CLOCK regulates mammary epithelial cell growth and differentiation

Abstract

Circadian clocks influence virtually all physiological processes, including lactation. Here, we investigate the role of the CLOCK gene in regulation of mammary epithelial cell growth and differentiation. Comparison of mammary morphology in late-pregnant wild-type and ClockΔ19 mice, showed that gland development was negatively impacted by genetic loss of a functional timing system. To understand whether these effects were due, in part, to loss of CLOCK function in the gland, the mouse mammary epithelial cell line, HC11, was transfected with short hairpin RNA that targeted Clock (shClock). Cells transfected with shClock expressed 70% less Clock mRNA than wild-type (WT) HC11 cultures, which resulted in significantly depressed levels of CLOCK protein (P < 0.05). HC11 lines carrying shClock had four-fold higher growth rates (P < 0.05), and the percentage of cells in G1 phase was significantly higher (90.1 ± 1.1% of shClock vs. 71.3 ± 3.6% of WT-HC11) following serum starvation. Quantitative-PCR (qPCR) analysis showed shClock had significant effects (P < 0.0001) on relative expression levels of Ccnd1, Wee1, and Tp63 qPCR analysis of the effect of shClock on Fasn and Cdh1 expression in undifferentiated cultures and cultures treated 96 h with dexamethasone, insulin, and prolactin (differentiated) found levels were reduced by twofold and threefold, respectively (P < 0.05), in shClock line relative to WT cultures. Abundance of CDH1 and TP63 proteins were significantly reduced in cultures transfected with shClock These data support how CLOCK plays a role in regulation of epithelial cell growth and differentiation in the mammary gland.

Exposure to chronic light–dark phase shifts during the prepartum nonlactating period attenuates circadian rhythms, decreases blood glucose, and increases milk yield in the subsequent lactation

“…A short-day photoperiod during the nonlactating dry period increased expression of genes that regulate proliferation and differentiation of cells in the mammary gland (Bentley et al, 2015); it also attenuated rhythms of expression of core clock genes in mammary tissue of goats (Casey et al, 2018). Our work in cells and rodents supports a role for the circadian clocks in regulation of mammary epithelial growth and differentiation (Casey et al, 2016). Thus, it is possible that attenuation of circadian rhythms in the PS treatment affected mammary-specific clocks in a manner that increased proliferation of the mammary tissue preparturition and is reflected as an increase in milk yield during lactation.…”

“…Human and rodent studies support a mechanistic link between circadian disruption and the development of metabolic diseases (Pillar and Shehadeh, 2008;Gangwisch, 2009;Möller-Levet et al, 2013). Of particular interest is that metabolism is altered and lactation performance is impaired in mice with mutations in the CLOCK gene, one of the genes that make up the core molecular clocks that generate circadian rhythms of gene expression (Dolatshad et al, 2006;Hoshino et al, 2006;Casey et al, 2016). Here our aim was to determine whether the circadian timing system plays a role in maintaining homeostasis and regulating the onset of lactation in transition dairy cows.…”

Administering xCT Inhibitors Based on Circadian Clock Improves Antitumor Effects

“…The treatment with high-serum concentrations induces robust circadian rhythms in the cultured cells (28,29). In addition, this treatment also affects cell-cycle synchronization (30). In this study, Wee1 expression did not show circadian rhythm between 24 and 48 hours after serum treatment, and we confirmed that Clock genes directly affect xCT levels without cell-cycle effects in serum-shocked colon 26 cells.…”

A molecular circadian clock operates in the parathyroid gland and is disturbed in chronic kidney disease associated bone and mineral disorder

“…29 The circadian clock has been implicated in gating control of the cell cycle at a number of targets: induction of Wee1 for inhibition of Cyclin B1 at the G 2 /M (Gap 2 /Mitosis) transition, 30 induction of p20 and p21 at the G 1 /S (Gap 1 /Synthesis) transition, 31,32 and regulation of p27, Cyclin D1, and c-Myc for progression through G 1 . [33][34][35][36] Cell turnover rate is very low in the parathyroid gland under normal conditions; with an estimated mean life span of normal parathyroid cells being 19 years in humans and 2 years in rats. 37 This rate increases dramatically in secondary hyperparathyroidism (sHPT) caused by chronic kidney disease (CKD).…”