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1.  Marked methylation changes in intestinal genes during the perinatal period of preterm neonates 
BMC Genomics  2014;15(1):716.
The serious feeding- and microbiota-associated intestinal disease, necrotizing enterocolitis (NEC), occurs mainly in infants born prematurely (5-10% of all newborns) and most frequently after formula-feeding. We hypothesized that changes in gene methylation is involved in the prenatal maturation of the intestine and its response to the first days of formula feeding, potentially leading to NEC in preterm pigs used as models for preterm infants.
Reduced Representation Bisulfite Sequencing (RRBS) was used to assess if changes in intestinal DNA methylation are associated with formula-induced NEC outbreak and advancing age from 10 days before birth to 4 days after birth. Selected key genes with differentially methylated gene regions (DMRs) between groups were further validated by HiSeq-based bisulfite sequencing PCR and RT-qPCR to assess methylation and expression levels. Consistent with the maturation of many intestinal functions in the perinatal period, methylation level of most genes decreased with advancing pre- and postnatal age. The highest number of DMRs was identified between the newborn and 4 d-old preterm pigs. There were few intestinal DMR differences between unaffected pigs and pigs with initial evidence of NEC. In the 4 d-old formula-fed preterm pigs, four genes associated with intestinal metabolism (CYP2W1, GPR146, TOP1MT, CEND1) showed significant hyper-methylation in their promoter CGIs, and thus, down-regulated transcription. Methylation-driven down-regulation of such genes may predispose the immature intestine to later metabolic dysfunctions and severe NEC lesions.
Pre- and postnatal changes in intestinal DNA methylation may contribute to high NEC sensitivity in preterm neonates. Optimizing gene methylation changes via environmental stimuli (e.g. diet, nutrition, gut microbiota), may help to make immature newborn infants more resistant to gut dysfunctions, both short and long term.
Electronic supplementary material
The online version of this article (doi:10.1186/1471-2164-15-716) contains supplementary material, which is available to authorized users.
PMCID: PMC4153944  PMID: 25163507
DNA methylation; Preterm neonates; Necrotizing enterocolitis
2.  MethylPurify: tumor purity deconvolution and differential methylation detection from single tumor DNA methylomes 
Genome Biology  2014;15(7):419.
We propose a statistical algorithm MethylPurify that uses regions with bisulfite reads showing discordant methylation levels to infer tumor purity from tumor samples alone. MethylPurify can identify differentially methylated regions (DMRs) from individual tumor methylome samples, without genomic variation information or prior knowledge from other datasets. In simulations with mixed bisulfite reads from cancer and normal cell lines, MethylPurify correctly inferred tumor purity and identified over 96% of the DMRs. From patient data, MethylPurify gave satisfactory DMR calls from tumor methylome samples alone, and revealed potential missed DMRs by tumor to normal comparison due to tumor heterogeneity.
Electronic supplementary material
The online version of this article (doi:10.1186/s13059-014-0419-x) contains supplementary material, which is available to authorized users.
PMCID: PMC4165374  PMID: 25103624
3.  Obligate mutualism within a host drives the extreme specialization of a fig wasp genome 
Genome Biology  2013;14(12):R141.
Fig pollinating wasps form obligate symbioses with their fig hosts. This mutualism arose approximately 75 million years ago. Unlike many other intimate symbioses, which involve vertical transmission of symbionts to host offspring, female fig wasps fly great distances to transfer horizontally between hosts. In contrast, male wasps are wingless and cannot disperse. Symbionts that keep intimate contact with their hosts often show genome reduction, but it is not clear if the wide dispersal of female fig wasps will counteract this general tendency. We sequenced the genome of the fig wasp Ceratosolen solmsi to address this question.
The genome size of the fig wasp C. solmsi is typical of insects, but has undergone dramatic reductions of gene families involved in environmental sensing and detoxification. The streamlined chemosensory ability reflects the overwhelming importance of females finding trees of their only host species, Ficus hispida, during their fleeting adult lives. Despite long-distance dispersal, little need exists for detoxification or environmental protection because fig wasps spend nearly all of their lives inside a largely benign host. Analyses of transcriptomes in females and males at four key life stages reveal that the extreme anatomical sexual dimorphism of fig wasps may result from a strong bias in sex-differential gene expression.
