The objective of this study was to determine the effects of winter environmental conditions and production systems on targeted mRNA expression profile of hair follicles (HF) in cattle, specifically focusing on genes regulating stress, inflammation, cellular integrity, and coat color. Two cattle breeds (Bos indicus, Brahman and Bos taurus, Angus) were subjected to two production systems without (NAT) or with a commonly used growth-promoting technology in commercial feedlots, TRT [100-200 mg of trenbolone acetate and 14-28 mg estradiol benzoate anabolic implants, in-feed monensin/tylosin (35/7 g/ton DM) and ractopamine hydrochloride (27 g/ton DM)] for a 180 day-period in Colorado State, USA (n = 50 steers/group). Growth performances were monitored and hair follicles were collected on a monthly basis from October 2023 to March 2024. Total RNA was extracted, reverse transcribed, and target genes were amplified by qPCR using 2^-ΔΔCt method. Angus cattle exhibited significantly higher ADG and FE from October through February compared to their Brahman counterparts, with the TRT production system further improving FE (P < 0.05) during November-December and January-February relative to the NAT system. A significantly higher gene expression of Cxcr2 and a lower expression of Asip were observed in the Angus breed compared to the Brahman counterparts. This breed-specific differential expression may reflect the differences in hair coat characteristics and physiological adaptive pathways. Compared to the NAT system, TRT treatment significantly upregulated the expression of Xcl1 and Asip while downregulating that of Cxcr2. These changes were consistent with the effects of anabolic steroids, antibiotics, and/or ractopamine. The expression of heat shock proteins (Hsp60, Hsp70, Hsp90), cytokines (IL6, IL18, Tnfα, Crp, IL10), chemokines and their related receptors (Xcl1, Cxcl12/14, Ccl2/4/5/20, Ccr2, Cxcr1/2), tight junction proteins (Cldn1, Ocln), and coat color-associated genes (Asip, Mc1r, Bmp2, Hoxc12, Lef1, Zhx3) followed seasonal patterns with peaks in October-November and/or December-February, which coincided with low temperatures. Collectively, to our knowledge, this is among the first studies showing a differential gene expression between breeds (Angus vs. Brahman), production systems (TRT vs. NAT), and season. By identifying hair follicle gene expression signatures associated with cold stress in beef cattle, this study provides a foundation for future mechanistic investigations.