Maternal dietary restriction and one-carbon metabolite supplementation modulate microRNA expression in bovine fetal skeletal muscle

Authors: DINIZ, WELLISON - Auburn University; BANERJEE, PRIYANKA - Auburn University; Crouse, Matthew; WARD, ALISON - University Of Saskatchewan; DAHLEN, CARL - North Dakota State University; SYRING, JESSICA - North Dakota State University; ENTZIE, YSSI - North Dakota State University; KING, LAYLA - University Of Minnesota Crookston; ANAS, MUHAMMAD - North Dakota State University; DANESHI, MOJTABA - North Dakota State University; HIRCHERT, MARA - North Dakota State University; SWANSON, REBECCA - North Dakota State University; BOROWICZ, PAWEL - North Dakota State University; CATON, JOEL - North Dakota State University

Submitted to: Plant and Animal Genome Conference
Publication Type: Abstract Only
Publication Acceptance Date: 11/11/2024
Publication Date: 1/2/2025
Citation: Diniz, W.J., Banerjee, P., Crouse, M.S., Ward, A.K., Dahlen, C.R., Syring, J.G., Entzie, Y.L., King, L.E., Anas, M., Daneshi, M., Hirchert, M.R., Swanson, R.M., Borowicz, P., Caton, J.S. 2025. Maternal dietary restriction and one-carbon metabolite supplementation modulate microRNA expression in bovine fetal skeletal muscle [abstract]. Plant and Animal Genome 32 Conference, January 10-15, 2025, San Diego, California. Poster Number P186.

Interpretive Summary:

Technical Abstract: Maternal nutrition influences fetal growth and development through epigenetic mechanisms. Yet, the effects of specific nutrients, including one-carbon metabolites (OCM), in fetal epigenetic programming remain unclear. This study evaluated the impact of maternal dietary restriction and strategic supplementation of OCM (folate, vitamin B12, methionine, and choline) on miRNA expression modulation of fetal muscle development. Crossbred Angus heifers were assigned in a 2 × 2 factorial arrangement to one of four treatments comprising the main effects of maternal body weight gain (Control - C, 0.45 kg/day ADG or Restricted - R, -0.23 kg/day ADG) and two levels of OCM supplementation (+OCM – daily rumen-protected methionine (7.4 g/d) and choline (44.4 g/d); weekly injections of 320 mg of folate and 20 mg of vitamin B12; or -OCM: control diet with no supplementation). Treatments were applied from breeding to day 62 of gestation, after which all animals were put on the control diet (adjusted for 0.6 kg/d gain) with no OCM supplementation. On day 161 of gestation, heifers were necropsied, and fetal Longissimus dorsi muscle was collected and used for total RNA isolation (n = 7 per group, except C+OCM, n = 8) and miRNA sequencing. MiRNAs were annotated via the mirDeep2 pipeline, and differentially expressed miRNAs (DEMs) were identified using DESeq2, revealing 76 unique DEMs across all six comparisons (FDR < 0.05). The main effect of the rate of body weight gain (R-OCM vs. C-OCM) without supplementation resulted in the greatest number of DEMs (n = 68), with 44 miRNAs downregulated in the fetuses from the R-OCM group. Among the upregulated miRNAs, bta-miR-1 promotes myogenesis by binding to the HDAC4 gene, enhancing myogenic gene expression to accelerate differentiation and reduce proliferation while also inhibiting Pax7 to reduce satellite cell proliferation. The bta-miR-206, upregulated in the R-OCM group, acts as a central regulator of skeletal muscle cell differentiation primarily by suppressing genes that inhibit myogenesis. Under a restricted diet and OCM supplementation (R+OCM vs. R-OCM), 11 miRNAs were upregulated. We used the miRNet tool to identify potential gene targets regulated by DEMs and the underlying pathways affected. From the R-OCM vs. C-OCM contrast, we identified 2,137 genes targeted by 56 miRNAs, which included myogenic genes and transcription factors such as MYOG, FOXO4, GATA2, and HAND1. We also identified the histone deacetylase 7 (HDAC7), which regulates myocyte migration and differentiation targeted by the mir-491, mir-339a, and miR-339b. The functional analysis of the gene targets retrieved KEGG pathways such as folate biosynthesis, vitamin digestion and absorption, and arginine and proline metabolism. Nutrient sensing-related pathways included the PI3K-Akt signaling, MAPK, and FoxO signaling pathways. Our results provide new insights into fetal epigenetic programming, showing that miRNAs play a central role in modulating skeletal muscle development in response to maternal diet and OCM availability, targeting myogenic transcription factors and chromatin remodelers.