Mammary transcriptome reveals cell maintenance and protein turnover support milk synthesis in early-lactation cows

A more complete understanding of the molecular mechanisms that support milk synthesis is needed to develop strategies to efficiently and sustainably meet the growing global demand for dairy products. With the postulate that coding gene transcript abundance reflects relative importance in supporting milk synthesis, we analyzed the global transcriptome of early lactation cows across magnitudes of normalized RNA-Seq read counts. Total RNA was isolated from milk samples collected from early-lactation cows (n = 6) following two treatment periods of postruminal lysine infusion of 0 or 63 g/day. Twelve libraries were prepared and sequenced on an Illumina NovaSeq6000 platform using paired end reads. Normalized read counts were averaged across both treatments, because EBseq analysis found no significant effect of lysine infusion. Approximately 10% of the total reads corresponded to 12,730 protein coding transcripts with a normalized read count mean ≥5. For functional annotation analysis, the protein coding transcripts were divided into nine categories by magnitude of reads. The 13 most abundant transcripts (≥50K reads) accounted for 67% of the 23M coding reads and included casein and whey proteins, regulators of fat synthesis and secretion, a ubiquitinating protein, and a tRNA transporter. Mammalian target of rapamycin, JAK/STAT, peroxisome proliferator-activated receptor alpha, and ubiquitin proteasome pathways were enriched with normalized reads ≥100 counts. Genes with ≤100 reads regulated tissue homeostasis and immune response. Enrichment in ontologies that reflect maintenance of translation, protein turnover, and amino acid recycling indicated that proteostatic mechanisms are central to supporting mammary function and primary milk component synthesis.