Shiow-Her Chiou

ATPase family AAA domain-containing 3A is a novel anti-apoptotic factor in lung adenocarcinoma cells

AAA domain-containing 3A (ATAD3A) is a member of the AAA-ATPase family. Three forms of ATAD3 have been identified: ATAD3A, ATAD3B and ATAD3C. In this study, we examined the type and expression of ATAD3 in lung adenocarcinoma (LADC). Expression of ATAD3A was detected by reverse transcription-polymerase chain reaction, immunoblotting, immunohistochemistry and confocal immunofluorescent microscopy. Our results show that ATAD3A is the major form expressed in LADC. Silencing of ATAD3A expression increased mitochondrial fragmentation and cisplatin sensitivity. Serum deprivation increased ATAD3A expression and drug resistance. These results suggest that ATAD3A could be an anti-apoptotic marker in LADC.

HIV-1 Vpr Triggers Mitochondrial Destruction by Impairing Mfn2-Mediated ER-Mitochondria Interaction

Human immunodeficiency virus 1 (HIV-1) viral protein R (Vpr) has been shown to induce host cell death by increasing the permeability of mitochondrial outer membrane (MOM). The mechanism underlying the damage to the mitochondria by Vpr, however, is not clearly illustrated. In this study, Vpr that is introduced, via transient transfection or lentivirus infection, into the human embryonic kidney cell line HEK293, human CD4+ T lymphoblast cell line SupT1, or human primary CD4+ T cells serves as the model system to study the molecular mechanism of Vpr-mediated HIV-1 pathogenesis. The results show that Vpr injures MOM and causes a loss in membrane potential (MMP) by posttranscriptionally reducing the expression of mitofusin 2 (Mfn2) via VprBP-DDB1-CUL4A ubiquitin ligase complex, gradually weakening MOM, and increasing mitochondrial deformation. Vpr also markedly decreases cytoplasmic levels of dynamin-related protein 1 (DRP1) and increases bulging in mitochondria-associated membranes (MAM), the specific regions of endoplasmic reticulum (ER) which form physical contacts with the mitochondria. Overexpression of Mfn2 and DRP1 significantly decreased the loss of MMP and apoptotic cell death caused by Vpr. Furthermore, by employing time-lapse confocal fluorescence microscopy, we identify the transport of Vpr protein from the ER, via MAM to the mitochondria. Taken together, our results suggest that Vpr-mediated cellular damage may occur on an alternative protein transport pathway from the ER, via MAM to the mitochondria, which are modulated by Mfn2 and DRP1.

Detection of severe acute respiratory syndrome-associated coronavirus in pneumocytes of the lung.

Abstract Previous reports have indicated that patients with severe acute respiratory syndrome (SARS)–associated coronavirus infection could develop atypical pneumonia with fulminant pulmonary edema. However, the target cells of SARS viral infection have not been characterized in detail. We report the pathologic findings of the lung in 3 cases of SARS. Chest radiographs at 2 to 3 weeks of infection revealed an atypical pneumonia with pulmonary consolidation, a clinical characteristic of SARS infection. The presence of the SARS virus was determined by nested reverse transcription–polymerase chain reaction (RT-PCR), and the infected cells were identified by in situ hybridization in open-lung biopsy and postmortem necropsy specimens. Expression of SARS virus–encoded RNA was detected in all 3 cases by RT-PCR, and the SARS viral signal was localized in pneumocytes by using in situ hybridization.

Sumoylation of eukaryotic elongation factor 2 is vital for protein stability and anti-apoptotic activity in lung adenocarcinoma cells

By screening mouse monoclonal antibody libraries for Kelch repeats, we serendipitously identified monoclonal antibodies to eukaryotic elongation factor 2 (eEF2). Interestingly, eEF2 was highly expressed in lung adenocarcinoma (LADC), but not in the neighboring non‐tumor lung tissue. Normally, eEF2 is involved in the peptidyl‐tRNA translocation during protein synthesis. Overexpression of eEF2 would implicate an association with disease progression of LADC. In the present study, we investigated the prognostic significance of eEF2 in patients with LADC. Expression of eEF2 was detected by immunoblotting, immunohistochemistry and confocal immunofluorescence microscopy. Our results show that patients with high eEF2 expression had a significantly higher incidence of early tumor recurrence (67.8%vs 18.2%, P = 0.016), and a significantly worse prognosis (P < 0.001). In an in vitro study, silencing of eEF2 expression increased mitochondrial elongation, cellular autophagy and cisplatin sensitivity. Moreover, eEF2 was sumoylated in LADC cells, and eEF2 sumoylation correlated with drug resistance. These results suggest that eEF2 is an anti‐apoptotic marker in LADC. However, biological function and involvement of eEF2 in the disease progression of LADC require further studies. (Cancer Sci 2011; 102: 1582–1589)

