Structural determinants in the C-terminal domain of apolipoprotein E mediating binding to the protein core of human aortic biglycan.

Abstract Apolipoprotein (apo) E-containing high density lipoprotein particles were reported to interact in vitrowith the proteoglycan biglycan (Bg), but the direct participation of apoE in this binding was not defined. To this end, we examined thein vitro binding of apoE complexed with dimyristoylphosphatidylcholine (DMPC) to human aortic Bg before and after glycosaminoglycan (GAG) depletion. In a solid-phase assay, apoE·DMPC bound to Bg and GAG-depleted protein core in a similar manner, suggesting a protein-protein mode of interaction. The binding was decreased in the presence of 1 m NaCl and was partially inhibited by either positively (0.2 m lysine, arginine) or negatively charged (0.2 m aspartic, glutamic) amino acids. A recombinant apoE fragment representing the C-terminal 10-kDa domain, complexed with DMPC, bound as efficiently as full-length apoE, whereas the N-terminal 22-kDa domain was inactive. Similar results were obtained with a gel mobility shift assay. Competition studies using a series of recombinant truncated apoEs showed that the charged segment in the C-terminal domain between residues 223 and 230 was involved in the binding. Overall, our results demonstrate that the C-terminal domain contains elements critical for the binding of apoE to the Bg protein core and that this binding is ionic in nature and independent of GAGs

Apical-Basal Polarity Signaling Components, Lgl1 and aPKCs, Control Glutamatergic Synapse Number and Function.

Normal synapse formation is fundamental to brain function. We show here that an apical-basal polarity (A-BP) protein, Lgl1, is present in the postsynaptic density and negatively regulates glutamatergic synapse numbers by antagonizing the atypical protein kinase Cs (aPKCs). A planar cell polarity protein, Vangl2, which inhibits synapse formation, was decreased in synaptosome fractions of cultured cortical neurons from Lgl1 knockout embryos. Conditional knockout of Lgl1 in pyramidal neurons led to reduction of AMPA/NMDA ratio and impaired plasticity. Lgl1 is frequently deleted in Smith-Magenis syndrome (SMS). Lgl1 conditional knockout led to increased locomotion, impaired novel object recognition and social interaction. Lgl1+/- animals also showed increased synapse numbers, defects in open field and social interaction, as well as stereotyped repetitive behavior. Social interaction in Lgl1+/- could be rescued by NMDA antagonists. Our findings reveal a role of apical-basal polarity proteins in glutamatergic synapse development and function and also suggest a potential treatment for SMS patients with Lgl1 deletion

ERG finally has something to YAP about in prostate cancer

SummaryThe significance of ERG in human prostate cancer is unclear because mouse prostate is resistant to ERG-mediated transformation. We determined that ERG activates the transcriptional program regulated by YAP1 of the Hippo signaling pathway and found that prostate-specific activation of either ERG or YAP1 in mice induces similar transcriptional changes and results in age-related prostate tumors. ERG binds to chromatin regions occupied by TEAD/YAP1 and transactivates Hippo target genes. In addition, in human luminal-type prostate cancer cells, ERG binds to the promoter of YAP1 and is necessary for YAP1 expression. These results provide direct genetic evidence of a causal role for ERG in prostate cancer and reveal a connection between ERG and the Hippo signaling pathway

Hepsin regulates TGF beta signaling via fibronectin proteolysis

Transforming growth factor-beta (TGF beta) is a multifunctional cytokine with a well-established role in mammary gland development and both oncogenic and tumor-suppressive functions. The extracellular matrix (ECM) indirectly regulates TGF beta activity by acting as a storage compartment of latent-TGF beta, but how TGF beta is released from the ECM via proteolytic mechanisms remains largely unknown. In this study, we demonstrate that hepsin, a type II transmembrane protease overexpressed in 70% of breast tumors, promotes canonical TGF beta signaling through the release of latent-TGF beta from the ECM storage compartment. Mammary glands in hepsin CRISPR knockout mice showed reduced TGF beta signaling and increased epithelial branching, accompanied by increased levels of fibronectin and latent-TGF beta 1, while overexpression of hepsin in mammary tumors increased TGF beta signaling. Cell-free and cell-based experiments showed that hepsin is capable of direct proteolytic cleavage of fibronectin but not latent-TGF beta and, importantly, that the ability of hepsin to activate TGF beta signaling is dependent on fibronectin. Altogether, this study demonstrates a role for hepsin as a regulator of the TGF beta pathway in the mammary gland via a novel mechanism involving proteolytic downmodulation of fibronectin.Peer reviewe

