Scribble is required for pregnancy-induced alveologenesis in the adult mammary gland

Abstract

The cell polarity protein scribble (SCRIB) is a crucial regulator of polarization, cell migration and tumorigenesis. Whereas SCRIB is known to regulate early stages of mouse mammary gland development, its function in the adult gland is not known. Using an inducible RNA interference (RNAi) mouse model for downregulating SCRIB expression, we report an unexpected role for SCRIB as a positive regulator of cell proliferation during pregnancy-associated mammary alveologenesis. SCRIB was required in the epithelial cell compartment of the mammary gland. Lack of SCRIB attenuated prolactin-induced activation of the JAK2-STAT5 signaling pathway. In addition, loss of SCRIB resulted in the downregulation of prolactin receptor (PRLR) at cell surface and its accumulation in intracellular structures that express markers of the Golgi complex and the recycling endosome. Unlike its role in virgin gland as a negative regulator cell proliferation, SCRIB is a positive regulator of mammary epithelial cell proliferation during pregnancy.

The Scribble family in cancer: twentieth anniversary

“…Finally, lano knock-out mice are viable and fertile with a normal Mendelian distribution [80]. However, as in scribble and erbin-deficient mice [76,81,82], we observed a delay in the development of the mammary epithelium in these animals (unpublished observations).…”

RAB6 GTPase regulates mammary secretory function by controlling the activation of STAT5

“…To investigate the possible molecular mechanisms underlying the reduced STAT5 activation observed upon Rab6a deletion, we took advantage of T47-D, a human breast cancer-derived cell line commonly used for studying PRL-induced signaling events ( Johnson et al, 2010 ; Baker et al, 2016 ; Oakes et al, 2017 ). Double staining for RAB6 and GM130 confirmed the expected localization of RAB6 in the Golgi of T47-D cells ( …”

“…Studies on PRLR signaling in the mouse mammary gland have been hampered by the lack of reliable antibodies working in immunohistochemistry, immunocytochemistry or immunoblotting ( Brisken and Ataca, 2015 ). To circumvent this difficulty, we used the well-characterized PRL-responding human T47-D cells ( Johnson et al, 2010 ; Baker et al, 2016 ; Oakes et al, 2017 ). Using siRNAs, we found that RAB6A depletion in this model resulted in decreased PRLR levels, altered membrane expression and reduced STAT5 activation downstream of PRL action.…”

“…The human mammary epithelial cell line T47-D was kindly provided by Dr V. Goffin (INSERM UMR_S1151-CNRS UMR8253, Institut Necker-Enfants Malades, Paris, France) and tested for contamination. Cells were maintained at 5% CO 2 in a 37°C incubator and grown in a phenol red-free RPMI 1640 medium containing 10% FBS and 5 µg/ml insulin, as previously described ( Baker et al, 2016 ).…”

“…Cells were washed in PBS 24 h before PRL treatment and medium was replaced by a serum-free medium. Cells were treated with human PRL (Sigma Aldrich, #L4021) at 250 ng/ml for 5, 15 and 45 min at 37°C in a medium supplemented with 5 µg/ml insulin and 1 µM dexamethasone, as previously described ( Baker et al, 2016 ).…”