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1.  The dynamic nature and territory of transcriptional machinery in the bacterial chromosome 
Our knowledge of the regulation of genes involved in bacterial growth and stress responses is extensive; however, we have only recently begun to understand how environmental cues influence the dynamic, three-dimensional distribution of RNA polymerase (RNAP) in Escherichia coli on the level of single cell, using wide-field fluorescence microscopy and state-of-the-art imaging techniques. Live-cell imaging using either an agarose-embedding procedure or a microfluidic system further underscores the dynamic nature of the distribution of RNAP in response to changes in the environment and highlights the challenges in the study. A general agreement between live-cell and fixed-cell images has validated the formaldehyde-fixing procedure, which is a technical breakthrough in the study of the cell biology of RNAP. In this review we use a systems biology perspective to summarize the advances in the cell biology of RNAP in E. coli, including the discoveries of the bacterial nucleolus, the spatial compartmentalization of the transcription machinery at the periphery of the nucleoid, and the segregation of the chromosome territories for the two major cellular functions of transcription and replication in fast-growing cells. Our understanding of the coupling of transcription and bacterial chromosome (or nucleoid) structure is also summarized. Using E. coli as a simple model system, co-imaging of RNAP with DNA and other factors during growth and stress responses will continue to be a useful tool for studying bacterial growth and adaptation in changing environment.
doi:10.3389/fmicb.2015.00497
PMCID: PMC4440401  PMID: 26052320
RNA polymerase; bacterial nucleolus; replisome; chromosome territories; growth rate regulation; stress responses; superresolution imaging; E. coli
2.  Dendritic cell CD83 homotypic interactions regulate inflammation and promote mucosal homeostasis 
Mucosal Immunology  2014;8(2):414-428.
Dendritic cells (DCs) form an extensive network in the intestinal lamina propria, which orchestrates the mucosal immune response. Alterations in DC function can predispose to inflammatory bowel disease, although by unknown mechanisms. We show that CD83, a highly regulated DC cell surface protein, modulates the immune response to prevent colitis. Mice with a conditional knockout of CD83 in DCs develop exacerbated colitis following dextran sodium sulfate challenge, whereas mucosal overexpression of CD83 inhibits DC inflammatory response and protects against colitis. These CD83 perturbations can be modeled in vitro where we show that CD83 homotypic interaction occurs via cell–cell contact and inhibits pro-inflammatory responses. CD83 knockdown or cytoplasmic truncation abrogates the effects of homotypic binding. We demonstrate that CD83 homotypic interaction regulates DC activation via the mitogen-activated protein kinase pathway by inhibiting p38α phosphorylation. Our findings indicate that CD83 homotypic interactions regulate DC activation and promote mucosal homeostasis.
doi:10.1038/mi.2014.79
PMCID: PMC4326976  PMID: 25204675
3.  Ubiquitination by SAG regulates macrophage survival/death and immune response during infection 
Cell Death and Differentiation  2014;21(9):1388-1398.
The checkpoint between the life and death of macrophages is crucial for the host's frontline immune defense during acute phase infection. However, the mechanism as to how the immune cell equilibrates between apoptosis and immune response is unclear. Using in vitro and ex vivo approaches, we showed that macrophage survival is synchronized by SAG (sensitive to apoptosis gene), which is a key member of the ubiquitin–proteasome system (UPS). When challenged by pathogen-associated molecular patterns (PAMPs), we observed a reciprocal expression profile of pro- and antiapoptotic factors in macrophages. However, SAG knockdown disrupted this balance. Further analysis revealed that ubiquitination of Bax and SARM (sterile α- and HEAT/armadillo-motif-containing protein) by SAG-UPS confers survival advantage to infected macrophages. SAG knockdown caused the accumulation of proapoptotic Bax and SARM, imbalance of Bcl-2/Bax in the mitochondria, induction of cytosolic cytochrome c and activation of caspase-9 and -3, all of which led to disequilibrium between life and death of macrophages. In contrast, SAG-overexpressing macrophages challenged with PAMPs exhibited upregulation of protumorigenic cytokines (IL-1β, IL-6 and TNF-α), and downregulation of antitumorigenic cytokine (IL-12p40) and anti-inflammatory cytokine (IL-10). This suggests that SAG-dependent UPS is a key switch between immune defense and apoptosis or immune overactivation and tumorigenesis. Altogether, our results indicate that SAG-UPS facilitates a timely and appropriate level of immune response, prompting future development of potential immunomodulators of SAG-UPS.
doi:10.1038/cdd.2014.54
PMCID: PMC4131172  PMID: 24786833
4.  LSD1-mediated epigenetic modification contributes to proliferation and metastasis of colon cancer 
British Journal of Cancer  2013;109(4):994-1003.
Background:
Emerging evidence has demonstrated that lysine-specific demethylase 1 (LSD1) has an important role in many pathological processes of cancer cells, such as carcinogenesis, proliferation and metastasis. In this study, we characterised the role and molecular mechanisms of LSD1 in proliferation and metastasis of colon cancer.
