The BH3-only protein BIM contributes to late-stage involution in the mouse mammary gland

Abstract

After cessation of lactation, involution of the mouse mammary gland proceeds in two distinct phases, a reversible and an irreversible one, which leads to the death and removal of alveolar cells. Cell death is preceded by the loss of STAT5 activity, which abrogates cell differentiation and gain of STAT3 activity. Despite early observations implicating BCL2 (B cell lymphoma 2) family proteins in this process, recent evidence suggests that STAT3-controlled cathepsin activity is most critical for cell death at the early stage of involution. Somewhat surprisingly, this cell death associates with but does not depend on the activation of pro-apoptotic effector caspases. However, transgenic overexpression of BCL2, that blocks caspase activation, delays involution while conditional deletion of BclX accelerates this process, suggesting that BCL2 family proteins are needed for the effective execution of involution. Here, we report on the transcriptional induction of multiple pro-apoptotic BCL2 family proteins of the ‘BH3-only' subgroup during involution and the rate-limiting role of BIM in this process. Loss of Bim delayed epithelial cell clearance during involution after forced weaning in mice, whereas the absence of related Bmf had minor and loss of Bad or Noxa no impact on this process. Consistent with a contribution of BCL2 family proteins to the second wave of cell death during involution, loss of Bim reduced the number of apoptotic cells in this irreversible phase. Notably, the expression changes observed within the BCL2 family did not depend on STAT3 signalling, in line with its initiating role early in the process, but rather appear to result from relief of repression by STAT5. Our findings support the existence of a signalling circuitry regulating the irreversible phase of involution in mice by engaging BH3-only protein-driven mitochondrial apoptosis.

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

“…Among known factors that control canonical mitochondrion-associated apoptosis, we identified the genes encoding the BH3-only proteins BMF and BIM (Bcl2l11) to be targets of JAK1 signaling. The results of analysis of involuting mammary glands of single-knockout mice confirm the findings presented in a recent report that showed that both proapoptotic proteins contribute to the initiation of apoptosis (58). More importantly, using a BMF/BIM-double-knockout transplant model, we now provide clear evidence that both of these JAK1 targets act synergistically in an epithelium-specific manner.…”

“…According to the ChIP-Seq and ChIP/qRT-PCR results, STAT3 is bound to the chromatin in proximity to the Bmf and Bim loci, and its association with GAS sequences near transcriptional start sites was increased during involution. This finding suggests that STAT3 might be directly involved in the transcription of these genes, and it is therefore puzzling that STAT3-knockout mice did not show the same deregulated expression in Bmf and Bim that was observed in the JAK1-knockout mice (58). One possible explanation is that other transcription factors whose expression levels are controlled by JAK1 (e.g., Runx1) also contribute to the expression of Bmf and Bim.…”

“…One possible explanation is that other transcription factors whose expression levels are controlled by JAK1 (e.g., Runx1) also contribute to the expression of Bmf and Bim. Whether STAT5 acts as a suppressor of Bim transcription, as suggested previously (58), warrants further validation using appropriate genetic in vivo models. The fact that the nuclear accumulation of pSTAT3 is not altered by the loss of BMF and BIM still supports the notion that these BH3-only proapoptotic factors function genetically downstream of the IL-6/JAK1/STAT3 axis.…”

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

“…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).…”

“…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...…”