Elf5 conditional knockout mice reveal its role as a master regulator in mammary alveolar development: Failure of Stat5 activation and functional differentiation in the absence of Elf5

Abstract

The transcription factor Elf5 plays an important role in mammary gland development. However, because of the embryonic lethality of Elf5 straight knockout mice, prior studies have been limited to experiments with Elf5 haploinsufficient animals, overexpression systems or transplants. Here, we have utilized K14-Cre to generate mammary-gland specific Elf5 conditional knockout mice. During pregnancy, Elf5-null mammary epithelium completely failed to initiate alveologenesis, and a characteristic of virgin ductal epithelial cells persisted postpartum. We demonstrate that the loss of Elf5 leads to the absence of alveolar secretory markers confirming previous published data. Interestingly, the developmental block due to a lack of Elf5 could not be restored by multiple gestations. Elf5-null mammary epithelial cells also display disorganized cell structures as evident by altered cell polarities, which might be the cause for collapsed lumina. We observe reduced levels of Stat5 and attenuated Stat5 activity as measured by p-Stat5 levels both in Elf5-null mammary glands as well as cultured mammary epithelial cells. This data suggests that the failure of alveolar and lactogenic differentiation due to the loss of Elf5 is mediated in part due to impaired Stat5 activity. In support of this hypothesis, we show by ChIP experiments that Stat5a promoter contains a conserved Elf5-binding site that is occupied by Elf5 in mammary glands. Mammary epithelia lacking Elf5 exhibited downregulation of several other critical genes involved in alveologenesis, suggesting Elf5 as a master regulator in alveolar development. We propose a model for Elf5-mediated alveolar development, in which Elf5 regulates the expression of key mediators of the PrlR/Jak2/Stat5 signaling pathway.

Transcription factor regulatory networks in mammary epithelial development and tumorigenesis

“…More recently, the importance of Elf-5 in lobuloalveolar differentiation was supported through the analysis of Elf-5 conditional knockout mice (Choi et al, 2009). Like STAT5A-deficient mammary glands, mammary-specific disruption of Elf-5 led to a complete block in alveolar differentiation (Choi et al, 2009). Taken together, these studies support Elf-5 as a master regulator of the alveolar switch that occurs during pregnancy.…”

“…In Elf-5 À/À transplants, an increase in CD61 þ luminal progenitor cells was observed, which positions Elf-5 as an important factor within the luminal alveolar lineage (Oakes et al, 2008). More recently, the importance of Elf-5 in lobuloalveolar differentiation was supported through the analysis of Elf-5 conditional knockout mice (Choi et al, 2009). Like STAT5A-deficient mammary glands, mammary-specific disruption of Elf-5 led to a complete block in alveolar differentiation (Choi et al, 2009).…”

“…Recent work demonstrates that deletion of STAT5A/5B in the mammary glands results in a severe defect in lobuloalveolar differentiation without effects on ductal outgrowth (Yamaji et al, 2009). Similarly, Elf-5 À/À glands are also capable of normal ductal elongation and branching morphogenesis but fail to undergo proper lobuloalveolar differentiation (Oakes et al, 2008;Choi et al, 2009). On its face, these results suggest that STAT5A/5B and Elf-5 control similar aspects of lobuloalveolar differentiation.…”

“…Indeed, the presence of STAT transcription factor-binding sites have been identified within the Elf-5 proximal promoter and Elf5 expression was lost in STAT5A/5B-deficient glands (Yamaji et al, 2009), arguing that STAT5A lies immediately upstream of Elf-5 in a transcription factor hierarchy. This regulation is likely more complex, given that the block in alveolar differentiation in Elf-5 null mammary glands may, in part, be attributed to decreased transcriptional activation of STAT5 through Elf-5 transcription binding sites within the STAT5A proximal promoter (Choi et al, 2009). However, one important difference between the phenotypes observed in STAT5A/5B and Elf-5 deficient mammary glands is that the Elf-5 deficient glands exhibit an increase in the CD61 þ luminal progenitor population (Oakes et al, 2008); whereas the STAT5A/5B-deficient glands exhibit a dramatic deficiency in CD61 þ progenitor cells.…”

“…Although these factors are clearly necessary for the proper development of the virgin mammary gland ductal tree and subsequent development during pregnancy, it is not yet clear how these proteins control the specification of distinct cell types within the emerging differentiation hierarchy of the mammary gland. In this respect, recent work has begun to focus on elucidating the roles and interactions of several key transcription factors during normal mammary epithelial differentiation of specific mammary epithelial populations Asselin-Labat et al, 2007;Oakes et al, 2008;Choi et al, 2009;Yamaji et al, 2009). In addition to terminal differentiation, another set of transcription factors are involved in the involution phase of mammary gland development, notably STAT3.…”