Methods:
We evaluated the correlation of LSD1, CDH-1 and CDH-2 with invasiveness of colon cancer cells, and investigated the roles of LSD1 in proliferation, invasion and apoptosis of colon cancer cells. We further investigated the mechanisms of LSD1-mediated metastasis of colon cancer.
Results:
Lysine-specific demethylase 1 was upregulated in colon cancer tissues, and the high LSD1 expression was significantly associated with tumour-node-metastasis (TNM) stages and distant metastasis. Functionally, inhibition of LSD1 impaired proliferation and invasiveness, and induced apoptosis of colon cancer cells in vitro. The LSD1 physically interacted with the promoter of CDH-1 and decreased dimethyl histone H3 lysine 4 (H3K4) at this region, downregulated CDH-1 expression, and consequently contributed to colon cancer metastasis.
Conclusion:
Lysine-specific demethylase 1 downregulates the expression of CDH-1 by epigenetic modification, and consequently promotes metastasis of colon cancer cells. The LSD1 antagonists might be a useful strategy to suppress metastasis of colon cancer.
doi:10.1038/bjc.2013.364
PMCID: PMC3749561  PMID: 23900215
lysine-specific demethylase 1; CDH-1; CDH-2; colon cancer; metastasis; histone modifications
5.  CLIC4 regulates TGF-β dependent myofibroblast differentiation to produce a cancer stroma 
Oncogene  2013;33(7):842-850.
Cancer stroma has a profound influence on tumor development and progression. The conversion of fibroblasts to activated myofibroblasts is a hallmark of reactive tumor stroma. Among a number of factors involved in this conversion, TGF-β has emerged as a major regulator. CLIC4, an integral protein in TGF-β signaling, is highly upregulated in stroma of multiple human cancers, and overexpression of CLIC4 in stromal cells enhances the growth of cancer xenografts. In this study we show that conditioned media from tumor cell lines induces expression of both CLIC4 and the myofibroblast marker alpha smooth muscle actin (α-SMA) in stromal fibroblasts via TGF-β signaling. Genetic ablation of CLIC4 in primary fibroblasts prevents or reduces constitutive or TGF-β induced expression of α-SMA and extracellular matrix components that are markers of myofibroblasts. CLIC4 is required for the activation of p38 Map Kinase by TGF-β, a pathway that signals myofibroblast conversion in stromal cells. This requirement involves the interaction of CLIC4 with PPM1a, the selective phosphatase of activated p-38. Conditioned media from fibroblasts overexpressing CLIC4 increases tumor cell migration and invasion in a TGF-β dependent manner and promotes epithelial to mesenchymal transition indicating that high stromal CLIC4 serves to enhance tumor invasiveness and progression. Thus, CLIC4 is significantly involved in the development of a nurturing tumor microenvironment by enhancing TGF-β signaling in a positive feedback loop. Targeting CLIC4 in tumor stroma should be considered as a strategy to mitigate some of the tumor enhancing effects of the cancer stroma.
doi:10.1038/onc.2013.18
PMCID: PMC3912213  PMID: 23416981
CLIC; α-SMA; p38; microenvironment; PPM1a
6.  THE IMPACT OF SARCOPENIA ON A PHYSICAL ACTIVITY INTERVENTION: THE LIFESTYLE INTERVENTIONS AND INDEPENDENCE FOR ELDERS PILOT STUDY (LIFE-P) 
Objective
To determine if sarcopenia modulates the response to a physical activity intervention in functionally limited older adults.
Design
Secondary analysis of a randomized controlled trial.
Setting
Three academic centers.
Participants
Elders aged 70 to 89 years at risk for mobility disability who underwent dual-energy x-ray absorptiometry (DXA) for body composition at enrollment and follow-up at twelve months (N = 177).
Intervention
Subjects participated in a physical activity program (PA) featuring aerobic, strength, balance, and flexibility training, or a successful aging (SA) educational program about healthy aging.
Measurements
Sarcopenia as determined by measuring appendicular lean mass and adjusting for height and total body fat mass (residuals method), Short Physical Performance Battery score (SPPB), and gait speed determined on 400 meter course.
Results
At twelve months, sarcopenic and non-sarcopenic subjects in PA tended to have higher mean SPPB scores (8.7±0.5 and 8.7±0.2 points) compared to sarcopenic and non-sarcopenic subjects in SA (8.3±0.5 and 8.4±0.2 points, p = 0.24 and 0.10), although the differences were not statistically significant. At twelve months, faster mean gait speeds were observed in PA: 0.93±0.4 and 0.95±0.03 meters/second in sarcopenic and non-sarcopenic PA subjects, and 0.89±0.4 and 0.91±0.03 meters/second in sarcopenic and non-sarcopenic SA subjects (p = 0.98 and 0.26), although not statistically significant. There was no difference between the sarcopenic and non-sarcopenic groups in intervention adherence or number of adverse events.
