Polarized deposition of basement membrane proteins depends on Phosphatidylinositol synthase and the levels of Phosphatidylinositol 4,5-bisphosphate

Abstract

The basement membrane (BM), a specialized sheet of the extracellular matrix contacting the basal side of epithelial tissues, plays an important role in the control of the polarized structure of epithelial cells. However, little is known about how BM proteins themselves achieve a polarized distribution. Here, we identify phosphatidylinositol 4,5-bisphosphate (PIP2) as a critical regulator of the polarized secretion of BM proteins. A decrease of PIP2 levels, in particular through mutations in Phosphatidylinositol synthase (Pis) and other members of the phosphoinositide pathway, leads to the aberrant accumulation of BM components at the apical side of the cell without primarily affecting the distribution of apical and basolateral polarity proteins. In addition, PIP2 controls the apical and lateral localization of Crag (Calmodulin-binding protein related to a Rab3 GDP/GTP exchange protein), a factor specifically required to prevent aberrant apical secretion of BM. We propose that PIP2, through the control of Crag's subcellular localization, restricts the secretion of BM proteins to the basal side.cell polarity | PTEN | PIK | Drosophila | oogenesis E pithelial cells are characterized by their polarized architecture, which enables them to exert their varied functions in embryonic and adult organisms. Epithelia exhibit a profound apical-basal polarity that is manifested in the cytoplasmic and surface organization of individual cells (1-3). Loss of apicalbasal cell polarity is often associated with carcinoma progression and tumor metastasis (4, 5). The establishment and maintenance of cell polarity relies on the transport of newly synthesized and recycled proteins to their correct destinations (6, 7). The lipid composition of the transport vesicles and of the plasma membrane is crucial for the establishment and maintenance of cell polarity (6-9). In particular, in 3D in vitro culture of Madin-Darby canine kidney (MDCK) cells, phosphatidylinositol 4,5-bisphosphate (PIP2) and phosphatidylinositol 3,4,5-trisphosphate (PIP3), two phosphoinositides (PtdIns) have been shown to play critical roles in polarized vesicle trafficking by mediating the recruitment of proteins to these different domains (10, 11).To set up a correct cell polarity, membrane asymmetry needs to be established. In cell culture, and likely during development of many tissues in multicellular organisms, this process is achieved by two external cues: one provided by the adjacent cells via cadherin-dependent adhesion and the other by interaction with the basement membrane (BM), a specialized sheet of the ECM secreted basally by the epithelial cells (12, 13). The main components of the BM are secreted glycoproteins, such as collagen IV (Coll IV), laminin, and the heparan sulfate proteoglycan perlecan (Pcan) (14), which interact with different membrane receptors, including integrin and dystroglycan (14,15). Previous studies in model organisms and 3D culture models have shown that BM secreted by the epithelial cells at their basal side plays a role as an initial ...

Companion Blood Cells Control Ovarian Stem Cell Niche Microenvironment and Homeostasis

“…Drosophila ColIV consists of two a chains, a-1 (Cg25c) and a-2 (Vkg), which form heterotrimers (Blumberg et al, 1988;Lunstrum et al, 1988) that are post-translationally modified by lysyl and prolyl hydroxylases enzymes, ultimately forming a functional BM (Bunt et al, 2010;Lerner et al, 2013;Pastor-Pareja and Xu, 2011). In adult ovaries, somatic follicular cells produce (legend continued on next page) ColIV, which is basally assembled into an egg chamber BM network through a Crag/Rab10/PIs-dependent mechanism (Denef et al, 2008;Devergne et al, 2014;Lerner et al, 2013;Medioni and Noselli, 2005). This specific ColIV plays an essential role in the establishment and maintenance of follicle cell polarity and in promoting egg elongation (Haigo and Bilder, 2011;Horne-Badovinac et al, 2012).…”

Building from the Ground up

“…This possibility is appealing as it aligns well with the likely function of Rab10. Alternatively, Scarface could function at the apical plasma membrane to prevent targeting of BM-containing vesicles to this domain, a function that has also been proposed for Crag (Denef et al, 2008; Devergne et al, 2014). Finally, Scarface could remove BM proteins from the apical surface either by stimulating a proteolytic cascade or via endocytosis.…”

“…Two additional proteins have been identified that are required for polarized BM secretion, although the mechanisms by which they do so are less well understood. These are Phosphatidylinositol synthase (Pis), an enzyme involved in the production of phosphoinositides (Devergne, Tsung, Barcelo, & Schupbach, 2014), and Scarface, a secreted serine protease-like protein that lacks catalytic activity (Sorrosal et al, 2010). …”

“…Various phosphoinositide isoforms are created by kinase- and phosphatase-mediated interconversion between different phosphorylation states of a common phospholipid backbone, phosphatidylinositol. Pis synthesizes phosphatidylinositol, and is therefore required for formation of all phosphoinositides; however, the authors focused their analyses on the role of phosphatidylinositol 4,5-bisphosphate (PIP2) in polarized BM secretion (Devergne et al, 2014). PIP2 functions as an apical determinant in the regulation of epithelial polarity (Martin-Belmonte et al, 2007).…”

“…In the follicular epithelium, PIP2 is enriched on apical and lateral membranes. Loss of Pis or other PIP2 biosynthetic enzymes decreased PIP2 levels and caused a loss of Crag from apical and lateral membranes (Devergne et al, 2014). How the lateral and apical Crag populations would feed into basal protein secretion is difficult to say.…”