An alternative import pathway of AIF to the mitochondria

In eukaryotic cells, transport of the newly synthesized proteins and phospholipids to the appropriate subcellular target compartments is essential for maintaining organelle morphology and cell survival. In animal cells, mitochondria are major organelles containing DNA genome that encodes only for a small fraction of their proteins, which are required for the organelle function. Most mitochondrial proteins are encoded by the nuclear genes and imported to the mitochondria following protein synthesis. Apoptosis-inducing factor (AIF), an essential FAD-dependent NADH oxidase for the oxidative phosphorylation, is located in the intermembranous space and contains mitochondrial localization signals. However, the import mechanism of AIF to the mitochondria is not yet studied. Using sucrose gradient ultracentrifugation and immunoblotting, AIF was detected in fractions of the endoplasmic reticulum, mitochondria-associated membranes (MAM) and mitochondria, and AIF from these fractions was resistant to trypsin in the absence of digitonin, suggesting that AIF could be protected by phospholipids. Knockdown of dynamin-related protein 1 (DRP1kd) expression reduced AIF levels in the mitochondria, but increased AIF concentrations in the MAM. Knockdown of mitofusin-2 (Mfn-2kd) or ATPase family AAA domain containing 3A (ATAD3Akd) expression, however, reduced AIF levels in the mitochondria and increased the number of transport vesicles that contained AIF in the cytosol, indicating that ATAD3A and Mfn-2 were respectively essential for the import and fusion of transport vesicles into the mitochondria. Here we show that AIF is imported from the endoplasmic reticulum to the mitochondria via mitochondria-associated membranes and transport vesicles.

Overexpression of aldo-keto reductase 1C2 is associated with disease progression in patients with prostatic cancer

Huang K‐H, Chiou S‐H, Chow K‐C, Lin T‐Y, Chang H‐W, Chiang I‐P & Lee M‐C
(2010) Histopathology 57, 384–394
Overexpression of aldo‐keto reductase 1C2 is associated with disease progression in patients with prostatic cancer

Mitochondrial protein ATPase family, AAA domain containing 3A correlates with radioresistance in glioblastoma

BACKGROUND ATPase-family, AAA domain containing 3A (ATAD3A) is located on human chromosome 1p36.33, and high endogenous expression may associate with radio- and chemosensitivity. This study was conducted to investigate the significance of ATAD3A in glioblastoma multiforme (GBM). METHODS Clinical significance of ATAD3A expression was assessed by immunohistochemistry in 67 GBM specimens, and prognostic value was assessed in 32 GBM patients statistically. To investigate in vitro phenotypic effects of ATAD3A, cell viability was measured using a clonogenic survival assay under either knockdown or ectopic expression of ATAD3A in GBM cell lines. The effects of ATAD3A knockdown on targeted DNA repair-associated proteins in T98G cells were evaluated using immunofluorescence and Western blotting. RESULTS Clinically, high expression of ATAD3A was independent of O(6)-DNA methylguanine-methyltransferase methylation status and correlated with worse prognosis. In vitro, high ATAD3A-expressing T98G cells were more resistant to radiation-induced cell death compared with control and low endogenous ATAD3A U87MG cells. After silencing ATAD3A, T98G cells became more sensitive to radiation. On the other hand, enforced ATAD3A expression in U87MG cells exhibited increased radioresistance. ATAD3A may coordinate with aldo-keto reductase genes and participate in bioactivation or detoxication of temozolomide. Surprisingly, deficient DNA repair after irradiation was observed in T98G/ATAD3A knockdown as a result of decreased nuclear ataxia telangiectasia mutated kinase and histones H2AX and H3, which was also evidenced by the sustained elevation of poly (ADP-ribose) polymerase prior to and after radiation treatment. CONCLUSION Our data suggest that high expression of ATAD3A is an independent biomarker for radioresistance in GBM. ATAD3A could be a potential target for therapy.

