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Purpose:Multiple myeloma is a malignancy of plasma cells. Extensive genetic and transcriptional characterization of myeloma has identified subtypes with prognostic and therapeutic implications. In contrast, relatively little is known about the myeloma epigenome.Experimental Design:CD138+CD38+ myeloma cells were isolated from fresh bone marrow aspirate or the same aspirate after freezing for 1–6 months. Gene expression and chromatin accessibility were compared between fresh and frozen samples by RNA sequencing (RNA-seq) and assay for transpose accessible chromatin sequencing (ATAC-seq). Chromatin accessible regions were used to identify regulatory RNA expression in more than 700 samples from newly diagnosed patients in the Multiple Myeloma Research Foundation CoMMpass trial (NCT01454297).Results:Gene expression and chromatin accessibility of cryopreserved myeloma recapitulated that of freshly isolated samples. ATAC-seq performed on a series of biobanked specimens identified thousands of chromatin accessible regions with hundreds being highly coordinated with gene expression. More than 4,700 of these chromatin accessible regions were transcribed in newly diagnosed myelomas from the CoMMpass trial. Regulatory element activity alone recapitulated myeloma gene expression subtypes, and in particular myeloma subtypes with immunoglobulin heavy chain translocations were defined by transcription of distal regulatory elements. Moreover, enhancer activity predicted oncogene expression implicating gene regulatory mechanisms in aggressive myeloma.Conclusions:These data demonstrate the feasibility of using biobanked specimens for retrospective studies of the myeloma epigenome and illustrate the unique enhancer landscapes of myeloma subtypes that are coupled to gene expression and disease progression.

Enrichment of transcription factor consensus binding motifs in regulatory elements predictive of gene expression associated with poor overall survival.

Correlation of regulatory element transcription and gene expression in CoMMpass specimens.

ATAC-seq data for Emory patients.

Correlation of chromatin accessibility and gene expression for the 5% most variably expressed genes.

Regulatory elements predictive of gene expression for genes prognostic of overall survival in CoMMpass.

Regulatory element transcription in newly diagnosed myelomas from CoMMpass.

Gene set enrichment analysis of transcribed regulatory elements in CoMMpass.

The public transport system is considered as one of the most important subsystems in metropolises for achieving sustainability objectives by mediating resources and travel demand. Representing the various urban transport networks is crucial in understanding travel behavior and the function of the transport system. However, previous studies have ignored the coupling relationships between multi-mode transport networks and travel flows. To address this problem, we constructed a multilayer network to illustrate two modes of transport (bus and metro) by assigning weights of travel flow and efficiency. We explored the scaling of the public transport system to validate the multilayer network and offered new visions for transportation improvements by considering population. The proposed methodology was demonstrated by using public transport datasets of Shanghai, China. For both the bus network and multilayer network, the scaling of node degree versus Population were explored at 1 km * 1 km urban cells. The results suggested that in the multilayer network, the scaling relations between node degree and population can provide valuable insights into quantifying the integration between the public transport system and urban land use, which will benefit sustainable improvements to cities.

In recent years, deploying Internet of Things (IoT) in electronic healthcare systems (EHS) has made great progress in healthcare detection. It is extremely important to reduce the cost of communication and ensure the authenticity and integrity of data. A linearly homomorphic signature scheme can solve the above problems. However, when the scale of EHS is too large, the transmission, storage and verification of signatures need a high cost. An aggregate signature can combine many signatures generated by many different users into a short one. Therefore, only one aggregate signature needs to be processed during verification, transmission and storage. Combining the advantages of aggregate signature and linearly homomorphic signature, this paper proposes an aggregate signature scheme based on a linearly homomorphic signature for EHS, which has both linear homomorphism and aggregation, and realizes double data compression. Moreover, our scheme can resist a potential real attack, named a coalition attack. The security of this scheme is rigorously demonstrated based on the computational Diffie–Hellman assumption in the random oracle model.

Purpose: Multiple myeloma is a malignancy of plasma cells. Extensive genetic and transcriptional characterization of myeloma has identified subtypes with prognostic and therapeutic implications. In contrast, relatively little is known about the myeloma epigenome. Experimental Design: CD138+CD38+ myeloma cells were isolated from fresh bone marrow aspirate or the same aspirate after freezing for 1–6 months. Gene expression and chromatin accessibility were compared between fresh and frozen samples by RNA sequencing (RNA-seq) and assay for transpose accessible chromatin sequencing (ATAC-seq). Chromatin accessible regions were used to identify regulatory RNA expression in more than 700 samples from newly diagnosed patients in the Multiple Myeloma Research Foundation CoMMpass trial (NCT01454297). Results: Gene expression and chromatin accessibility of cryopreserved myeloma recapitulated that of freshly isolated samples. ATAC-seq performed on a series of biobanked specimens identified thousands of chromatin accessible regions with hundreds being highly coordinated with gene expression. More than 4,700 of these chromatin accessible regions were transcribed in newly diagnosed myelomas from the CoMMpass trial. Regulatory element activity alone recapitulated myeloma gene expression subtypes, and in particular myeloma subtypes with immunoglobulin heavy chain translocations were defined by transcription of distal regulatory elements. Moreover, enhancer activity predicted oncogene expression implicating gene regulatory mechanisms in aggressive myeloma. Conclusions: These data demonstrate the feasibility of using biobanked specimens for retrospective studies of the myeloma epigenome and illustrate the unique enhancer landscapes of myeloma subtypes that are coupled to gene expression and disease progression.

