Heme oxygenase-1 attenuates the cisplatin-induced apoptosis of auditory cells via down-regulation of reactive oxygen species generation

Heme oxygenase-1 (HO-1), the rate-limiting enzyme of heme catabolism, is known to modulate various cellular functions, including cytokine production, cell proliferation, and apoptosis, in stress-related conditions. However, the role of HO-1 in the auditory system remains elusive. Herein, we demonstrate that pharmacologic induction of HO-1 along with catalytic activation significantly suppressed apoptosis of HEI-OC1 cells induced by cisplatin. Studies of ectopic expression of pcDNA3-HO-1 and siRNA of HO-1 further revealed the protective role of HO-1 against cisplatin in HEI-OC1 cells. Among the catabolic metabolites of HO-1, both carbon monoxide (CO) and bilirubin were directly involved in the protective role of HO-1 against cisplatin through inhibition of reactive oxygen species generation. Furthermore, pharmacological induction of HO-1 completely prevented the destruction of outer hair cell arrays by cisplatin through a CO-dependent mechanism in organotrophic culture of the rat primary organ of Corti explants. These results suggest that HO-1 may serve as a safeguard of auditory sensory hair cells against a variety of challenges of oxidative stress, including noise trauma, presbycusis, and ototoxic drugs, respectively.     

Self-illuminating quantum dot conjugates for in vivo imaging

Fluorescent semiconductor quantum dots hold great potential for molecular imaging in vivo. However, the utility of existing quantum dots for in vivo imaging is limited because they require excitation from external illumination sources to fluoresce, which results in a strong autofluorescence background and a paucity of excitation light at nonsuperficial locations. Here we present quantum dot conjugates that luminesce by bioluminescence resonance energy transfer in the absence of external excitation. The conjugates are prepared by coupling carboxylate-presenting quantum dots to a mutant of the bioluminescent protein Renilla reniformis luciferase. We show that the conjugates emit long-wavelength (from red to near-infrared) bioluminescent light in cells and in animals, even in deep tissues, and are suitable for multiplexed in vivo imaging. Compared with existing quantum dots, self-illuminating quantum dot conjugates have greatly enhanced sensitivity in small animal imaging, with an in vivo signal-to-background ratio of > 10(3) for 5 pmol of conjugate.     

Human C5aR knock-in mice facilitate the production and assessment of anti-inflammatory monoclonal antibodies

Complement component C5a binds C5a receptor (C5aR) and facilitates leukocyte chemotaxis and release of inflammatory mediators. We used neutrophils from human C5aR knock-in mice, in which the mouse C5aR coding region was replaced with that of human C5aR, to immunize wild-type mice and to generate high-affinity antagonist monoclonal antibodies (mAbs) to human C5aR. These mAbs blocked neutrophil migration to C5a in vitro and, at low doses, both prevented and reversed inflammatory arthritis in the murine K/BxN model. Of approximately 40 mAbs generated to C5aR, all potent inhibitors recognized a small region of the second extracellular loop that seems to be critical for regulation of receptor activity. Human C5aR knock-in mice not only facilitated production of high-affinity mAbs against an important human therapeutic target but were also useful in preclinical validation of the potency of these antagonists. This strategy should be applicable to other important mAb therapeutics.     

AnArabidopsis short root and dwarfism mutant defines a novel locus that mediates both cell division and elongation

Arabidopsis leaf morphology is determined by the coordinated action of cell division and elongation. Of all the hormones that control leaf shape, the brassinosteroids (BRs) are active components in this process. BRs are a group of plant-originated steroidal compounds that induce growth along the long axes of organs. Here, we report the isolation and characterization of a novel mutant,short root and dwarfism (srd). Its dwarf phenotype includes round and curled leaves, reduced fertility, and short hypocotyls in the light and dark. Dwarfism in the aerial portions and a short-root morphology are not rescued by exogenous application of BRs, suggesting thatsrd is not impaired in BR metabolic pathways. Anatomical analysis revealed thatsrd roots are much shorter and thicker than the wild type due to additional layers of cortical cells. A lack of cell elongation but an increase in division results in these short but horizontally swollen roots. A double mutantsrd/bri1-5 also displays the short-root phenotype, implying thatsrd is epistatic tobri1. Cloning and further characterization ofSRD should provide additional information about its role in the determination of leaf shape and root elongation.     

