Regulation of formin INF2 and its alteration in INF2-linked inherited disorders.

Formins are proteins that catalyze the formation of linear filaments made of actin. INF2, a formin, is crucial for correct vesicular transport, microtubule stability and mitochondrial division. Its activity is regulated by a complex of cyclase-associated protein and lysine-acetylated G-actin (KAc-actin), which helps INF2 adopt an inactive conformation through the association of its N-terminal diaphanous inhibitory domain (DID) with its C-terminal diaphanous autoinhibitory domain.

INF2 formin variants linked to human inherited kidney disease reprogram the transcriptome, causing mitotic chaos and cell death.

Mutations in the human INF2 gene cause autosomal dominant focal segmental glomerulosclerosis (FSGS)-a condition characterized by podocyte loss, scarring, and subsequent kidney degeneration. To understand INF2-linked pathogenicity, we examined the effect of pathogenic INF2 on renal epithelial cell lines and human primary podocytes. Our study revealed an increased incidence of mitotic cells with surplus microtubule-organizing centers fostering multipolar spindle assembly, leading to nuclear abnormalities, particularly multi-micronucleation.

The MAL Family of Proteins: Normal Function, Expression in Cancer, and Potential Use as Cancer Biomarkers.

The MAL family of integral membrane proteins consists of MAL, MAL2, MALL, PLLP, CMTM8, MYADM, and MYADML2. The best characterized members are elements of the machinery that controls specialized pathways of membrane traffic and cell signaling. This review aims to help answer the following questions about the -family genes: (i) is their expression regulated in cancer and, if so, how? (ii) What role do they play in cancer? (iii) Might they have biomedical applications? Analysis of large-scale gene expression datasets indicated altered levels of -family transcripts in specific cancer types.

Structure and function of the N-terminal extension of the formin INF2.

In INF2-a formin linked to inherited renal and neurological disease in humans-the DID is preceded by a short N-terminal extension of unknown structure and function. INF2 activation is achieved by Ca-dependent association of calmodulin (CaM). Here, we show that the N-terminal extension of INF2 is organized into two α-helices, the first of which is necessary to maintain the perinuclear F-actin ring and normal cytosolic F-actin content.

MALL, a membrane-tetra-spanning proteolipid overexpressed in cancer, is present in membraneless nuclear biomolecular condensates.

Proteolipids are proteins with unusual lipid-like properties. It has long been established that PLP and plasmolipin, which are two unrelated membrane-tetra-spanning myelin proteolipids, can be converted in vitro into a water-soluble form with a distinct conformation, raising the question of whether these, or other similar proteolipids, can adopt two different conformations in the cell to adapt their structure to distinct environments. Here, we show that MALL, another proteolipid with a membrane-tetra-spanning structure, distributes in membranes outside the nucleus and, within the nucleus, in membrane-less, liquid-like PML body biomolecular condensates.

Plasmolipin regulates basolateral-to-apical transcytosis of ICAM-1 and leukocyte adhesion in polarized hepatic epithelial cells.

Apical localization of Intercellular Adhesion Receptor (ICAM)-1 regulates the adhesion and guidance of leukocytes across polarized epithelial barriers. Here, we investigate the molecular mechanisms that determine ICAM-1 localization into apical membrane domains of polarized hepatic epithelial cells, and their effect on lymphocyte-hepatic epithelial cell interaction. We had previously shown that segregation of ICAM-1 into apical membrane domains, which form bile canaliculi and bile ducts in hepatic epithelial cells, requires basolateral-to-apical transcytosis.

The MAL Protein, an Integral Component of Specialized Membranes, in Normal Cells and Cancer.

The gene encodes a 17-kDa protein containing four putative transmembrane segments whose expression is restricted to human T cells, polarized epithelial cells and myelin-forming cells. The MAL protein has two unusual biochemical features. First, it has lipid-like properties that qualify it as a member of the group of proteolipid proteins.

A Model for Primary Cilium Biogenesis by Polarized Epithelial Cells: Role of the Midbody Remnant and Associated Specialized Membranes.

Primary cilia are solitary, microtubule-based protrusions surrounded by a ciliary membrane equipped with selected receptors that orchestrate important signaling pathways that control cell growth, differentiation, development and homeostasis. Depending on the cell type, primary cilium assembly takes place intracellularly or at the cell surface. The intracellular route has been the focus of research on primary cilium biogenesis, whereas the route that occurs at the cell surface, which we call the "alternative" route, has been much less thoroughly characterized.

Midbody Remnant Inheritance Is Regulated by the ESCRT Subunit CHMP4C.

The inheritance of the midbody remnant (MBR) breaks the symmetry of the two daughter cells, with functional consequences for lumen and primary cilium formation by polarized epithelial cells, and also for development and differentiation. However, despite its importance, neither the relationship between the plasma membrane and the inherited MBR nor the mechanism of MBR inheritance is well known. Here, the analysis by correlative light and ultra-high-resolution scanning electron microscopy reveals a membranous stalk that physically connects the MBR to the apical membrane of epithelial cells.

Compensatory increase of VE-cadherin expression through ETS1 regulates endothelial barrier function in response to TNFα.

VE-cadherin plays a central role in controlling endothelial barrier function, which is transiently disrupted by proinflammatory cytokines such as tumor necrosis factor (TNFα). Here we show that human endothelial cells compensate VE-cadherin degradation in response to TNFα by inducing VE-cadherin de novo synthesis. This compensation increases adherens junction turnover but maintains surface VE-cadherin levels constant.

Caveolin-1α regulates primary cilium length by controlling RhoA GTPase activity.