Conclusion
These data suggest that older adults with sarcopenia, who represent a vulnerable segment of the elder population, are capable of improvements in physical performance after a physical activity intervention.
doi:10.1007/s12603-013-0369-0
PMCID: PMC4111145  PMID: 24402391
Sarcopenia; physical activity; gait speed; short physical performance battery
7.  Pericardial adipose tissue and coronary artery calcification in The Multi-Ethnic Study of Atherosclerosis (MESA) 
Obesity (Silver Spring, Md.)  2013;21(5):1056-1063.
We examined the relationship of pericardial adipose tissue (PAT) with coronary artery calcification in MESA, a large cohort in which associations by race/ethnicity can be compared. The baseline cohort comprised 6,814 Caucasian (38%), African American (28%), Chinese American (12%) and Hispanic (22%) adults aged 45–84, without known clinical cardiovascular disease. Cardiac CT was used to measure PAT (cm3) and calcification (Agatston score). We examined cross-sectional associations of PAT with the presence (score>0) and severity (continuous score if >0) of calcification using prevalence ratio (PR) (n=6,672) and linear regression (n=3,362), respectively. Main models were adjusted for age, age2, gender, race/ethnicity, field site, smoking, physical activity, alcohol and education. PAT volume (adjusted for age, height, weight and site) was greatest in Chinese males, while Black males had less PAT than all but Black females. PAT was associated with presence [PR per standard deviation (SD): 1.06 (95% CI: 1.04, 1.08)] and severity [difference in log Agatston score per SD: 0.15 (0.09, 0.21)] of calcification, but neither association varied by race/ethnicity. Adjustment for generalized adiposity attenuated but did not eliminate the associations. With further adjustment for traditional risk factors and inflammatory markers, only the association with severity remained statistically significant [PR: 1.02 (1.00, 1.04), difference: 0.10 (0.03, 0.17)]. Heterogeneity by sex was observed for presence of calcification (PR in men: 1.04; in women: 1.08; p for interaction<0.0001). Pericardial adipose tissue was associated with the presence and severity of coronary artery calcification in this cohort, but despite differences in PAT volumes and calcification across race/ethnic groups, neither association varied by race/ethnicity.
doi:10.1002/oby.20090
PMCID: PMC4042681  PMID: 23784910
Coronary artery calcification; pericardial fat; subclinical atherosclerosis risk factors; obesity; epidemiology
8.  SYMPTOMS OF DEPRESSION BUT NOT ANXIETY ARE ASSOCIATED WITH CENTRAL OBESITY AND CARDIOVASCULAR DISEASE IN PEOPLE WITH TYPE 2 DIABETES: THE EDINBURGH TYPE 2 DIABETES STUDY 
Diabetologia  2009;53(3):467-471.
Aims/hypothesis
To identify risk factors for depression and anxiety in a well-characterised cohort of subjects with type 2 diabetes mellitus.
Methods
We used baseline data from participants (n=1066, 48.7% women, aged 67.9 ± 4.2 years) from the Edinburgh Type 2 Diabetes Study. Symptoms of anxiety and depression were assessed using the Hospital Anxiety and Depression Scale (HADS). Obesity was characterised according to both overall (body mass index, fat mass) and abdominal (waist circumference) measurement. Cardiovascular disease was assessed by questionnaire, physical examination and review of medical records. Stepwise multiple linear regression was performed to identify explanatory variables related to either anxiety or depression HADS scores.
Results
Abdominal obesity (waist circumference) and cardiovascular disease (ischaemic heart disease and ankle-brachial pressure index) were related to depression but not anxiety. Lifetime history of severe hypoglycaemia was associated with anxiety. Other cardiovascular risk factors or microvascular complications were not related to either anxiety or depressive symptoms.
Conclusion/interpretation
Depression but not anxiety is associated with abdominal obesity and cardiovascular disease in people with type 2 diabetes mellitus. This knowledge may help to identify depressive symptoms among patients with type 2 diabetes who are at greatest risk.
doi:10.1007/s00125-009-1628-9
PMCID: PMC3977034  PMID: 20012009
depression; anxiety; obesity; diabetes; cortisol
9.  Bacteriophage λ N protein inhibits transcription slippage by Escherichia coli RNA polymerase 
Nucleic Acids Research  2014;42(9):5823-5829.
Transcriptional slippage is a class of error in which ribonucleic acid (RNA) polymerase incorporates nucleotides out of register, with respect to the deoxyribonucleic acid (DNA) template. This phenomenon is involved in gene regulation mechanisms and in the development of diverse diseases. The bacteriophage λ N protein reduces transcriptional slippage within actively growing cells and in vitro. N appears to stabilize the RNA/DNA hybrid, particularly at the 5′ end, preventing loss of register between transcript and template. This report provides the first evidence of a protein that directly influences transcriptional slippage, and provides a clue about the molecular mechanism of transcription termination and N-mediated antitermination.
doi:10.1093/nar/gku203
PMCID: PMC4027172  PMID: 24711367
10.  Tanshinone-1 induces tumor cell killing, enhanced by inhibition of secondary activation of signaling networks 
Cell Death & Disease  2013;4(11):e905-.