Our comparison of the C. solmsi genome with other insects provides new insights into the evolution of obligate mutualism. The draft genome of the fig wasp, and transcriptomic comparisons between both sexes at four different life stages, provide insights into the molecular basis for the extreme anatomical sexual dimorphism of this species.
PMCID: PMC4053974  PMID: 24359812
4.  Molecular footprints of domestication and improvement in soybean revealed by whole genome re-sequencing 
BMC Genomics  2013;14:579.
Artificial selection played an important role in the origin of modern Glycine max cultivars from the wild soybean Glycine soja. To elucidate the consequences of artificial selection accompanying the domestication and modern improvement of soybean, 25 new and 30 published whole-genome re-sequencing accessions, which represent wild, domesticated landrace, and Chinese elite soybean populations were analyzed.
A total of 5,102,244 single nucleotide polymorphisms (SNPs) and 707,969 insertion/deletions were identified. Among the SNPs detected, 25.5% were not described previously. We found that artificial selection during domestication led to more pronounced reduction in the genetic diversity of soybean than the switch from landraces to elite cultivars. Only a small proportion (2.99%) of the whole genomic regions appear to be affected by artificial selection for preferred agricultural traits. The selection regions were not distributed randomly or uniformly throughout the genome. Instead, clusters of selection hotspots in certain genomic regions were observed. Moreover, a set of candidate genes (4.38% of the total annotated genes) significantly affected by selection underlying soybean domestication and genetic improvement were identified.
Given the uniqueness of the soybean germplasm sequenced, this study drew a clear picture of human-mediated evolution of the soybean genomes. The genomic resources and information provided by this study would also facilitate the discovery of genes/loci underlying agronomically important traits.
PMCID: PMC3844514  PMID: 23984715
Artificial selection; Evolution; Genetic diversity; Population genomics; Soybean
5.  A rare Von Hippel–Lindau disease that mimics acute myelitis: case report and review of the literature 
Neurological Sciences  2010;32(2):305-307.
Von Hippel–Lindau disease (VHL) comprises a series of complicated clinical manifestations. We hereby report one unique case of VHL with a natural history that mimics acute myelitis. MRI and biopsy in this patient showed multiple solid hemangioblastomas of the central nervous system and kidney. This study further confirmed that VHL is of highly clinical, imaging, and pathological heterogeneity. Diagnosis for VHL should be based on combination of clinical, radiological, pathological, and genetic data.
PMCID: PMC3056988  PMID: 20927563
Von Hippel–Lindau disease; Hemangioblastoma; Clinical manifestation; Magnetic resonance image; Histopathology
6.  The Phosphodiesterase-4 (PDE4) Inhibitor Rolipram Decreases Ethanol Seeking and Consumption in Alcohol-preferring Fawn-Hooded Rats 
Alcohol dependence is a complex psychiatric disorder demanding development of novel pharmacotherapies. Since the cyclic AMP (cAMP) signaling cascade has been implicated in mediating behavioral responses to alcohol, key components in this cascade may serve as potential treatment targets. Phosphodiesterase 4 (PDE4), an enzyme that specifically catalyzes the hydrolysis of cAMP, represents as a key point in regulating intracellular cAMP levels. Thus, it was of interest to determine whether PDE4 was involved in the regulation of alcohol use and abuse.
Male Fawn-Hooded (FH/Wjd) rats were tested for 5% (v/v) ethanol and 10% (w/v) sucrose operant oral self-administration following treatment with the selective PDE4 inhibitor rolipram (0.0125, 0.025, or 0.05 mg/kg, s.c.); rolipram at higher doses (0.05, 0.1, and 0.2 mg/kg, s.c.) was tested to determine its impact on the intake of ethanol, sucrose, or water using the two-bottle choice drinking paradigm. Subsequent open-field testing was performed to evaluate the influence of higher doses of rolipram on locomotor activity.
Acute administration of rolipram dose-dependently reduced operant self-administration of 5% ethanol, but had no effect on 10% sucrose responding. Time-course assessment revealed significant decreases in ethanol consumption after rolipram (0.1, 0.2 mg/kg) treatment in continuous- and intermittent-access to ethanol at 5% or 10%, respectively. Moreover, chronic rolipram treatment time-dependently decreased 5% ethanol consumption and preference during treatment days and after the termination of rolipram administration. Rolipram at the highest doses (0.1 and 0.2 mg/kg) did decrease locomotor activity, but the effect lasted only 10 and 20 min, respectively, which did not likely alter long-term ethanol drinking.