Overexpression of optic atrophy 1 protein increases cisplatin resistance via inactivation of caspase-dependent apoptosis in lung adenocarcinoma cells

Optic atrophy 1 protein, a 112-kd guanosine triphosphatase, is involved in the mitochondrial inner membrane fusion and anticancer drug-mediated cytotoxicity, which implicate an association with disease progression of the cancer. In this study, we investigated the prognostic value of optic atrophy 1 expression in patients with lung adenocarcinoma. Using immunohistochemical staining, expression of optic atrophy 1 was determined in 286 lung adenocarcinoma patients. Expression of optic atrophy 1 was confirmed by immunoblotting. The relationship between optic atrophy 1 expression and clinicopathological parameters was analyzed statistically by comparing survival between different groups using the log-rank test. The results showed that optic atrophy 1 overexpression was detected in 219 (76.6%) of lung adenocarcinoma patients. A significant difference was found in cumulative survival between patients with high optic atrophy 1 levels and those with low optic atrophy 1 levels (P = .0016). In the in vitro experiments with cell lines, silencing of optic atrophy 1 expression reduced cisplatin resistance, which was further shown via increased release of cytochrome c and activation of caspase-dependent apoptotic pathway. In conclusion, optic atrophy 1 is highly expressed in lung adenocarcinoma and indicates poor prognosis.

Characterization of interleukin-1β mRNA expression in chicken macrophages in response to avian reovirus

Inhibitors of viral disassembly or RNA and protein synthesis, viral disassembly intermediates (infectious subviral particles, ISVP), binary ethylenimine-inactivated virions, and viral particles lacking genomic double-stranded (ds) RNA (empty particles) were used to assess the expression of interleukin-1beta (IL-1beta) mRNA in chicken (chIL-1beta) macrophages in response to avian reovirus. The results demonstrate that two distinct expression patterns of chIL-1beta mRNA mediated by different steps in viral replication were found. Viral disassembly was required for the induction of a rapid, transient expression pattern of chIL-1beta mRNA that was rapidly induced at 30 min, with maximal levels reached by 2 h, and fell to a low level within 6 h post-inoculation, while viral RNA synthesis rather than protein translation, which was subsequent to membrane penetration, was required to induce a stable, sustained expression pattern of chIL-1beta mRNA that occurred at and after 6 h post-inoculation. In addition, the induction of chIL-1beta mRNA expression by the empty particles and ISVP was extremely weak, compared with the active dsRNA(+) virions or binary ethylenimine-inactivated virions, suggesting that the presence of dsRNA, even if transcriptionally inactive, may be an important factor in this response.

Pet dogs owned by lupus patients are at a higher risk of developing lupus

The aim of this study is to determine whether pet dogs owned by patients with systemic lupus erythematosus (SLE) are at a higher risk of developing SLE. Diagnosis of canine SLE was mainly based on the 11 diagnostic criteria for human SLE and two marked immunological features of canine SLE. Among 59 pet dogs owned by 37 SLE patients, 11 (18.64%) were ANA positive, and three (5.08%) had SLE. In contrast, of 187 pet dogs owned by non-SLE households, nine (4.81%) were ANA positive, and none (0%) had SLE. Among 650 outpatient dogs registered in the veterinary hospital, 34 (5.23%) were ANA positive, and six (0.92%) had SLE. Frequency of ANA and SLE among pet dogs owned by SLE patients was significantly higher than in pet dogs owned by non-SLE households (P 1/4 0.001 for ANA; P 1/4 0.013 for SLE) and in outpatient dogs (P, 0.001 for ANA; P 1/4 0.032 for SLE). With respect to canine SLE development, the relative risk or risk ratio (R)of human SLE contact varied from 5.5 (compared with outpatient dogs) to near the infinite (compared with dogs owned by non-SLE households). The prevalence of canine SLE among pet dogs of SLE patients was therefore estimated to be 508 per 10 000 [95% confidence interval (95% CI), 0-1068]. In conclusion, pet dogs with human SLE contact were at a higher risk of developing SLE. Our results indicate that a common environmental factor or zoonotic agent may be involved in the development of human and canine SLE.