Tumor multidrug resistance (MDR) can result from overexpression of drug transporters and deregulation of cellular signaling transduction. New agents and strategies are required for overcoming MDR. Here, we report that tanshinone-1, a bioactive ingredient in traditional Chinese medicine, directly killed MDR tumor cells and their corresponding parental cells, which was potentiated by inhibition of secondary activation of signaling networks. Tanshinone-1 was slightly more potent at inducing cytotoxicity and apoptosis in MDR cells than in corresponding parental cells. Tanshinone-1-induced MDR cell killing was independent of the function and expression of drug transporters but was partially correlated with the phosphatase-dependent reduction of phospho-705-Stat3, which secondarily activated p38-, AKT-, and ERK-involved signaling networks. Cotreatments with p38, AKT, and ERK inhibitors potentiated the anti-MDR effects of tanshinone-1. Our study presents a model for MDR cell killing using a compound of natural origin. This model could lead to new therapeutic strategies for targeting signaling network(s) in MDR cancers as well as new strategies for multitarget design.
doi:10.1038/cddis.2013.443
PMCID: PMC3847321  PMID: 24201804
tanshinone-1; multidrug resistance; Stat3; p38; AKT; ERK
11.  A Cross-Talk Between NFAT and NF-κB Pathways is Crucial for Nickel-Induced COX-2 Expression in Beas-2B Cells 
Current cancer drug targets  2011;11(5):548-559.
Cyclooxygenase-2 (COX-2) is a critical enzyme implicated in chronic inflammation-associated cancer development. Our studies have shown that the exposure of Beas-2B cells, a human bronchial epithelial cell line, to lung carcinogenic nickel compounds results in increased COX-2 expression. However, the signaling pathways leading to nickel-induced COX-2 expression are not well understood. In the current study, we found that the exposure of Beas-2B cells to nickel compounds resulted in the activation of both nuclear factor of activated T cell (NFAT) and nuclear factor-κB (NF-κB). The expression of COX-2 induced upon nickel exposure was inhibited by either a NFAT pharmacological inhibitor or the knockdown of NFAT3 by specific siRNA. We further found that the activation of NFAT and NF-κB was dependent on each other. Since our previous studies have shown that NF-κB activation is critical for nickel-induced COX-2 expression in Beas-2B cells exposed to nickel compounds under same experimental condition, we anticipate that there might be a cross-talk between the activation of NFAT and NF-κB for the COX-2 induction due to nickel exposure in Beas-2B cells. Furthermore, we showed that the scavenging of reactive oxygen species (ROS) by introduction of mitochondrial catalase inhibited the activation of both NFAT and NF-κB, and the induction of COX-2 due to nickel exposure. Taken together, our results defining the evidence showing a key role of the cross-talk between NFAT and NF-κB pathways in regulating nickel-induced COX-2 expression, further provide insight into the understanding of the molecular mechanisms linking nickel exposure to its lung carcinogenic effects.
PMCID: PMC3759234  PMID: 21486220
Beas-2B cells; COX-2; nickel; NFAT; NF-κB; ROS
12.  Efficient use of longitudinal CD4 counts and viral load measures in survival analysis 
Statistics in medicine  2012;31(19):2086-2097.
CD4 counts and viral loads are dynamic quantities that change with time in HIV-infected persons. Commonly used single summary measures, such as viral load set point or early CD4 count do not explicitly account for changes in viral load or CD4 counts or other features of the overall time course of these measures. However, the efficient use of all repeated measurements within each subject is often a challenge made more difficult by sparse and irregular sampling over time. Here we illustrate how functional principal component (FPC) analysis provides an effective statistical approach for exploiting the patterns in CD4 count and viral load data over time. The method is demonstrated using data from Kenyan women who acquired HIV-1 during follow-up in a high risk cohort and were subsequently followed prospectively from early infection. The FPC scores for each woman obtained by this method serve as informative summary statistics for the CD4-count and viral-load trajectories. Similar to baseline CD4 count or viral set point, the first FPC score can be interpreted as a single-value summary measure of an individual's overall CD4 count or viral load. However, unlike most single-value summaries of CD4-count or viral-load trajectories, the first FPC score summarizes the dynamics of these quantities and is seen to reveal specific features of the trajectories associated with mortality in this cohort. Moreover, FPC scores are shown to be a more powerful prognostic factor than other common summaries when used in survival analysis.
doi:10.1002/sim.5318
PMCID: PMC3641845  PMID: 22415871
longitudinal data; functional principal components; CD4 counts; viral loads
13.  Genome conformation capture reveals that the Escherichia coli chromosome is organized by replication and transcription 
Nucleic Acids Research  2013;41(12):6058-6071.