These results suggest that PDE4 plays a role in alcohol seeking and consumption behavior. Drugs interfering with PDE4 may be a potential pharmacotherapy for alcohol dependence.
PMCID: PMC4335658  PMID: 22671516
Cyclic AMP Signaling; Phosphodiesterase-4 (PDE4); Rolipram; FH/Wjd Rat; Ethanol Intake
7.  MicroRNA-133a Regulates Insulin-like Growth Factor-1 Receptor Expression and Vascular Smooth Muscle Cell Proliferation in Murine Atherosclerosis 
Atherosclerosis  2013;232(1):171-179.
MicroRNA-133a (miR-133a) and insulin-like growth factor-1 (IGF-1) are two different molecules known to regulate cardiovascular cell proliferation. This study tested whether miR-133a affects expression of IGF-1 receptor (IGF-1R) and proliferation of IGF-1-stimulated vascular smooth muscle cells (VSMC) in a murine model of atherosclerosis.
Methods and Results
Expression of IGF-1R was analyzed by immuno-fluorescence and immuno-blotting, and miR-133a by qRT-PCR in the aortas of wild-type C57BL/6J (WT) and apolipoprotein-E deficient (ApoE−/−) mice. Compared to those in WT aortas, the IGF-1R and miR-133a levels were lower in ApoE−/− aortas. ApoE−/− VSMC grew slower than WT cells in the cultures with IGF-1-containing medium. MiR-133a-specific inhibitor decreased miR-133a, IGF-1R expression, IGF-1-stimulated VSMC growth in lipoprotein-deficient media. By contrast, miR-133a precursor increased IGF-1R levels and promoted IGF-1-induced VSMC proliferation. In the luciferase-IGF-1R 3’UTR reporter system, the reporter luciferase activity was not inhibited in VSMC with miR-133a overexpression. IGF-1R mRNA half-life in ApoE−/− VSMC was shorter than that in WT VSMC. MiR-133a inhibitor reduced but precursor increased the mRNA half-life, although the effects appeared less striking in ApoE−/− VSMC than in WT cells.
MiR-133a serves as a stimulatory factor for IGF-1R expression through prolonging IGF-1R mRNA half-life. In atherosclerosis induced by ApoE deficiency, reduced miR-133a expression is associated with lower IGF-1R levels and suppressive VSMC growth. Administration of miR-133a precursor may potentiate IGF-1 stimulated VSMC survival and growth.
PMCID: PMC4334121  PMID: 24401233
MicroRNA; Insulin-like growth factor; Artery; Smooth muscle cell; Atherosclerosis
9.  TBX6 Null Variants and a Common Hypomorphic Allele in Congenital Scoliosis 
The New England journal of medicine  2015;372(4):341-350.
Congenital scoliosis is a common type of vertebral malformation. Genetic susceptibility has been implicated in congenital scoliosis.
We evaluated 161 Han Chinese persons with sporadic congenital scoliosis, 166 Han Chinese controls, and 2 pedigrees, family members of which had a 16p11.2 deletion, using comparative genomic hybridization, quantitative polymerase-chain-reaction analysis, and DNA sequencing. We carried out tests of replication using an additional series of 76 Han Chinese persons with congenital scoliosis and a multi-center series of 42 persons with 16p11.2 deletions.
We identified a total of 17 heterozygous TBX6 null mutations in the 161 persons with sporadic congenital scoliosis (11%); we did not observe any null mutations in TBX6 in 166 controls (P<3.8×10−6). These null alleles include copy-number variants (12 instances of a 16p11.2 deletion affecting TBX6) and single-nucleotide variants (1 nonsense and 4 frame-shift mutations). However, the discordant intrafamilial phenotypes of 16p11.2 deletion carriers suggest that heterozygous TBX6 null mutation is insufficient to cause congenital scoliosis. We went on to identify a common TBX6 haplotype as the second risk allele in all 17 carriers of TBX6 null mutations (P<1.1×10−6). Replication studies involving additional persons with congenital scoliosis who carried a deletion affecting TBX6 confirmed this compound inheritance model. In vitro functional assays suggested that the risk haplotype is a hypomorphic allele. Hemivertebrae are characteristic of TBX6-associated congenital scoliosis.
