Transcriptome response of oocytes to seasonal heat stress in beef cows
Introduction: Reduced reproductive performance due to seasonal heat stress is a major problem in the beef and dairy industries. Examining the effects of heat stress on the molecular mechanism of bovine oocytes can help to better understand their alterations on a transcriptional level and to plan and mitigate those effects. We aimed to study the response of oocytes to seasonal thermal stress.  Materials and Methods: Multiparous angus dry beef cows (n?=?11) were kept together during the study and subjected to synchronisation and stimulation of follicular growth using a 5-day CIDR and follicle-stimulating hormone (FSH) protocol. Ovum pick-up (OPU) was conducted on all animals in the winter (January) and summer (August). Cumulus-oocyte-complexes (COCs) were isolated from the follicular fluid aspirated during the OPU procedure. Denuded oocytes were further isolated from the COCs, snap frozen, and stored at -80?°C until further use for library sequencing, RNA sequencing, bioinformatics and gene enrichment analysis. Additionally, rectal temperatures were recorded on the day of each OPU. Environmental data were collected daily three weeks before the day of each OPU using the Florida Automated Weather Network. Statistical analysis for average rectal temperature included overall mean (average rectal temperature), treatment effects (season), and the residual. Treatment effect was considered significant at P?=?0.05. Data are presented as mean?±?standard error of the mean. RNA was extracted from five biological replicates/pools of oocytes (each containing n?=?2) followed by library preparation and sequencing (NovaSeq; Illumina).  Results: As expected, environmental conditions were contrasting [average air temperature (11.5?°C vs 27.5?°C), average max air temperature (16.9?°C vs 33.7?°C), relative humidity (83.5% vs 82.3%), and temperature-humidity index (53.39 vs 79.16) for winter and summer, respectively]. Average rectal temperature was greater (P?=?0.03) in summer (39.2?±?0.2?°C) than in the winter (38.8?±?0.2?°C). Data analysis revealed an up-regulation of 446 transcripts and a down-regulation of 940 transcripts in oocytes collected during summer compared to winter (Fold Change?=?2; FDR P-value?=?0.05). Upregulated transcripts are involved in protein digestion and absorption, ATP-binding cassette transporter, oocyte meiosis, and progesterone-mediated oocyte maturation pathways. Conversely, down-regulated transcripts are involved in pathways related to extracellular matrix-receptor interaction, focal adhesion, and PI3K-Akt signaling.  Conclusions: In conclusion, exposure of cows to thermal stress can significantly alter the transcriptome of oocytes, which may negatively impact subsequent developmental competence. Overall design: Multiparous angus dry beef cows (n?=?11) were kept together during the study and subjected to synchronisation and stimulation of follicular growth using a 5-day CIDR and follicle-stimulating hormone (FSH) protocol. Ovum pick-up (OPU) was conducted on all animals in the winter (January) and summer (August). Cumulus-oocyte-complexes (COCs) were isolated from the follicular fluid aspirated during the OPU procedure. Denuded oocytes were further isolated from the COCs, snap frozen, and stored at -80?°C until further use for RNA sequencing