Janus Kinase 1 Is Essential for Inflammatory Cytokine Signaling and Mammary Gland Remodeling

Abstract

Despite a wealth of knowledge about the significance of individual signal transducers and activators of transcription (STATs), essential functions of their upstream Janus kinases (JAKs) during postnatal development are less well defined. Using a novel mammary gland-specific JAK1 knockout model, we demonstrate here that this tyrosine kinase is essential for the activation of STAT1, STAT3, and STAT6 in the mammary epithelium. The loss of JAK1 uncouples interleukin-6-class ligands from their downstream effector, STAT3, which leads to the decreased expression of STAT3 target genes that are associated with the acutephase response, inflammation, and wound healing. Consequently, JAK1-deficient mice exhibit impaired apoptosis and a significant delay in mammary gland remodeling. Using RNA sequencing, we identified several new JAK1 target genes that are upregulated during involution. These include Bmf and Bim, which are known regulators of programmed cell death. Using a BMF/BIMdouble-knockout epithelial transplant model, we further validated that the synergistic action of these proapoptotic JAK1 targets is obligatory for the remodeling of the mammary epithelium. The collective results of this study suggest that JAK1 has nonredundant roles in the activation of particular STAT proteins and this tyrosine kinase is essential for coupling inflammatory cytokine signals to the cell death machinery in the differentiated mammary epithelium.T he postnatal growth, functional differentiation, and postlactational remodeling of the epithelial compartment of the mammary gland are controlled by hormones and locally synthesized cytokines (1). While estrogen is primarily required for the elongation of mammary ducts after the onset of puberty (2, 3), progesterone and prolactin orchestrate the specification, proliferation, and differentiation of secretory alveolar cells during pregnancy (4-6). Following lactation and the weaning of the young, circulating levels of prolactin (PRL) decline and milk stasis induces the local production of interleukin-6 (IL-6)-class cytokines, in particular, IL-6, leukemia inhibitory factor (LIF), and oncostatin M (OSM). These IL-6-class cytokines trigger a cascade of intracellular events that orchestrate a process known as mammary gland remodeling, which entails the death and selective removal of terminally differentiated alveolar cells (7-9).PRL and IL-6-class cytokines signal through their corresponding receptors and utilize Janus kinases (JAKs) to activate downstream signal transducers and activators of transcription (STATs) that subsequently alter the transcriptional profile, growth, and homeostasis of mammary epithelial cells. Five of the seven STAT proteins that are known in mammals (i.e., STAT1, STAT3, STAT5a, STAT5b, and STAT6) have been found to be sequentially activated at defined stages of mammary gland development (10, 11). The levels of phosphorylated STAT1 have been reported to be elevated in nulliparous females, but this particular transcription factor seems to be largely dispensable for normal ma...

Crosstalk between STAT5 activation and PI3K/AKT functions in normal and transformed mammary epithelial cells

“…The activation of these STATs as defined by their tyrosine phosphorylation and nuclear translocation occurs in a sequential manner in response to the major cytokines that control a particular developmental program. The highest levels of STAT1 activation in the mammary epithelium were observed in nulliparous and nonpregnant parous females after postlactational remodeling as well as briefly during mid-pregnancy (Philp et al, 1996; Sakamoto et al, 2016b). STAT6 was reported to have a biologically significant role as a downstream effector of IL-4 and IL-13 in developing alveoli of pregnant females (Khaled et al, 2007).…”

Interrogating the relevance of mitochondrial apoptosis for vertebrate development and postnatal tissue homeostasis

“…Simultaneous pharmacological inhibition of caspases and cathepsins delayed mesodermal cell death in chicken limbs more potently than caspase inhibition alone (Zuzarte-Luis et al 2007). Furthermore, a critical role of cathepsins B and L has been reported in mammary gland remodeling during involution (Kreuzaler et al 2011), although its physiological role remains disputed (Sakamoto et al 2016). Regardless, these observations again support the idea that mitochondrial apoptosis is not solely responsible for developmentally programmed cell death in vertebrates but works in conjunction with, or can be compensated for by, other cell killing mechanisms.…”

“…Additionally, increased lymphadenopathy, higher serum Ig levels, and a higher incidence of glomerulonephritis and malignant disease, leading to premature lethality, were observed when compared with BIM mutant mice (Labi et al 2014;Woess et al 2015). BMF also supports BIM in promoting mammary epithelial cell death during organ morphogenesis Schmelzle et al 2007) and the late stage of involution after the cessation of lactation, a classic example of developmental apoptosis (Sakamoto et al 2016;Schuler et al 2016).…”

“…Phenocopies Bim −/− mice regarding development; infertile males due to defective spermatogenesis (later than in Baxdeficient mice) Coultas et al 2005 Bim/Bmf Born at reduced frequency, similar to Bim −/− mice; webbed feet; increased incidence of vaginal atresia (65%) and malocclusion of the incisors (25%); increased resistance of Hubner et al 2010;Labi et al 2014;Woess et al 2015;Sakamoto et al 2016 Continued Tuzlak et al et al 2009). As a consequence, hepatocytes or pancreatic β cells from BID-deficient mice are insensitive to Fas ligation in vivo (Yin et al 1999).…”

“…As such it connects the extrinsic and intrinsic apoptosis signaling pathways in certain cell types that rely on efficient neutralization of XIAP to generate above-threshold caspase activity upon death receptor (DR) ligation (Jost Bouillet et al 1999Bouillet et al , 2002Hildeman et al 2002;Enders et al 2003;Pellegrini et al 2003;Villunger et al 2003b;Fischer et al 2007;Mailleux et al 2007;Schuler et al 2016 Bmf Normal embryonic development; lymphocytes and MEFs partially protected from glucocorticoid or histone deacetylase inhibition (HDACi)-induced apoptosis; mild Bcell hyperplasia; potential overlap with BIM in mammary gland involution Labi et al 2008;Hubner et al 2010;Sakamoto et al 2016;Schuler et al 2016 Bid Normal embryonic development; hepatocytes resist FasLinduced and TNF-induced hepatitis; delayed apoptosis upon FasL or TNF-α treatment in MEFs; pancreatic β cells resist death receptor (DR) killing; reduced response to the DNAdamaging agents (controversial); neurons more protected from ischemic cell death (controversial) Yin et al 1999;Plesnila et al 2001;Sax et al 2002;Kamer et al 2005;Zinkel et al 2005;Kaufmann et al 2007Kaufmann et al , 2009McKenzie et al 2008;Engel et al 2010 Bad Normal embryonic development; minor protection from apoptotic stimuli; modest reduction in B-cell proliferation in response to mitogens; decreased production of IgG upon immunization; prolonged platelet life span; reduced sensitivity to TNF-induced hepatitis (controversial); impaired glucose stimulated insulin secretion and glucokinase activity Danial et al 2003;Ranger et al 2003;Kelly et al 2010;Yan et al 2013;Ottina et al 2015 Puma Normal embryonic development; hematopoietic and nonhematopoietic cell types highly resistant to genotoxic stress and γ irradiation as well as growth factor deprivation; high-fat-diet-fed as well as chow diet-fed mice show decreased food intake and ambulatory capacity Jeffers et al 20...…”