Compound inheritance of a rare null mutation and a hypomorphic allele of TBX6 accounted for up to 11% of congenital scoliosis cases in the series that we analyzed.
PMCID: PMC4326244  PMID: 25564734
10.  An abundance of rare functional variants in 202 drug target genes sequenced in 14,002 people 
Science (New York, N.Y.)  2012;337(6090):100-104.
Rare genetic variants contribute to complex disease risk; however, the abundance of rare variants in human populations remains unknown. We explored this spectrum of variation by sequencing 202 genes encoding drug targets in 14,002 individuals. We find rare variants are abundant (one every 17 bases) and geographically localized, such that even with large sample sizes, rare variant catalogs will be largely incomplete. We used the observed patterns of variation to estimate population growth parameters, the proportion of variants in a given frequency class that are putatively deleterious, and mutation rates for each gene. Overall we conclude that, due to rapid population growth and weak purifying selection, human populations harbor an abundance of rare variants, many of which are deleterious and have relevance to understanding disease risk.
PMCID: PMC4319976  PMID: 22604722
11.  Ghrelin Inhibits the Differentiation of T Helper 17 Cells through mTOR/STAT3 Signaling Pathway 
PLoS ONE  2015;10(2):e0117081.
Enhanced activity of interleukin 17 (IL-17) producing T helper 17 (Th17) cells plays an important role in autoimmune and inflammatory diseases. Significant loss of body weight and appetite is associated with chronic inflammation and immune activation, suggesting the cross talk between immune and neuroendocrine systems. Ghrelin has been shown to regulate the organism immune function. However, the effects of ghrelin on the differentiation of Th17 cells remain elusive. In the present study, we observed the enhanced differentiation of Th17 cells in spleens of growth hormone secretagogue receptor 1a (GHSR1a)-/- mice. Treatment of ghrelin repressed Th17 cell differentiation in a time- and concentration-dependent manner. Phosphorylation of mammalian target of rapamycin (mTOR) and signal transducer and activator of transcription 3 (STAT3) was increased in the spleens of GHSR1a-/- mice. Activation of mTOR signaling by injection of Cre-expressiong adenovirus into tuberous sclerosis complex 1 (TSC1) loxp/loxp mice increased the differentiation of Th17 cells in spleen, which was associated with an increment in the phosphorylation of STAT3. Activation of mTOR signaling by leucine or overexpression of p70 ribosome protein subunit 6 kinase 1 (S6K1) activated mTOR signaling in isolated T cells, while reversed the ghrelin-induced inhibition of iTh17 cell differentiation. In conclusion, mTOR mediates the inhibitory effect of ghrelin on the differentiation of Th17 cells by interacting with STAT3.
PMCID: PMC4319964  PMID: 25658305
12.  Alterations in the Rat Serum Proteome Induced by Prepubertal Exposure to Bisphenol A and Genistein 
Journal of Proteome Research  2014;13(3):1502-1514.
Humans are exposed to an array of chemicals via the food, drink and air, including a significant number that can mimic endogenous hormones. One such chemical is Bisphenol A (BPA), a synthetic chemical that has been shown to cause developmental alterations and to predispose for mammary cancer in rodent models. In contrast, the phytochemical genistein has been reported to suppress chemically induced mammary cancer in rodents, and Asians ingesting a diet high in soy containing genistein have lower incidence of breast and prostate cancers. In this study, we sought to: (1) identify protein biomarkers of susceptibility from blood sera of rats exposed prepubertally to BPA or genistein using Isobaric Tandem Mass Tags quantitative mass spectrometry (TMT-MS) combined with MudPIT technology and, (2) explore the relevance of these proteins to carcinogenesis. Prepubertal exposures to BPA and genistein resulted in altered expression of 63 and 28 proteins in rat sera at postnatal day (PND) 21, and of 9 and 18 proteins in sera at PND35, respectively. This study demonstrates the value of using quantitative proteomic techniques to explore the effect of chemical exposure on the rat serum proteome and its potential for unraveling cellular targets altered by BPA and genistein involved in carcinogenesis.
