Impact of serine protease inhibitor alpha1-antitrypsin on expression of endoplasmic reticulum stress-induced proinflammatory factors in adipocytes

Obesity-induced endoplasmic reticulum (ER) stress contributes to low-grade chronic inflammation in adipose tissue and may cause metabolic disorders such as diabetes mellitus and dyslipidemia. Identification of high serpina A1 (alpha-1 antitrypsin, A1AT) expression in mouse adipose tissue and adipocytes prompted us to explore the role of A1AT in the inflammatory response of adipocytes under ER stress. We aimed to determine the role of A1AT expression in adipocytes with ER stress during regulation of adipocyte homeostasis and inflammation. To this end, we chemically induced ER stress in A1AT small interfering RNA-transfected differentiating adipocytes using thapsigargin. Induction of CCAAT-enhancer-binding protein homologous protein (CHOP), an ER stress marker, by thapsigargin was lower in A1AT-deficient SW872 adipocytes. Thapsigargin or the proinflammatory cytokine tumor necrosis factor (TNF)α increased basal expression of cytokines such as interleukin (IL)-1β and IL-8 in both SW872 and primary omental adipocytes. This thapsigargin- or TNFα-induced expression of proinflammatory genes was increased by A1AT deficiency. These findings indicate that adipose A1AT may suppress the ER stress response to block excessive expression of proinflammatory factors, which suggests that A1AT protects against adipose tissue dysfunction associated with ER stress activation.     

The Deubiquitinating Enzyme AMSH1 and the ESCRT-III Subunit VPS2.1 Are Required for Autophagic Degradation in Arabidopsis

In eukaryotes, posttranslational modification by ubiquitin regulates the activity and stability of many proteins and thus influences a variety of developmental processes as well as environmental responses. Ubiquitination also plays a critical role in intracellular trafficking by serving as a signal for endocytosis. We have previously shown that the Arabidopsis thaliana ASSOCIATED MOLECULE WITH THE SH3 DOMAIN OF STAM3 (AMSH3) is a deubiquitinating enzyme (DUB) that interacts with ENDOSOMAL COMPLEX REQUIRED FOR TRANSPORT-III (ESCRT-III) and is essential for intracellular transport and vacuole biogenesis. However, physiological functions of AMSH3 in the context of its ESCRT-III interaction are not well understood due to the severe seedling lethal phenotype of its null mutant. In this article, we show that Arabidopsis AMSH1, an AMSH3-related DUB, interacts with the ESCRT-III subunit VACUOLAR PROTEIN SORTING2.1 (VPS2.1) and that impairment of both AMSH1 and VPS2.1 causes early senescence and hypersensitivity to artificial carbon starvation in the dark similar to previously reported autophagy mutants. Consistent with this, both mutants accumulate autophagosome markers and accumulate less autophagic bodies in the vacuole. Taken together, our results demonstrate that AMSH1 and the ESCRT-III-subunit VPS2.1 are important for autophagic degradation and autophagy-mediated physiological processes.     

Review ArticleNaphthalene Combretastatin Analogues: Synthesis,Cytotoxicity and Antitubulin Activity

Synthesis and evaluation of new combretastatin analogues with varied modifications on the bridge and the aromatic rings, have shown that the 2-naphthyl moiety is a good surrogate for the 3-hydroxy-4-methoxyphenyl (B-ring) of combretastatin A-4. Other bicyclic systems, such as 6(7)-quinolyl and 5-indolyl, can replace the B-ring, but they produce less potent analogues in the cytotoxicity and tubulin polymerization inhibition assays. Other modifications are detrimentral for the potency of the studied analogues. The 2-naphthyl combretastatin 53 and the related 6-quinolyl combretastatin 106 analogues are the most potent among the derivatives of this type, whereas 92 and 95 are the most potent among the naphthalene derivatives with a heterocycle in the bridge. Previous and new results in this family of combretastatin analogues are discussed.     