To fit within the confines of the cell, bacterial chromosomes are highly condensed into a structure called the nucleoid. Despite the high degree of compaction in the nucleoid, the genome remains accessible to essential biological processes, such as replication and transcription. Here, we present the first high-resolution chromosome conformation capture-based molecular analysis of the spatial organization of the Escherichia coli nucleoid during rapid growth in rich medium and following an induced amino acid starvation that promotes the stringent response. Our analyses identify the presence of origin and terminus domains in exponentially growing cells. Moreover, we observe an increased number of interactions within the origin domain and significant clustering of SeqA-binding sequences, suggesting a role for SeqA in clustering of newly replicated chromosomes. By contrast, ‘histone-like’ protein (i.e. Fis, IHF and H-NS) -binding sites did not cluster, and their role in global nucleoid organization does not manifest through the mediation of chromosomal contacts. Finally, genes that were downregulated after induction of the stringent response were spatially clustered, indicating that transcription in E. coli occurs at transcription foci.
doi:10.1093/nar/gkt325
PMCID: PMC3695519  PMID: 23632166
14.  Anti-Hepatitis C Virus Activity and Toxicity of Type III Phosphatidylinositol-4-Kinase Beta Inhibitors 
Antimicrobial Agents and Chemotherapy  2012;56(10):5149-5156.
Type III phosphatidylinositol-4-kinase beta (PI4KIIIβ) was previously implicated in hepatitis C virus (HCV) replication by small interfering RNA (siRNA) depletion and was therefore proposed as a novel cellular target for the treatment of hepatitis C. Medicinal chemistry efforts identified highly selective PI4KIIIβ inhibitors that potently inhibited the replication of genotype 1a and 1b HCV replicons and genotype 2a virus in vitro. Replicon cells required more than 5 weeks to reach low levels of 3- to 5-fold resistance, suggesting a high resistance barrier to these cellular targets. Extensive in vitro profiling of the compounds revealed a role of PI4KIIIβ in lymphocyte proliferation. Previously proposed functions of PI4KIIIβ in insulin secretion and the regulation of several ion channels were not perturbed with these inhibitors. Moreover, PI4KIIIβ inhibitors were not generally cytotoxic as demonstrated across hundreds of cell lines and primary cells. However, an unexpected antiproliferative effect in lymphocytes precluded their further development for the treatment of hepatitis C.
doi:10.1128/AAC.00946-12
PMCID: PMC3457382  PMID: 22825118
15.  T-cell death following immune activation is mediated by mitochondria-localized SARM 
Cell Death and Differentiation  2012;20(3):478-489.
Following acute-phase infection, activated T cells are terminated to achieve immune homeostasis, failure of which results in lymphoproliferative and autoimmune diseases. We report that sterile α- and heat armadillo-motif-containing protein (SARM), the most conserved Toll-like receptors adaptor, is proapoptotic during T-cell immune response. SARM expression is significantly reduced in natural killer (NK)/T lymphoma patients compared with healthy individuals, suggesting that decreased SARM supports NK/T-cell proliferation. T cells knocked down of SARM survived and proliferated more significantly compared with wild-type T cells following influenza infection in vivo. During activation of cytotoxic T cells, the SARM level fell before rising, correlating inversely with cell proliferation and subsequent T-cell clearance. SARM knockdown rescued T cells from both activation- and neglect-induced cell deaths. The mitochondria-localized SARM triggers intrinsic apoptosis by generating reactive oxygen species and depolarizing the mitochondrial potential. The proapoptotic function is attributable to the C-terminal sterile alpha motif and Toll/interleukin-1 receptor domains. Mechanistically, SARM mediates intrinsic apoptosis via B cell lymphoma-2 (Bcl-2) family members. SARM suppresses B cell lymphoma-extra large (Bcl-xL) and downregulates extracellular signal-regulated kinase phosphorylation, which are cell survival effectors. Overexpression of Bcl-xL and double knockout of Bcl-2 associated X protein and Bcl-2 homologous antagonist killer substantially reduced SARM-induced apoptosis. Collectively, we have shown how T-cell death following infection is mediated by SARM-induced intrinsic apoptosis, which is crucial for T-cell homeostasis.
doi:10.1038/cdd.2012.144
PMCID: PMC3569988  PMID: 23175186
intrinsic T-cell death by SARM; influenza infection; adoptive transfer mouse model; neglect- and activation-induced cell death; NK/T-cell lymphoma
16.  Anomalous exchange bias at collinear/noncollinear spin interface 
Scientific Reports  2013;3:1094.