PMCID: PMC3993963  PMID: 24552547
serum; proteomics; BPA; genistein; carcinogenesis
13.  De Novo Design of Self-Assembling Foldamers That Inhibit Heparin–Protein Interactions 
ACS Chemical Biology  2014;9(4):967-975.
A series of self-associating foldamers have been designed as heparin reversal agents, as antidotes to prevent bleeding due to this potent antithrombotic agent. The foldamers have a repeating sequence of Lys-Sal, in which Sal is 5-amino-2-methoxy-benzoic acid. These foldamers are designed to self-associate along one face of an extended chain in a β-sheet-like interaction. The methoxy groups were included to form intramolecular hydrogen bonds that preclude the formation of very large amyloid-like aggregates, while the positively charged Lys side chains were introduced to interact electrostatically with the highly anionic heparin polymer. The prototype compound (Lys-Sal)4 carboxamide weakly associates in aqueous solution at physiological salt concentration in a monomer-dimer-hexamer equilibrium. The association is greatly enhanced at either high ionic strength or in the presence of a heparin derivative, which is bound tightly. Variants of this foldamer are active in an antithrombin III–factor Xa assay, showing their potential as heparin reversal agents.
PMCID: PMC4324449  PMID: 24491145
14.  Field-based evidence for consistent responses of bacterial communities to copper contamination in two contrasting agricultural soils 
Copper contamination on China's arable land could pose severe economic, ecological and healthy consequences in the coming decades. As the drivers in maintaining ecosystem functioning, the responses of soil microorganisms to long-term copper contamination in different soil ecosystems are still debated. This study investigated the impacts of copper gradients on soil bacterial communities in two agricultural fields with contrasting soil properties. Our results revealed consistent reduction in soil microbial biomass carbon (SMBC) with increasing copper levels in both soils, coupled by significant declines in bacterial abundance in most cases. Despite of contrasting bacterial community structures between the two soils, the bacterial diversity in the copper-contaminated soils showed considerably decreasing patterns when copper levels elevated. High-throughput sequencing revealed copper selection for major bacterial guilds, in particular, Actinobacteria showed tolerance, while Acidobacteria and Chloroflexi were highly sensitive to copper. The thresholds that bacterial communities changed sharply were 800 and 200 added copper mg kg−1 in the fluvo-aquic soil and red soil, respectively, which were similar to the toxicity thresholds (EC50 values) characterized by SMBC. Structural equation model (SEM) analysis ascertained that the shifts of bacterial community composition and diversity were closely related with the changes of SMBC in both soils. Our results provide field-based evidence that copper contamination exhibits consistently negative impacts on soil bacterial communities, and the shifts of bacterial communities could have largely determined the variations of the microbial biomass.
PMCID: PMC4313605
copper contamination; soil bacterial community; diversity; abundance; community composition; soil microbial biomass carbon; field experiment
15.  Deep Sequencing of HIV-Infected Cells: Insights into Nascent Transcription and Host-Directed Therapy 
Journal of Virology  2014;88(16):8768-8782.
Polyadenylated mature mRNAs are the focus of standard transcriptome analyses. However, the profiling of nascent transcripts, which often include nonpolyadenylated RNAs, can unveil novel insights into transcriptional regulation. Here, we separately sequenced total RNAs (Total RNAseq) and mRNAs (mRNAseq) from the same HIV-1-infected human CD4+ T cells. We found that many nonpolyadenylated RNAs were differentially expressed upon HIV-1 infection, and we identified 8 times more differentially expressed genes at 12 h postinfection by Total RNAseq than by mRNAseq. These expression changes were also evident by concurrent changes in introns and were recapitulated by later mRNA changes, revealing an unexpectedly significant delay between transcriptional initiation and mature mRNA production early after HIV-1 infection. We computationally derived and validated the underlying regulatory programs, and we predicted drugs capable of reversing these HIV-1-induced expression changes followed by experimental confirmation. Our results show that combined total and mRNA transcriptome analysis is essential for fully capturing the early host response to virus infection and provide a framework for identifying candidate drugs for host-directed therapy against HIV/AIDS.
