We found that the autophagic machinery could effectively eliminate pathogenic group A Streptococcus (GAS) within nonphagocytic cells. After escaping from endosomes into the cytoplasm, GAS became enveloped by autophagosome-like compartments and were killed upon fusion of these compartments with lysosomes. In autophagy-deficient Atg5-/- cells, GAS survived, multiplied, and were released from the cells. Thus, the autophagic machinery can act as an innate defense system against invading pathogens.

Mutant ␣ 1 -antitrypsin Z (␣ 1 -ATZ) protein, which has a tendency to form aggregated polymers as it accumulates within the endoplasmic reticulum of the liver cells, is associated with the development of chronic liver injury and hepatocellular carcinoma in hereditary ␣ 1 -antitrypsin (␣ 1 -AT) deficiency. Previous studies have suggested that efficient intracellular degradation of ␣ 1 -ATZ is correlated with protection from liver disease in ␣ 1 -AT deficiency and that the ubiquitin-proteasome system accounts for a major route, but not the sole route, of ␣ 1 -ATZ disposal. Yet another intracellular degradation system, autophagy, has also been implicated in the pathophysiology of ␣ 1 -AT deficiency. To provide genetic evidence for autophagy-mediated disposal of ␣ 1 -ATZ, here we used cell lines deleted for the Atg5 gene that is necessary for initiation of autophagy. In the absence of autophagy, the degradation of ␣ 1 -ATZ was retarded, and the characteristic cellular inclusions of ␣ 1 -ATZ accumulated. In wild-type cells, colocalization of the autophagosomal membrane marker GFP-LC3 and ␣ 1 -ATZ was observed, and this colocalization was enhanced when clearance of autophagosomes was prevented by inhibiting fusion between autophagosome and lysosome. By using a transgenic mouse with liver-specific inducible expression of ␣ 1 -ATZ mated to the GFP-LC3 mouse, we also found that expression of ␣ 1 -ATZ in the liver in vivo is sufficient to induce autophagy. These data provide definitive evidence that autophagy can participate in the quality control/degradative pathway for ␣ 1 -ATZ and suggest that autophagic degradation plays a fundamental role in preventing toxic accumulation of ␣ 1 -ATZ.3 a monomeric 394-amino acid glycoprotein, is synthesized and secreted primarily by liver cells. It is a prototypic member of serine protease inhibitor (serpin) superfamily proteins and the most abundant of the circulating serpins. The principal role of ␣ 1 -AT in serum is to protect lung tissues from destructive proteases (elastase, cathepsin G, and proteinase 3) released by neutrophils during inflammation. Some genetic alterations in ␣ 1 -AT are responsible for defective secretion and thus cause serum ␣ 1 -AT deficiency (1-3). The most common causal mutation found in Caucasian populations is the replacement of Glu-342 by Lys that characterizes the Z mutant of ␣ 1 -AT (␣ 1 -ATZ). This substitution is sufficient to cause an abnormality in folding early in the secretory pathway with retention of the mutant ␣ 1 -ATZ molecule in the ER of liver cells. Homozygotes for the ␣ 1 -ATZ mutation (PIZZ) are characterized by serum levels of ␣ 1 -AT that are ϳ10 -15% of those in the general population and are susceptible to two major target organ injuries. Destructive lung disease/emphysema in adults is due to a loss-of-function mechanism. Chronic liver disease often first discovered in childhood, but also affecting adults, is due to a gain-of-toxic-function mechanism in which liver cell injury results from the hepatotoxic effects of retained ␣ 1 -ATZ...

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BackgroundMicroRNA (miRNA) is an emerging subclass of small non-coding RNAs that regulates gene expression and has a pivotal role for many physiological processes including cancer development. Recent reports revealed the role of miRNAs as ideal biomarkers and therapeutic targets due to their tissue- or disease-specific nature. Head and neck cancer (HNC) is a major cause of cancer-related mortality and morbidity, and laryngeal cancer has the highest incidence in it. However, the molecular mechanisms involved in laryngeal cancer development remain to be known and highly sensitive biomarkers and novel promising therapy is necessary.Methodology/Principal FindingsTo explore laryngeal cancer-specific miRNAs, RNA from 5 laryngeal surgical specimens including cancer and non-cancer tissues were hybridized to microarray carrying 723 human miRNAs. The resultant differentially expressed miRNAs were further tested by using quantitative real time PCR (qRT-PCR) on 43 laryngeal tissue samples including cancers, noncancerous counterparts, benign diseases and precancerous dysplasias. Significant expressional differences between matched pairs were reproduced in miR-133b, miR-455-5p, and miR-196a, among which miR-196a being the most promising cancer biomarker as validated by qRT-PCR analyses on additional 84 tissue samples. Deep sequencing analysis revealed both quantitative and qualitative deviation of miR-196a isomiR expression in laryngeal cancer. In situ hybridization confirmed laryngeal cancer-specific expression of miR-196a in both cancer and cancer stroma cells. Finally, inhibition of miR-196a counteracted cancer cell proliferation in both laryngeal cancer-derived cells and mouse xenograft model.Conclusions/SignificanceOur study provided the possibilities that miR-196a might be very useful in diagnosing and treating laryngeal cancer.

We have cloned human dymple, a novel dynamin family member. The full-length cDNA sequence encodes a protein composed of 736 amino acids with a molecular mass of 80 kDa. This amino acid sequence most resembles yeast DNM1P and VPS1P. Dymple lacks a prolinerich carboxyl-terminal domain through which dynamin binds to SH3 domains to be activated. Northern blot analysis revealed two transcript sizes of 2.5 and 4.2 kilobases with alternative polyadenylation at the highest levels in brain, skeletal muscle, and testis. It was further established that there are three patterns of alternative splicing producing in-frame deletions in the coding sequence of dymple in a tissue-specific manner. When overexpressed, wild-type dymple exhibited a punctate perinuclear cytoplasmic pattern, whereas an amino-terminal deletion mutant formed large aggregates bounded by a trans-Golgi network marker. Since dynamin participates in clathrin-mediated endocytosis through a wellcharacterized mechanism, the existence of a dynaminlike molecule in each specific vesicle transport pathway has been predicted. Our findings suggest that dymple may be the first example of such a subfamily in mammalian cells other than dynamin itself, although its precise role and membrane localization remain to be resolved.

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Diesel fuel ignition delay times were characterized in a rapid compression machine (RCM) using cylinder ambient gas temperature and pressure measurements as diagnostics. The objective of the study was to investigate the dependency of ignition delay time on: (a) cylinder ambient gas temperature, (b) cylinder ambient gas pressure, (c) injection pressure, (d) injector nozzle orifice diameter, (e) base fuel cetane number and (f) 2-ethylhexyl nitrate (2-EHN) cetane improver additive. The results presented here show that diesel ignition delay times can be shortened by increasing cylinder gas ambient temperatures and pressures, injection pressures and base fuel cetane number, either through blend components or by addition of cetane improver. Decreasing the injector nozzle orifice diameter also decreases the ignition delay time. It was also found that ignition chemistry is rate controlled by the molar concentration of the cylinder gas oxygen.