We report on the interfacial magnetic coupling in manganite bilayers of collinear ferromagnetic La0.7Sr0.3MnO3 and noncollinear multiferroic TbMnO3. Exchange bias is observed at the Néel temperature of TbMnO3 (~41 K) due to the onset of long-range antiferromagnetic ordering in the Mn spin sublattice. Interestingly, an anomalous plateau of exchange bias emerges at the ordering temperature of Tb spins (~10 K), and we ascribe this unique feature to the strong coupling between Tb and Mn spin sublattices in TbMnO3, which in turn influences the magnetic coupling across the interface. On the other hand, the enhancement of coercivity in La0.7Sr0.3MnO3/TbMnO3 shows monotonous temperature dependence. Our results illustrate a strong interfacial magnetic interaction at the La0.7Sr0.3MnO3/TbMnO3 interface, highlighting the roles of competing spin orders, magnetic frustration, and coupling between multiple spin sublattices in artificial collinear/noncollinear spin heterostructures.
doi:10.1038/srep01094
PMCID: PMC3549540
17.  Free Sialic Acid Storage Disease without sialuria 
Annals of neurology  2009;65(6):753-757.
We performed high resolution in vitro proton nuclear magnetic resonance spectroscopy on CSF and urine samples of 44 patients with leukodystrophies of unknown cause. Free sialic acid was elevated in CSF of two siblings with mental retardation and mild hypomyelination. By contrast, urinary excretion of free sialic acid in urine was normal on repeated testing by two independent methods. Both patients were homozygous for the K136E mutation in SLC17A5, the gene responsible for the free sialic acid storage diseases. Our findings demonstrate that mutations in the SLC17A5 gene have to be considered in patients with hypomyelination, even in the absence of sialuria.
doi:10.1002/ana.21624
PMCID: PMC3508714  PMID: 19557856
Free Sialic Acid Storage Diseases; Leukodystrophy; NMR Spectroscopy
18.  Investigation of the non-volatile resistance change in noncentrosymmetric compounds 
Scientific Reports  2012;2:587.
Coexistence of polarization and resistance-switching characteristics in single compounds has been long inspired scientific and technological interests. Here, we report the non-volatile resistance change in noncentrosymmetric compounds investigated by using defect nanotechnology and contact engineering. Using a noncentrosymmetric material of ZnO as example, we first transformed ZnO into high resistance state. Then ZnO electrical polarization was probed and its domains polarized 180° along the [001]-axis with long-lasting memory effect (>25 hours). Based on our experimental observations, we have developed a vacancy-mediated pseudoferroelectricity model. Our first-principle calculations propose that vacancy defects initiate a spontaneous inverted domains nucleation at grain boundaries, and then they grow in the presence of an electrical field. The propagation of inverted domains follows the scanning tip motion under applied electrical field, leading to the growth of polarized domains over large areas.
doi:10.1038/srep00587
PMCID: PMC3421435  PMID: 22905318
19.  Identification of Toyocamycin, an agent cytotoxic for multiple myeloma cells, as a potent inhibitor of ER stress-induced XBP1 mRNA splicing 
Blood Cancer Journal  2012;2(7):e79-.
The IRE1α-XBP1 pathway, a key component of the endoplasmic reticulum (ER) stress response, is considered to be a critical regulator for survival of multiple myeloma (MM) cells. Therefore, the availability of small-molecule inhibitors targeting this pathway would offer a new chemotherapeutic strategy for MM. Here, we screened small-molecule inhibitors of ER stress-induced XBP1 activation, and identified toyocamycin from a culture broth of an Actinomycete strain. Toyocamycin was shown to suppress thapsigargin-, tunicamycin- and 2-deoxyglucose-induced XBP1 mRNA splicing in HeLa cells without affecting activating transcription factor 6 (ATF6) and PKR-like ER kinase (PERK) activation. Furthermore, although toyocamycin was unable to inhibit IRE1α phosphorylation, it prevented IRE1α-induced XBP1 mRNA cleavage in vitro. Thus, toyocamycin is an inhibitor of IRE1α-induced XBP1 mRNA cleavage. Toyocamycin inhibited not only ER stress-induced but also constitutive activation of XBP1 expression in MM lines as well as primary samples from patients. It showed synergistic effects with bortezomib, and induced apoptosis of MM cells including bortezomib-resistant cells at nanomolar levels in a dose-dependent manner. It also inhibited growth of xenografts in an in vivo model of human MM. Taken together, our results suggest toyocamycin as a lead compound for developing anti-MM therapy and XBP1 as an appropriate molecular target for anti-MM therapy.
doi:10.1038/bcj.2012.26
PMCID: PMC3408640  PMID: 22852048
multiple myeloma; ER stress; IRE1α; XBP1; toyocamycin; adenosine analog
20.  Dietary Fat and Cholesterol and Risk of Cardiovascular Disease in Older Adults: the Health ABC Study 
BACKGROUND AND AIMS
Although dietary fats and cholesterol have previously been associated with risk of cardiovascular disease (CVD) in middle aged populations, less is known among older adults. The purpose of this study was to determine the association between dietary fats, cholesterol, and eggs and CVD risk among community-dwelling adults aged 70–79 in the Health, Aging and Body Composition Study.