IMPORTANCE In this study, we used mass sequencing to identify genes differentially expressed in CD4+ T cells during HIV-1 infection. To our surprise, we found many differentially expressed genes early after infection by analyzing both newly transcribed unprocessed pre-mRNAs and fully processed mRNAs, but not by analyzing mRNAs alone, indicating a significant delay between transcription initiation and mRNA production early after HIV-1 infection. These results also show that important findings could be missed by the standard practice of analyzing mRNAs alone. We then derived the regulatory mechanisms driving the observed expression changes using integrative computational analyses. Further, we predicted drugs that could reverse the observed expression changes induced by HIV-1 infection and showed that one of the predicted drugs indeed potently inhibited HIV-1 infection. This shows that it is possible to identify candidate drugs for host-directed therapy against HIV/AIDS using our genomics-based approach.
PMCID: PMC4136300  PMID: 24850744
16.  Novel variation and de novo mutation rates in population-wide de novo assembled Danish trios 
Nature Communications  2015;6:5969.
Building a population-specific catalogue of single nucleotide variants (SNVs), indels and structural variants (SVs) with frequencies, termed a national pan-genome, is critical for further advancing clinical and public health genetics in large cohorts. Here we report a Danish pan-genome obtained from sequencing 10 trios to high depth (50 × ). We report 536k novel SNVs and 283k novel short indels from mapping approaches and develop a population-wide de novo assembly approach to identify 132k novel indels larger than 10 nucleotides with low false discovery rates. We identify a higher proportion of indels and SVs than previous efforts showing the merits of high coverage and de novo assembly approaches. In addition, we use trio information to identify de novo mutations and use a probabilistic method to provide direct estimates of 1.27e−8 and 1.5e−9 per nucleotide per generation for SNVs and indels, respectively.
The generation of a national pan-genome, a population-specific catalogue of genetic variation, may advance the impact of clinical genetics studies. Here the Besenbacher et al. carry out deep sequencing and de novo assembly of 10 parent–child trios to generate a Danish pan-genome that provides insight into structural variation, de novo mutation rates and variant calling.
PMCID: PMC4309431  PMID: 25597990
17.  Stigma against HIV/AIDS among female sex workers and general migrant women in eastern China 
BMC Women's Health  2015;15:2.
HIV related stigma is a recognized barrier to early detection of HIV and causes great suffering for those affected. However, data regarding HIV related stigma among female sex workers (FSW) in China was limited, with none for comparison between FSW and general migrant women (GMW). Therefore, the aim of this study was to examine HIV related stigma among FSW and GMW in Shanghai, China.
A community based cross-sectional study with face-to-face interviews was conducted in Shanghai (September 2011 through December 2012), using a structured questionnaire.HIV related stigma scores were examined graphically using boxplot. A logistic regression analysis with the proportional odds model was employed to identify factors affecting HIV related stigma scores.
A total of 1,396 subjects, including 721 FSW and 675 GMW, were recruited in the present study. Both groups had substantial misconceptions about HIV/AIDS, although FSW had slightly higher scores on average. Both groups showed a medium level of HIV related stigma (38.34 ± 6.21 and 38.35 ± 6.86 for FSW and GMW, respectively). For the FSW, higher levels of stigma were observed for those who were in the older age groups (age 26-35 years, OR, 2.06, 95% CI 1.06-4.01), those who were married (OR, 1.62, 95% CI 1.03-2.54), and those who were working at lower-level sex service sites (OR, 1.60, 95% CI 1.06-2.43). Conversely, HIV knowledge was inversely associated with the level of HIV related stigma (OR, 0.93, 95% CI 0.87-0.98).Among GMW participating in the study, those age in the 26-35 years were more likely to show higher level of stigma (OR, 2.61, 95% CI 1.03-2.54), and HIV knowledge was found to be inversely associated with the HIV related stigma level as well (OR, 0.89, 95% CI 0.84-0.95).
The present study suggests that there is an urgent need for the development of appropriate education strategies to reduce HIV related stigma among FSW and GMW in Shanghai, China. In particular, older women, less educated women, and women that have lived in Shanghai a relatively long time should be targeted in future stigma reduction programs.
Electronic supplementary material
The online version of this article (doi:10.1186/s12905-014-0160-3) contains supplementary material, which is available to authorized users.
PMCID: PMC4316807  PMID: 25608866
18.  Characterizing protein crystal contacts and their role in crystallization: rubredoxin as a case study 
Soft matter  2014;10(2):290-302.