The conserved oligomeric Golgi complex is involved in penetration resistance of barley to the barley powdery mildew fungus

Membrane trafficking is vital to plant development and adaptation to the environment. It is suggested that post‐Golgi vesicles and multivesicular bodies are essential for plant defence against directly penetrating fungal parasites at the cell wall. However, the actual plant proteins involved in membrane transport for defence are largely unidentified. We applied a candidate gene approach and single cell transient‐induced gene silencing for the identification of membrane trafficking proteins of barley involved in the response to the fungal pathogen Blumeria graminis f.sp. hordei. This revealed potential components of vesicle tethering complexes [putative exocyst subunit HvEXO70F‐like and subunits of the conserved oligomeric Golgi (COG) complex] and Golgi membrane trafficking (COPIγ coatomer and HvYPT1‐like RAB GTPase) as essential for resistance to fungal penetration into the host cell.     

Negative Regulation of Notch Signaling by Xylose

The Notch signaling pathway controls a large number of processes during animal development and adult homeostasis. One of the conserved post-translational modifications of the Notch receptors is the addition of an O-linked glucose to epidermal growth factor-like (EGF) repeats with a C-X-S-X-(P/A)-C motif by Protein O-glucosyltransferase 1 (POGLUT1; Rumi in Drosophila). Genetic experiments in flies and mice, and in vivo structure-function analysis in flies indicate that O-glucose residues promote Notch signaling. The O-glucose residues on mammalian Notch1 and Notch2 proteins are efficiently extended by the addition of one or two xylose residues through the function of specific mammalian xylosyltransferases. However, the contribution of xylosylation to Notch signaling is not known. Here, we identify the Drosophila enzyme Shams responsible for the addition of xylose to O-glucose on EGF repeats. Surprisingly, loss- and gain-of-function experiments strongly suggest that xylose negatively regulates Notch signaling, opposite to the role played by glucose residues. Mass spectrometric analysis of Drosophila Notch indicates that addition of xylose to O-glucosylated Notch EGF repeats is limited to EGF14–20. A Notch transgene with mutations in the O-glucosylation sites of Notch EGF16–20 recapitulates the shams loss-of-function phenotypes, and suppresses the phenotypes caused by the overexpression of human xylosyltransferases. Antibody staining in animals with decreased Notch xylosylation indicates that xylose residues on EGF16–20 negatively regulate the surface expression of the Notch receptor. Our studies uncover a specific role for xylose in the regulation of the Drosophila Notch signaling, and suggest a previously unrecognized regulatory role for EGF16–20 of Notch.     

Fibronectin conformation regulates the proangiogenic capability of tumor-associated adipogenic stromal cells

Background

Changes in fibronectin (Fn) matrix remodeling contribute to mammary tumor angiogenesis and are related to altered behavior of adipogenic stromal cells; yet, the underlying mechanisms remain unclear due in part to a lack of reductionist model systems that allow the inherent complexity of cell-derived extracellular matrices (ECMs) to be deciphered. In particular, breast cancer-associated adipogenic stromal cells not only enhance the composition, quantity, and rigidity of deposited Fn, but also partially unfold these matrices. However, the specific effect of Fn conformation on tumor angiogenesis is undefined.

Methods

Decellularized matrices and a conducting polymer device consisting of poly(3,4-ethylenedioxythiophene) doped with poly(styrenesulfonate) (PEDOT:PSS) were used to examine the effect of Fn conformation on the behavior of 3T3-L1 preadipocytes. Changes in cell adhesion and proangiogenic capability were tested via cell counting and by quantification of vascular endothelial growth factor (VEGF) secretion, respectively. Integrin-blocking antibodies were utilized to examine varied integrin specificity as a potential mechanism.

Results

Our findings suggest that tumor-associated partial unfolding of Fn decreases adhesion while enhancing VEGF secretion by breast cancer-associated adipogenic precursor cells, and that altered integrin specificity may underlie these changes.

Conclusions and general significance

These results not only have important implications for our understanding of tumorigenesis, but also enhance knowledge of cell-ECM interactions that may be harnessed for other applications including advanced tissue engineering approaches. This article is part of a Special Issue entitled Organic Bioelectronics — Novel Applications in Biomedicine.