METHODS AND RESULTS
Diet was assessed using an interviewer-administered 108-item food frequency questionnaire (n=1,941). CVD events were defined as a confirmed myocardial infarction, coronary death, or stroke. Relative rates of CVD over 9 years of follow-up were estimated using Cox proportional hazards models. During follow-up, there were 203 incident cases of CVD. There were no significant associations between dietary fats and CVD risk. Dietary cholesterol (HR (95% CI): 1.47 (0.93, 2.32) for the upper vs. lower tertile; P for trend, 0.10) and egg consumption (HR (95% CI): 1.68 (1.12, 2.51) for 3+/week vs. <1/week); P for trend, 0.01) were associated with increased CVD risk. However, in subgroup analyses, dietary cholesterol and egg consumption were associated with increased CVD risk only among older adults with type 2 diabetes (HR (95% CI): 3.66 (1.09, 12.29) and 5.02 (1.63, 15.52), respectively, for the upper vs. lower tertile/group).
CONCLUSIONS
Dietary cholesterol and egg consumption were associated with increased CVD risk among older, community-dwelling adults with type 2 diabetes. Further research on the biological mechanism(s) for the increased CVD risk with higher dietary cholesterol and frequent egg consumption among older adults with diabetes is warranted.
doi:10.1016/j.numecd.2009.11.007
PMCID: PMC2911502  PMID: 20338738
21.  Genetic Differentiation of Chinese Indigenous Meat Goats Ascertained Using Microsatellite Information 
To investigate the genetic diversity of seven Chinese indigenous meat goat breeds (Tibet goat, Guizhou white goat, Shannan white goat, Yichang white goat, Matou goat, Changjiangsanjiaozhou white goat and Anhui white goat), explain their genetic relationship and assess their integrity and degree of admixture, 302 individuals from these breeds and 42 Boer goats introduced from Africa as reference samples were genotyped for 11 microsatellite markers. Results indicated that the genetic diversity of Chinese indigenous meat goats was rich. The mean heterozygosity and the mean allelic richness (AR) for the 8 goat breeds varied from 0.697 to 0.738 and 6.21 to 7.35, respectively. Structure analysis showed that Tibet goat breed was genetically distinct and was the first to separate and the other Chinese goats were then divided into two sub-clusters: Shannan white goat and Yichang white goat in one cluster; and Guizhou white goat, Matou goat, Changjiangsanjiaozhou white goat and Anhui white goat in the other cluster. This grouping pattern was further supported by clustering analysis and Principal component analysis. These results may provide a scientific basis for the characteristization, conservation and utilization of Chinese meat goats.
doi:10.5713/ajas.2011.11308
PMCID: PMC4093133  PMID: 25049548
Meat Goat; Genetic Diversity; Genetic Differentiation; Microsatellite
22.  Genomic biomarkers and cellular pathways of ischemic stroke by RNA gene expression profiling 
Neurology  2010;75(11):1009-1014.
Objective:
The objective of this study was to provide insight into the molecular mechanisms of acute ischemic cerebrovascular syndrome (AICS) through gene expression profiling and pathway analysis.
Methods:
Peripheral whole blood samples were collected from 39 MRI-diagnosed patients with AICS and 25 nonstroke control subjects ≥18 years of age. Total RNA was extracted from whole blood stabilized in Paxgene RNA tubes, amplified, and hybridized to Illumina HumanRef-8v2 bead chips. Gene expression was compared in a univariate manner between stroke patients and control subjects using t test in GeneSpring. The significant genes were tested in a logistic regression model controlling for age, hypertension, and dyslipidemia. Inflation of type 1 error was corrected by Bonferroni and Ingenuity Systems Pathway analysis was performed. Validation was performed by QRT-PCR using Taqman gene expression assays.
Results:
A 9-gene profile was identified in the whole blood of ischemic stroke patients using gene expression profiling. Five of these 9 genes were identified in a previously published expression profiling study of stroke and are therefore likely biomarkers of stroke. Pathway analysis revealed toll-like receptor signaling as a highly significant canonical pathway present in the peripheral whole blood of patients with AICS.
Conclusions:
Our study highlights the relevance of the innate immune system through toll-like receptor signaling as a mediator of response to ischemic stroke and supports the claim that gene expression profiling can be used to identify biomarkers of ischemic stroke. Further studies are needed to validate and refine these biomarkers for their diagnostic potential.