Modulation of airway remodeling and airway inflammation by peroxisome proliferator-activated receptor gamma in a murine model of toluene diisocyanate-induced asthma

Toluene diisocyanate (TDI) is a leading cause of occupational asthma. Although considerable controversy remains regarding its pathogenesis, TDI-induced asthma is an inflammatory disease of the airways characterized by airway remodeling. Peroxisome proliferator-activated receptor gamma (PPARgamma) has been shown to play a critical role in the control of airway inflammatory responses. However, no data are available on the role of PPARgamma in TDI-induced asthma. We have used a mouse model for TDI-induced asthma to determine the effect of PPARgamma agonist, rosiglitazone, or pioglitazone, and PPARgamma on TDI-induced bronchial inflammation and airway remodeling. This study with the TDI-induced model of asthma revealed the following typical pathophysiological features: increased numbers of inflammatory cells of the airways, airway hyperresponsiveness, increased levels of Th2 cytokines (IL-4, IL-5, and IL-13), adhesion molecules (ICAM-1 and VCAM-1), chemokines (RANTES and eotaxin), TGF-beta1, and NF-kappaB in nuclear protein extracts. In addition, the mice exposed to TDI developed features of airway remodeling, including thickening of the peribronchial smooth muscle layer, subepithelial collagen deposition, and increased airway mucus production. Administration of PPARgamma agonists or adenovirus carrying PPARgamma2 cDNA reduced the pathophysiological symptoms of asthma and decreased the increased levels of Th2 cytokines, adhesion molecules, chemokines, TGF-beta1, and NF-kappaB in nuclear protein extracts after TDI inhalation. In addition, inhibition of NF-kappaB activation decreased the increased levels of Th2 cytokines, adhesion molecules, chemokines, and TGF-beta1 after TDI inhalation. These findings demonstrate a protective role of PPARgamma in the pathogenesis of the TDI-induced asthma phenotype.     

Bis(O2S2 macrocycle) complexes of palladium(II)

Two isomeric dibenzo-O₂S₂ macrocycles L¹ and L² have been synthesised and their coordination chemistry towards palladium(II) has been investigated. Two-step approaches via reactions of 1:1-type complexes, [cis-Cl₂LPd] (1a: L = L¹, 1b: L = L²), with different O₂S₂ macrocycle systems (L¹ and L²) have led to the isolation of the following bis(O₂S₂ macrocycle) palladium(II) complexes in the solid state: [Pd(L¹)₂](ClO₄)₂ (2a) and a mixture of [Pd(L¹)₂](ClO₄)₂ (2a) + [Pd(L²)₂](ClO₄)₂ (2b).     

Autophosphorylation at serine 1981 stabilizes ATM at DNA damage sites

Ataxia telangiectasia mutated (ATM) plays a critical role in the cellular response to DNA damage. In response to DNA double-strand breaks (DSBs), ATM is autophosphorylated at serine 1981. Although this autophosphorylation is widely considered a sign of ATM activation, it is still not clear if autophosphorylation is required for ATM functions including localization to DSBs and activation of ATM kinase activity. In this study, we show that localization of ATM to DSBs is differentially regulated with the initial localization requiring the MRE11–RAD50–NBS1 complex and sustained retention requiring autophosphorylation of ATM at serine 1981. Autophosphorylated ATM interacts with MDC1 and the latter is required for the prolonged association of ATM to DSBs. Ablation of ATM autophosphorylation or knock-down of MDC1 protein affects the ability of ATM to phosphorylate downstream substrates and confer radioresistance. Together, these data suggest that autophosphorylation at serine 1981 stabilizes ATM at the sites of DSBs, and this is required for a proper DNA damage response.     

Bcl-rambo induces apoptosis via interaction with the adenine nucleotide translocator

The Bcl-2 family proteins plays a central role in apoptosis. The pro- or anti-apoptotic activities of Bcl-2 family are dependent on the Bcl-2 homology (BH) regions. Bcl-rambo, a new pro-apoptotic member, is unusual in that its pro-apoptotic activity is independent of its BH domains. However, the mechanism underlying Bcl-rambo-induced apoptosis is largely unknown. Mitochondrial localization is indispensable for the pro-apoptotic function of Bcl-rambo. Bcl-rambo interacts physically with the adenine nucleotide translocator (ANT), suppresses the ADT/ATP-dependent translocation activity of ANT. Collectively, our data indicate Bcl-rambo is a pro-apoptotic member of the Bcl-2 family, induces the permeability transition via interaction with ANT.

Structured summary of protein interactions:

Bcl-Rambo and HSP60colocalize by fluorescence microscopy (View interaction)Bcl-rambobinds to ANT1 by pull down (View interaction)     

G Protein-coupled receptor kinase 5 is localized to centrosomes and regulates cell cycle progression

G protein-coupled receptor kinases (GRKs) are important regulators of G protein-coupled receptor function and mediate receptor desensitization, internalization, and signaling. While GRKs also interact with and/or phosphorylate many other proteins and modify their function, relatively little is known about the cellular localization of endogenous GRKs. Here we report that GRK5 co-localizes with γ-tubulin, centrin, and pericentrin in centrosomes. The centrosomal localization of GRK5 is observed predominantly at interphase and although its localization is not dependent on microtubules, it can mediate microtubule nucleation of centrosomes. Knockdown of GRK5 expression leads to G2/M arrest, characterized by a prolonged G2 phase, which can be rescued by expression of wild type but not catalytically inactive GRK5. This G2/M arrest appears to be due to increased expression of p53, reduced activity of aurora A kinase and a subsequent delay in the activation of polo-like kinase 1. Overall, these studies demonstrate that GRK5 is localized in the centrosome and regulates microtubule nucleation and normal cell cycle progression.