The primary cilium is a single non-motile protrusion of the plasma membrane of most types of mammalian cell. The structure, length and function of the primary cilium must be tightly controlled because their dysfunction is associated with disease. Caveolin 1 (Cav1), which is best known as a component of membrane invaginations called caveolae, is also present in non-caveolar membrane domains whose function is beginning to be understood.

The actin-MRTF-SRF transcriptional circuit controls tubulin acetylation via gene expression.

The role of formins in microtubules is not well understood. In this study, we have investigated the mechanism by which INF2, a formin mutated in degenerative renal and neurological hereditary disorders, controls microtubule acetylation. We found that silencing of INF2 in epithelial RPE-1 cells produced a dramatic drop in tubulin acetylation, increased the G-actin/F-actin ratio, and impaired myocardin-related transcription factor (MRTF)/serum response factor (SRF)-dependent transcription, which is known to be repressed by increased levels of G-actin.

Alternative polyadenylation in a family of paralogous EPB41 genes generates protein 4.1 diversity.

Alternative polyadenylation (APA) is a step in mRNA 3'-end processing that contributes to the complexity of the transcriptome by generating isoforms that differ in either their coding sequence or their 3'-untranslated regions (UTRs). The EPB41 genes, EPB41, EPB41L2, EPB41L3 and EPB41L1, encode an impressively complex array of structural adaptor proteins (designated 4.1R, 4.

Novel role for the midbody in primary ciliogenesis by polarized epithelial cells.

The primary cilium is a membrane protrusion that is crucial for vertebrate tissue homeostasis and development. Here, we investigated the uncharacterized process of primary ciliogenesis in polarized epithelial cells. We show that after cytokinesis, the midbody is inherited by one of the daughter cells as a remnant that initially locates peripherally at the apical surface of one of the daughter cells.

RhoB controls endothelial barrier recovery by inhibiting Rac1 trafficking to the cell border.

Endothelial barrier dysfunction underlies chronic inflammatory diseases. In searching for new proteins essential to the human endothelial inflammatory response, we have found that the endosomal GTPase RhoB is up-regulated in response to inflammatory cytokines and expressed in the endothelium of some chronically inflamed tissues. We show that although RhoB and the related RhoA and RhoC play additive and redundant roles in various aspects of endothelial barrier function, RhoB specifically inhibits barrier restoration after acute cell contraction by preventing plasma membrane extension.

The MAL protein is crucial for proper membrane condensation at the ciliary base, which is required for primary cilium elongation.

The base of the primary cilium contains a zone of condensed membranes whose importance is not known. Here, we have studied the involvement of MAL, a tetraspanning protein that exclusively partitions into condensed membrane fractions, in the condensation of membranes at the ciliary base and investigated the importance of these membranes in primary cilium formation. We show that MAL accumulates at the ciliary base of epithelial MDCK cells.

Apicobasal polarity controls lymphocyte adhesion to hepatic epithelial cells.

Loss of apicobasal polarity is a hallmark of epithelial pathologies. Leukocyte infiltration and crosstalk with dysfunctional epithelial barriers are crucial for the inflammatory response. Here, we show that apicobasal architecture regulates the adhesion between hepatic epithelial cells and lymphocytes.

Protein 4.1R binds to CLASP2 and regulates dynamics, organization and attachment of microtubules to the cell cortex.

The microtubule (MT) cytoskeleton is essential for many cellular processes, including cell polarity and migration. Cortical platforms, formed by a subset of MT plus-end-tracking proteins, such as CLASP2, and non-MT binding proteins such as LL5β, attach distal ends of MTs to the cell cortex. However, the mechanisms involved in organizing these platforms have not yet been described in detail.

MYADM controls endothelial barrier function through ERM-dependent regulation of ICAM-1 expression.

The endothelium maintains a barrier between blood and tissue that becomes more permeable during inflammation. Membrane rafts are ordered assemblies of cholesterol, glycolipids, and proteins that modulate proinflammatory cell signaling and barrier function. In epithelial cells, the MAL family members MAL, MAL2, and myeloid-associated differentiation marker (MYADM) regulate the function and dynamics of ordered membrane domains.

INF2 promotes the formation of detyrosinated microtubules necessary for centrosome reorientation in T cells.

T cell antigen receptor-proximal signaling components, Rho-family GTPases, and formin proteins DIA1 and FMNL1 have been implicated in centrosome reorientation to the immunological synapse of T lymphocytes. However, the role of these molecules in the reorientation process is not yet defined. Here we find that a subset of microtubules became rapidly stabilized and that their α-tubulin subunit posttranslationally detyrosinated after engagement of the T cell receptor.

Crosstalk between reticular adherens junctions and platelet endothelial cell adhesion molecule-1 regulates endothelial barrier function.

Objective: Endothelial cells provide a barrier between the blood and tissues, which is reduced during inflammation to allow selective passage of molecules and cells. Adherens junctions (AJ) play a central role in regulating this barrier. We aim to investigate the role of a distinctive 3-dimensional reticular network of AJ found in the endothelium.

Protein 4.1R regulates cell migration and IQGAP1 recruitment to the leading edge.

In red blood cells, multifunctional protein 4.1R stabilizes the spectrin-actin network and anchors it to the plasma membrane. To contribute to the characterization of functional roles of 4.

MYADM regulates Rac1 targeting to ordered membranes required for cell spreading and migration.

Membrane organization into condensed domains or rafts provides molecular platforms for selective recruitment of proteins. Cell migration is a general process that requires spatiotemporal targeting of Rac1 to membrane rafts. The protein machinery responsible for making rafts competent to recruit Rac1 remains elusive.