GLOSSARY
= acute ischemic cerebrovascular syndrome;
= blood–brain barrier;
= Ingenuity Systems Pathway analysis;
= peripheral blood mononuclear cell;
= recombinant tissue plasminogen activator;
= toll-like receptor.
doi:10.1212/WNL.0b013e3181f2b37f
PMCID: PMC2942033  PMID: 20837969
23.  Evidence for altered Wnt signaling in psoriatic skin 
The Wnt gene family encodes a set of highly conserved secreted signaling proteins that have major roles in embryogenesis and tissue homeostasis. Yet the expression of this family of important mediators in psoriasis, a disease characterized by marked changes in keratinocyte growth and differentiation, is incompletely understood. We subjected 58 paired biopsies from lesional and uninvolved psoriatic skin and 64 biopsies from normal skin to global gene expression profiling. WNT5A transcripts were up-regulated 5-fold in lesional skin, accompanied by increased Wnt-5a protein levels. Notably, WNT5A mRNA was markedly induced by IL-1α, TNF-α, IFN-γ and TGF-α in cultured keratinocytes. FZD2 and FZD5, which encode receptors for Wnt5A, were also increased in lesional psoriatic skin. In contrast, expression of WIF1 mRNA, encoding a secreted antagonist of the Wnt proteins, was down-regulated >10-fold in lesional skin, along with decreased WIF-1 immunostaining. Interestingly, pathway analysis along with reduced AXIN2 expression and lack of nuclear translocation of beta-catenin indicated a suppression of canonical Wnt signaling in lesional skin.
Our results suggest a shift away from canonical Wnt signaling towards non-canonical pathways driven by interactions between Wnt-5a and its cognate receptors in psoriasis, accompanied by impaired homeostatic inhibition of Wnt signaling by WIF-1 and Dkk.
doi:10.1038/jid.2010.67
PMCID: PMC2886156  PMID: 20376066
Wnt-signaling; psoriasis; WIF-1; Wnt proteins; keratinocytes
24.  Wogonin and related natural flavones are inhibitors of CDK9 that induce apoptosis in cancer cells by transcriptional suppression of Mcl-1 
Cell Death & Disease  2011;2(7):e182-.
The wogonin-containing herb Scutellaria baicalensis has successfully been used for curing various diseases in traditional Chinese medicine. Wogonin has been shown to induce apoptosis in different cancer cells and to suppress growth of human cancer xenografts in vivo. However, its direct targets remain unknown. In this study, we demonstrate for the first time that wogonin and structurally related natural flavones, for example, apigenin, chrysin and luteolin, are inhibitors of cyclin-dependent kinase 9 (CDK9) and block phosphorylation of the carboxy-terminal domain of RNA polymerase II at Ser2. This effect leads to reduced RNA synthesis and subsequently rapid downregulation of the short-lived anti-apoptotic protein myeloid cell leukemia 1 (Mcl-1) resulting in apoptosis induction in cancer cells. We show that genetic inhibition of Mcl-1 or CDK9 expression by siRNA is sufficient to mimic flavone-induced apoptosis. Pull-down and in silico docking studies demonstrate that wogonin directly binds to CDK9, presumably to the ATP-binding pocket. In contrast, wogonin does not inhibit CDK2, CDK4 and CDK6 at doses that inhibit CDK9 activity. Furthermore, we show that wogonin preferentially inhibits CDK9 in malignant compared with normal lymphocytes. Thus, our study reveals a new mechanism of anti-cancer action of natural flavones and supports CDK9 as a therapeutic target in oncology.
doi:10.1038/cddis.2011.66
PMCID: PMC3199715  PMID: 21776020
anti-cancer drug; apoptosis; CDK9; Mcl-1; transcription
25.  Cell-type-specific regulation of raft-associated Akt signaling 
Liu, Y | Yang, G | Bu, X | Liu, G | Ding, J | Li, P | Jia, W
Cell Death & Disease  2011;2(4):e145-.
20S-protopanaxadiol (aPPD) is a metabolite of ginseng saponins, which is reported to be pro-apoptotic in some cells but anti-apoptotic in neuronal cells by regulating Akt signaling. Owing to its cholesterol-like structure, we hypothesized that aPPD may regulate Akt signaling by interacting with lipid rafts. Here, we compared Akt signaling in glioblastoma U87MG and neuroblastoma Neuro-2a cells treated with aPPD. aPPD did not change Akt activity in the total plasma membranes of each cell type, but drastically altered the activity of raft-associated Akt. Strikingly, Akt activity was decreased in the rafts of U87MG cells but increased in N2a cells by aPPD through regulating raft-associated dephosphorylation. The bidirectional regulation of raft-associated Akt signaling by aPPD enhanced the chemotoxicity of Paclitaxel or Vinblastine in U87MG cells but attenuated the excitotoxicity of N-methyl--aspartate in N2a cells. Our results demonstrated that the activity of raft-associated but not total membrane Akt determines its cellular functions. Lipid rafts differ in different types of cells, which allows for the possibility of cell-type-specific targeting for which aPPD might prove to be a useful agent.
doi:10.1038/cddis.2011.28
PMCID: PMC3122059  PMID: 21490677
20S-protopanaxadiol; Akt; apoptosis; lipid rafts

Results 1-25 (51)