Single-nucleus RNA sequencing reveals cell type-specific response to heat stress in bovine mammary gland

Gene-set enrichment analyses revealed increased ribosomal activities across HS-responsive clusters and enrichment of protein folding and metabolic pathways in luminal alveolar cells, suggesting elevated proteostasis demands under stress. Pseudotime analysis positioned luminal cells along a progenitor-to-secretory trajectory under TN, accompanied by increased casein gene expression, whereas under HS, mature luminal cells shifted toward a homeostasis regulatory state.

Cell-cell communication analysis demonstrated HS-induced remodeling of interepithelial signaling, including altered ERBB4-mediated signaling from luminal hormone-sensing to alveolar lineages. Finally, transcription factor activity profiling highlighted cell-type-specific HS-activated regulators and their downstream target genes. Together, this cell type-resolved atlas delineates how HS alters bovine mammary epithelial function, developmental state, and intercellular crosstalk.

These findings point to proteostasis pressure, disrupted signaling pathways, and rewired regulatory networks as mechanistic contributors to reduced lactational performance under HS, offering insights for improving heat resilience in dairy cattle.