The fields of structural biology and soft matter have independently sought out fundamental principles to rationalize protein crystallization. Yet the conceptual differences and the limited overlap between the two disciplines have thus far prevented a comprehensive understanding of the phenomenon to emerge. We conduct a computational study of proteins from the rubredoxin family that bridges the two fields. Using atomistic simulations, we characterize their crystal contacts, and accordingly parameterize patchy particle models. Comparing the phase diagrams of these schematic models with experimental results enables us to critically examine the assumptions behind the two approaches. The study also reveals features of protein-protein interactions that can be leveraged to crystallize proteins more generally.
PMCID: PMC3907588  PMID: 24489597
19.  Luminescence signature of free exciton dissociation and liberated electron transfer across the junction of graphene/GaN hybrid structure 
Scientific Reports  2015;5:7687.
Large-area graphene grown on Cu foil with chemical vapor deposition was transferred onto intentionally undoped GaN epilayer to form a graphene/GaN Schottky junction. Optical spectroscopic techniques including steady-state and time-resolved photoluminescence (PL) were employed to investigate the electron transfer between graphene and n-type GaN at different temperatures. By comparing the near-band-edge excitonic emissions before and after the graphene covering, some structures in the excitonic PL spectra are found to show interesting changes. In particular, a distinct “dip” structure is found to develop at the center of the free exciton emission peak as the temperature goes up. A mechanism that the first dissociation of some freely moveable excitons at the interface was followed by transfer of liberated electrons over the junction barrier is proposed to interpret the appearance and development of the “dip” structure. The formation and evolution process of this “dip” structure can be well resolved from the measured time-resolved PL spectra. First-principles simulations provide clear evidence of finite electron transfer at the interface between graphene and GaN.
PMCID: PMC4286737  PMID: 25567005
20.  Complete Genome Sequence of the Fish Pathogen Yersinia ruckeri Strain SC09, Isolated from Diseased Ictalurus punctatus in China 
Genome Announcements  2015;3(1):e01327-14.
Yersinia ruckeri SC09 is a Gram-negative bacterium isolated from a moribund Ictalurus punctatus collected in Jianyang, China. Here, we report the complete genome sequence of this microorganism to facilitate the investigation of its pathogenicity and to reevaluate its taxonomic position.
PMCID: PMC4290980  PMID: 25573927
21.  Flipping in the Pore: Discovery of Dual Inhibitors That Bind in Different Orientations to the Wild-Type versus the Amantadine-Resistant S31N Mutant of the Influenza A Virus M2 Proton Channel 
Journal of the American Chemical Society  2014;136(52):17987-17995.
Influenza virus infections lead to numerous deaths and millions of hospitalizations each year. One challenge facing anti-influenza drug development is the heterogeneity of the circulating influenza viruses, which comprise several strains with variable susceptibility to antiviral drugs. For example, the wild-type (WT) influenza A viruses, such as the seasonal H1N1, tend to be sensitive to antiviral drugs, amantadine and rimantadine, while the S31N mutant viruses, such as the pandemic 2009 H1N1 (H1N1pdm09) and seasonal H3N2, are resistant to this class of drugs. Thus, drugs targeting both WT and the S31N mutant are highly desired. We report our design of a novel class of dual inhibitors along with their ion channel blockage and antiviral activities. The potency of the most active compound 11 in inhibiting WT and the S31N mutant influenza viruses is comparable with that of amantadine in inhibiting WT influenza virus. Solution NMR studies and molecular dynamics (MD) simulations of drug-M2 interactions supported our design hypothesis: namely, the dual inhibitor binds in the WT M2 channel with an aromatic group facing down toward the C-terminus, while the same drug binds in the S31N M2 channel with its aromatic group facing up toward the N-terminus. The flip-flop mode of drug binding correlates with the structure–activity relationship (SAR) and has paved the way for the next round of rational design of broad-spectrum antiviral drugs.
PMCID: PMC4286326  PMID: 25470189
22.  Targeted Therapy Resistance Mediated by Dynamic Regulation of Extrachromosomal Mutant EGFR DNA 
Science (New York, N.Y.)  2013;343(6166):72-76.
