The endosomal trafficking regulator LITAF controls the cardiac Nav1.5 channel via the ubiquitin ligase NEDD4-2

The QT interval is a recording of cardiac electrical activity. Previous genome-wide association studies identified genetic variants that modify the QT interval upstream of LITAF (lipopolysaccharide-induced tumor necrosis factor-α factor), a protein encoding a regulator of endosomal trafficking. However, it was not clear how LITAF might impact cardiac excitation. We investigated the effect of LITAF on the voltage-gated sodium channel Nav1.5, which is critical for cardiac depolarization. We show that overexpressed LITAF resulted in a significant increase in the density of Nav1.5-generated voltage-gated sodium current I_Na and Nav1.5 surface protein levels in rabbit cardiomyocytes and in HEK cells stably expressing Nav1.5. Proximity ligation assays showed co-localization of endogenous LITAF and Nav1.5 in cardiomyocytes, whereas co-immunoprecipitations confirmed they are in the same complex when overexpressed in HEK cells. In vitro data suggest that LITAF interacts with the ubiquitin ligase NEDD4-2, a regulator of Nav1.5. LITAF overexpression down-regulated NEDD4-2 in cardiomyocytes and HEK cells. In HEK cells, LITAF increased ubiquitination and proteasomal degradation of co-expressed NEDD4-2 and significantly blunted the negative effect of NEDD4-2 on I_Na. We conclude that LITAF controls cardiac excitability by promoting degradation of NEDD4-2, which is essential for removal of surface Nav1.5. LITAF-knockout zebrafish showed increased variation in and a nonsignificant 15% prolongation of action potential duration. Computer simulations using a rabbit-cardiomyocyte model demonstrated that changes in Ca²⁺ and Na⁺ homeostasis are responsible for the surprisingly modest action potential duration shortening. These computational data thus corroborate findings from several genome-wide association studies that associated LITAF with QT interval variation.     

Bioactive molecules from Nocardia: diversity,bioactivities and biosynthesis

Journal of Industrial Microbiology & Biotechnology - Nocardia spp. are catalase positive, aerobic, and non-motile Gram-positive filamentous bacteria. Many Nocarida spp. have been reported as...     

Removal characteristics of metal ions using degreased coffee beans: adsorption equilibrium of cadmium(II)

The feasibility of using coffee beans after being dripped and degreased (DCB) as an adsorbent for base metals such as copper(II), zinc(II), lead(II), iron(III) and cadmium(II) were examined. The compositions of the DCB were characterized by Fourier transform infrared spectroscopy, scanning electronic micrograph and fluorescent X-ray. It was found that DCB contain sulfur and calcium from the analysis using fluorescent X-ray. The plant cell wall in DCB has the porous structure from the scanning electron microscopy (SEM) analysis, and the specific surface area was determined to be 1.2 m2/g using the specific surface area analyzer. Batch adsorption experiments on DCB were carried out at various pHs in order to elucidate the selectivity of metal ions. All metals were adsorbed at low pH region (3.0-5.0). Of particular interest was the adsorption characteristics of cadmium(II) on DCB. The adsorption isotherm for cadmium(II) at pH 8 fitted with a Langmuir equation to yield an adsorption equilibrium constant of 55.2 mmol dm(-3) and an adsorption capacity of 5.98 x 10(-2) mmol g(-1). The desorption of cadmium(II) was easily achieved over 90% by a single batchwise treatment with an aqueous solution of hydrochloric acid or nitric acid at more than 0.01 mol dm(-3). These results suggested that DCB behaves as a cation exchanger.     

Bone marrow-derived IL-13Rα1-positive thymic progenitors are restricted to the myeloid lineage

The earliest thymic progenitors (ETPs) were recently shown to give rise to both lymphoid and myeloid cells. Whereas the majority of ETPs are derived from IL-7Rα-positive cells and give rise exclusively to T cells, the origin of the myeloid cells remains undefined. In this study, we show both in vitro and in vivo that IL-13Rα1(+) ETPs yield myeloid cells with no potential for maturation into T cells, whereas IL-13Rα1(-) ETPs lack myeloid potential. Moreover, transfer of lineage-negative IL-13Rα1(+) bone marrow stem cells into IL-13Rα1-deficient mice reconstituted thymic IL-13Rα1(+) myeloid ETPs. Myeloid cells or macrophages in the thymus are regarded as phagocytic cells whose function is to clear apoptotic debris generated during T cell development. However, the myeloid cells derived from IL-13Rα1(+) ETPs were found to perform Ag-presenting functions. Thus, IL-13Rα1 defines a new class of myeloid restricted ETPs yielding APCs that could contribute to development of T cells and the control of immunity and autoimmunity.     

