# Integrated fetal testicular transcriptomic and epigenomic profiles during maternal nutrient restriction with dietary melatonin intervention

## TL;DR

This study shows how melatonin can help counteract the negative effects of poor maternal nutrition on fetal testis development by altering gene activity and DNA methylation.

## Contribution

The study integrates transcriptomic and epigenomic data to reveal how melatonin supplementation affects fetal testis development under maternal nutrient restriction.

## Key findings

- Melatonin supplementation altered gene expression and DNA methylation in fetal testes under nutrient restriction.
- Specific genes like DAAM1 and KYAT1 showed significant expression changes with melatonin.
- Nutrient restriction alone caused 370 DMRs without corresponding gene expression changes.

## Abstract

Fetal development is a critical period that establishes reproductive efficiency and herd performance depending on in-utero epigenetic modifications. Dietary restrictions may affect fetal testis development and the offspring fertility. Several studies have connected genetic instability to circadian cycle disruptions, including epigenetic modifications to melatonin, a key regulator. On day 160 of gestation, 17 male-bearing Brangus heifers were assigned to one of four groups in a 2 × 2 factorial treatment arrangement: adequately fed (ADQ; 100% NRC recommendation, n = 3), nutrient restricted (RES; 60% NRC recommendation, n = 5), or ADQ or RES supplemented with 20 mg/d melatonin (ADQ-MEL, n = 5; RES-MEL, n = 4). On day 240 of gestation, heifers underwent Cesarean sections to collect fetuses and testicular tissues. The fetal testicular tissue was processed and analyzed using the Methyl-MiniSeq Service: Genome-wide bisulfite sequencing (Methyl MiniSeq-GWBS). Sequence reads from Methyl Mini-Seq libraries were identified using standard Illumina platform calling software for methylome profile. RNA-Seq libraries were then sequenced on the Illumina platform for transcriptome profile. The common genes between differentially methylated regions (DMRs) and differentially expressed genes (DEGs) across different treatment groups were identified by an overlap analysis using bedtools v2.31.1. There were 413 DMRs in RES-CON and ADQ-CON testicular tissues, without differential gene expression. Compared with the ADQ-CON group, the ADQ-MEL group showed 411 DMRs and a higher KYAT1 gene expression (P-adj <0.05) without methylation changes. Comparing RES-MEL with RES-CON showed that 9 genes (DAAM1, COL28A1, RPL10, TRPM3, SLIT, ARHGEF40, SYT1, TMEM35B, CSPG4B) were expressed more in the former (P-adj <0.05). The only hypomethylated gene was DAAM1 located on chromosome 10. However, 13 genes (PTPRU, snRNP-E, TMEM59L, MUC5B, ANAPC15, FAM221A, SHCBPiL, PAQR5, PPP4R3C, DTNB, LncRNA, SHANK2, RIC3) showed increased expression in RES-CON vs. RES-MEL without differential methylation alterations, yet there were 370 DMRs. Five genes showed increased expression in RES-MEL compared with ADQ-MEL (P-adj <0.05), including histone H2B on chromosome 23. Two genes (PTPRU, TDRD10) showed increased expression in ADQ-MEL compared with RES-MEL (P-adj <0.05) without affecting methylation and 344 DMRs. In conclusion, dietary melatonin supplementation to nutrient restricted dams may influence fetal development as epigenomic and transcriptomic regulators are altered.

Epigenetics offers a new way to study embryonic development, as it is affected by environmental influences and the causes of illness. Understanding how dietary limitation and/or melatonin supplementation alter epigenetic modifications would help us optimize pregnant cow management and male offspring development through nutritional manipulation, offering a physiological alternative to pharmacotherapies.