Intratumoral heterogeneity contributes to cancer drug resistance, but the underlying mechanisms are not understood. Single-cell analyses of patient-derived models and clinical samples from glioblastoma patients treated with epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKIs) demonstrate that tumor cells reversibly up-regulate or suppress mutant EGFR expression, conferring distinct cellular phenotypes to reach an optimal equilibrium for growth. Resistance to EGFR TKIs is shown to occur by elimination of mutant EGFR from extrachromosomal DNA. After drug withdrawal, reemergence of clonal EGFR mutations on extrachromosomal DNA follows. These results indicate a highly specific, dynamic, and adaptive route by which cancers can evade therapies that target oncogenes maintained on extrachromosomal DNA.
PMCID: PMC4049335  PMID: 24310612
23.  Numerical Analysis of the Effect of T-tubule Location on Calcium Transient in Ventricular Myocytes 
Bio-medical materials and engineering  2014;24(1):10.3233/BME-130932.
Intracellular calcium (Ca2+) signaling in cardiac myocytes is vital for proper functioning of the heart. Understanding the intracellular Ca2+ dynamics would give an insight into the functions of normal and diseased hearts. In the current study, spatiotemporal Ca2+ dynamics is investigated in ventricular myocytes by considering Ca2+ release and re-uptake via sarcolemma and transverse tubules (T-tubules), Ca2+ diffusion and buffering in the cytosol, and the blockade of Ca2+ activities associated with the sarcoplasmic reticulum. This study is carried out using a three dimensional (3D) geometric model of a branch of T-tubule extracted from the electron microscopy (EM) images of a partial ventricular myocyte. Mathematical modeling is done by using a system of partial differential equations involving Ca2+ , buffers, and membrane channels. Numerical simulation results suggest that a lack of T-tubule structure at the vicinity of the cell surface could increase the peak time of Ca2+ concentration in myocytes. The results also show that T-tubules and mobile buffers play an important role in the regulation of Ca2+ transient in ventricular myocytes.
PMCID: PMC3835764  PMID: 24212025
Cardiac myocytes; calcium dynamics; reaction-diffusion equations; numerical analysis; finite element method
24.  New Software Developments for Quality Mesh Generation and Optimization from Biomedical Imaging Data 
Computer methods and programs in biomedicine  2013;113(1):10.1016/j.cmpb.2013.08.009.
In this paper we present a new software toolkit for generating and optimizing surface and volumetric meshes from three-dimensional (3D) biomedical imaging data, targeted at image-based finite element analysis of some biomedical activities in a single material domain. Our toolkit includes a series of geometric processing algorithms including surface re-meshing and quality-guaranteed tetrahedral mesh generation and optimization. All methods described have been encapsulated into a user-friendly graphical interface for easy manipulation and informative visualization of biomedical images and mesh models. Numerous examples are presented to demonstrate the effectiveness and efficiency of the described methods and toolkit.
PMCID: PMC3836056  PMID: 24252469
Geometric modeling; tetrahedral mesh generation; tetrahedral mesh smoothing; surface re-meshing; biomedical images
25.  Targeted next-generation sequencing as a comprehensive test for patients with and female carriers of DMD/BMD: a multi-population diagnostic study 
Duchenne and Becker muscular dystrophies (DMD/BMD) are the most commonly inherited neuromuscular disease. However, accurate and convenient molecular diagnosis cannot be achieved easily because of the enormous size of the dystrophin gene and complex causative mutation spectrum. Such traditional methods as multiplex ligation-dependent probe amplification plus Sanger sequencing require multiple steps to fulfill the diagnosis of DMD/BMD. Here, we introduce a new single-step method for the genetic analysis of DMD patients and female carriers in real clinical settings and demonstrate the validation of its accuracy. A total of 89 patients, 18 female carriers and 245 non-DMD patients were evaluated using our targeted NGS approaches. Compared with traditional methods, our new method yielded 99.99% specificity and 98.96% sensitivity for copy number variations detection and 100% accuracy for the identification of single-nucleotide variation mutations. Additionally, this method is able to detect partial deletions/duplications, thus offering precise personal DMD gene information for gene therapy. We detected novel partial deletions of exons in nine samples for which the breakpoints were located within exonic regions. The results proved that our new method is suitable for routine clinical practice, with shorter turnaround time, higher accuracy, and better insight into comprehensive genetic information (detailed breakpoints) for ensuing gene therapy.
PMCID: PMC3865410  PMID: 23756440
targeted NGS; genetic diagnosis; Duchenne and Becker muscular dystrophies; statistical analysis for CNVs; breakpoints

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