progress. We will assess the relevance of mRNA translation for allergies and tolerance in animal models.

Funding Sources:

NIFA, NIH, Gates Foundation, PureTech, Inc. and USDA Hatch and Multistate. J.Z. is a consultant for PureTech.

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