Cadherin signaling: keeping cells in touch [v1; ref status: indexed, http://f1000r.es/5c4]

Cadherin-catenin complexes are critical for the assembly of cell-cell adhesion structures known as adherens junctions. In addition to the mechanical linkage of neighboring cells to each other, these cell-cell adhesion protein complexes have recently emerged as important sensors and transmitters of the extracellular cues inside the cell body and into the nucleus. In the past few years, multiple studies have identified a connection between the cadherin-catenin protein complexes and major intracellular signaling pathways. Those studies are the main focus of this review

Cadherin-catenin proteins in vertebrate development.

Cadherin-catenin adhesion is pivotal for the development of multicellular organisms. Features such as a large repertoire of homotypically interacting cadherins, rapid assembly and disassembly, and a connection to a force-generating actin cytoskeleton make cadherin-mediated junctions ideal structures for the execution of complex changes in cell and tissue morphology during development. Recent findings highlight the role of cadherin-catenin proteins as critical regulators of major developmental pathways. We re-evaluate the significance of cadherin-catenin adhesion structures and propose that in addition to intercellular adhesion, they may be used as biosensors of the external cellular environment that help adjust the behavior of individual cells to ensure survival of the entire organism

Characterization of reactions catalysed by yeast phosphatidylinositol synthase

The nature of reactions catalysed by yeast phosphatidylinositol synthase expressed in E. coli has been investigated. The single enzyme is shown to carry both CDP-diacylglycerol-dependent incorporation of inositol into phosphatidylinositol (Km for inositol of 0.090 mM) and a CDP-diacylglycerol-independent exchange reaction between phosphatidylinositol and inositol (Km for inositol of 0.066 mM). The exchange reaction and reversal of phosphatidylinositol synthase were both stimulated by CMP, but had different optimum pH and requirements for substrates. These results suggest that CMP-stimulated exchange and CMP-dependent reverse reactions are distinct processes catalysed by the same enzyme, phosphatidylinositol synthase

alphaE-catenin is not a significant regulator of beta-catenin signaling in the developing mammalian brain.

beta-Catenin is a crucial mediator of the canonical Wnt-signaling pathway. alpha-catenin is a major beta-catenin-binding protein, and overexpressed alpha-catenin can negatively regulate beta-catenin activity. Thus, alpha-catenin may be an important modulator of the Wnt pathway. We show here that endogenous alpha-catenin has little impact on the transcriptional activity of beta-catenin in developing mammalian organisms. We analyzed beta-catenin signaling in mice with conditional deletion of alphaE-catenin (Ctnna1) in the developing central nervous system. This mutation results in brain hyperplasia and we investigated whether activation of beta-catenin signaling may be at least partially responsible for this phenotype. To reveal potential quantitative or spatial changes in beta-catenin signaling, we used mice carrying a beta-catenin-signaling reporter transgene. In addition, we analyzed the expression of known endogenous targets of the beta-catenin pathway and the amount and localization of beta-catenin in mutant progenitor cells. We found that although loss of alphaE-catenin resulted in disruption of intercellular adhesion and hyperplasia in the developing brain, beta-catenin signaling was not altered. We conclude that endogenous alphaE-catenin has no significant impact on beta-catenin transcriptional activities in the developing mammalian brain

alphaE-catenin controls cerebral cortical size by regulating the hedgehog signaling pathway.

During development, cells monitor and adjust their rates of accumulation to produce organs of predetermined size. We show here that central nervous system-specific deletion of the essential adherens junction gene, alphaE-catenin, causes abnormal activation of the hedgehog pathway, resulting in shortening of the cell cycle, decreased apoptosis, and cortical hyperplasia. We propose that alphaE-catenin connects cell-density-dependent adherens junctions with the developmental hedgehog pathway and that this connection may provide a negative feedback loop controlling the size of developing cerebral cortex