Expression of cyclooxygenase-2 in invasive breast carcinomas and its prognostic impact

Representative tumour sections from 468 patients with invasive breast cancer were immunostained for cyclooxygenase-2 (COX-2) and evaluated. The relationships between COX-2 expression, clinical outcome and various clinicopathological variables, including tumour vascularity and disseminated tumour cells (DTC) in the bone marrow were examined. COX-2 expression in invasive breast carcinoma cells was positively associated with oestrogen receptor and/or progesterone receptor positivity (p<0.001). Triple-negative tumours showed no/low COX-2 expression more frequently than other tumour types (p<0.001). Expression of COX-2 was not associated with breast cancer-specific survival (p=0.49, log-rank) or distant disease-free survival (p=0.67, log-rank) for all patients, including lymph node-negative, untreated patients (p>0.14, log-rank). There was also no significant association between COX-2 expression and histological grade, tumour size, nodal status, DTC in bone marrow, p53, HER2, or tumour vascularity. In conclusion, COX-2 expression in this series was associated with the presence of hormone receptors. Low COX-2 expression was observed in triple-negative breast carcinomas.     

The UPEC pore-forming toxin α-hemolysin triggers proteolysis of host proteins to disrupt cell adhesion, inflammatory, and survival pathways

Uropathogenic Escherichia coli (UPEC), which are the leading cause of both acute and chronic urinary tract infections, often secrete a labile pore-forming toxin known as α-hemolysin (HlyA). We show that stable insertion of HlyA into epithelial cell and macrophage membranes triggers degradation of the cytoskeletal scaffolding protein paxillin and other host regulatory proteins, as well as components of the proinflammatory NFκB signaling cascade. Proteolysis of these factors requires host serine proteases, and paxillin degradation specifically involves the serine protease mesotrypsin. The induced activation of mesotrypsin by HlyA is preceded by redistribution of mesotrypsin precursors from the cytosol into foci along microtubules and within nuclei. HlyA intoxication also stimulated caspase activation, which occurred independently of effects on host serine proteases. HlyA-induced proteolysis of host proteins likely allows UPEC to not only modulate epithelial cell?functions, but also disable macrophages and suppress inflammatory responses.     

Intramolecular folding in human ILPR fragment with three C-rich repeats

Enrichment of four tandem repeats of guanine (G) rich and cytosine (C) rich sequences in functionally important regions of human genome forebodes the biological implications of four-stranded DNA structures, such as G-quadruplex and i-motif, that can form in these sequences. However, there have been few reports on the intramolecular formation of non-B DNA structures in less than four tandem repeats of G or C rich sequences. Here, using mechanical unfolding at the single-molecule level, electrophoretic mobility shift assay (EMSA), circular dichroism (CD), and ultraviolet (UV) spectroscopy, we report an intramolecularly folded non-B DNA structure in three tandem cytosine rich repeats, 5'-TGTC4ACAC4TGTC4ACA (ILPR-I3), in the human insulin linked polymorphic region (ILPR). The thermal denaturation analyses of the sequences with systematic C to T mutations have suggested that the structure is linchpinned by a stack of hemiprotonated cytosine pairs between two terminal C4 tracts. Mechanical unfolding and Br(2) footprinting experiments on a mixture of the ILPR-I3 and a 5'-C4TGT fragment have further indicated that the structure serves as a building block for intermolecular i-motif formation. The existence of such a conformation under acidic or neutral pH complies with the strand-by-strand folding pathway of ILPR i-motif structures.     

Effect of different biosynthetic precursors on the production of nargenicin A1 from metabolically engineered Nocardia sp. CS682

Nargenicin A1 is a 28-membered polyketide macrolide, with antibacterial activity against methicillin-resistant Staphylococcus aureus, produced by Nocardia sp. CS682. In this study, the production of nargenicin A1 was improved by enhancing the supply of different biosynthetic precursors. In Nocardia sp. CS682 (KCTC11297BP), this improvement was ~4.62-fold with the supplementation of 30 mM methyl oleate, 4.25-fold with supplementation of 15 mM sodium propionate, and 2.81-fold with supplementation of 15 mM sodium acetate. In Nocardia sp. metK18 and Nocardia sp. CS682 expressing S-adenosylmethionine synthetase (MetK), the production of nargenicin A1 was improved by ~5.57-fold by supplementation with 30 mM methyl oleate, 5.01-fold by supplementation with 15 mM sodium propionate, and 3.64-fold by supplementation with 15 mM sodium acetate. Furthermore, supplementing the culture broth of Nocardia sp. ACC18 and Nocardia sp. CS682 expressing the acetyl-CoA carboxylase complex (AccA2 and AccBE) with 30 mM methyl oleate, 15 mM sodium propionate, or 15 mM sodium acetate resulted in ~6.99-, 6.46-, and 5.58-fold increases, respectively, in nargenicin A1 production. Our overall results showed that among the supplements, methyl oleate was the most effective precursor supporting the highest titers of nargenicin A1 in Nocardia sp. CS682, Nocardia sp. metK18, and Nocardia sp. ACC18.