Background Clostridium saccharoperbutylacetonicum N1-4 (HMT) is a strictly anaerobic, spore-forming Gram-positive bacterium capable of hyper-butanol production through the well-known acetone–butanol–ethanol fermentation process. Recently, five putative RRNPP-type QSSs (here designated as QSS1 to QSS5) were predicted in this bacterial strain, each of which comprises a putative RRNPP-type regulator (QssR1 to QssR5) and a cognate signaling peptide precursor (QssP1 to QssP5). In addition, both proteins are encoded by the same operon. The functions of these multiple RRNPP-type QSSs are unknown. Results To elucidate the function of multiple RRNPP-type QSSs as related to cell metabolism and solvent production in N1-4 (HMT), we constructed qssR-deficient mutants ΔR1, ΔR2, ΔR3 and ΔR5 through gene deletion using CRISPR–Cas9 and N1-4-dcas9-R4 (with the QssR4 expression suppressed using CRISPR–dCas9). We also constructed complementation strains by overexpressing the corresponding regulator gene. Based on systematic characterization, results indicate that QSS1, QSS2, QSS3, and QSS5 positively regulate the sol operon expression and thus solvent production, but they likely negatively regulate cell motility. Consequently, QSS4 might not directly regulate solvent production, but positively affect cell migration. In addition, QSS3 and QSS5 appear to positively regulate sporulation efficiency. Conclusions Our study provides the first insights into the roles of multiple RRNPP-type QSSs of C. saccharoperbutylacetonicum for the regulation of solvent production, cell motility, and sporulation. Results of this study expand our knowledge of how multiple paralogous QSSs are involved in the regulation of essential bacterial metabolism pathways.

Bioproduction of renewable chemicals is considered as an urgent solution for fossil energy crisis. However, despite tremendous efforts, it is still challenging to generate microbial strains that can produce target biochemical to high levels. Here, we report an example of biosynthesis of high-value and easy-recoverable derivatives built upon natural microbial pathways, leading to improvement in bioproduction efficiency. By leveraging pathways in solventogenic clostridia for co-producing acyl-CoAs, acids and alcohols as precursors, through rational screening for host strains and enzymes, systematic metabolic engineering-including elimination of putative prophages, we develop strains that can produce 20.3 g/L butyl acetate and 1.6 g/L butyl butyrate. Techno-economic analysis results suggest the economic competitiveness of our developed bioprocess. Our principles of selecting the most appropriate host for specific bioproduction and engineering microbial chassis to produce high-value and easy-separable end products may be applicable to other bioprocesses., Esters can be used as fuels and specialty chemicals for food flavoring, cosmetic and pharmaceutical industries. Here, the authors systematically engineer clostridia, including discovery and deletion of prophages to increase strain stability, for the production of butyl acetate and butyl butyrate from corn stover at low cost.

Fine-grained subjective partitioning of urban space using human activity flows reveals actual human activity spaces with high resolution, which has great implications for the development and validation of planning strategies. This paper presents a new method for fine-grained subjective partitioning of urban space based on the combination of network analysis and human interactions from social media. Three main procedures are involved in this method: 1) a cut-off point for hierarchical partitioning is determined by fitting the probability distribution function of human activity patterns; 2) based on this cut-off point, improved hierarchical weighted-directed spatial networks are constructed by incorporating a gravity model into conventional spatial networks to take into account the importance of the attraction of nodes in shaping urban space; and 3) the hierarchical and fine-grained partitioning results, which reveal the actual human activity spaces with high resolution at multiscale are obtained by implementing a spatial community detection algorithm in these networks. A case study, using a real-world dataset from the capital of China validates the effectiveness of the proposed method. By analyzing the results from Beijing, we concluded that the social media, a gravity model, and the hierarchical subjective communities detected from the hierarchical human activity networks are all outstanding contributors to the fine-grained subjective partitioning of urban spaces.

Clostridium saccharoperbutylacetonicum N1-4 (HMT) is known as a hyper acetone-butanol-ethanol (ABE) producing strain. Within its genome, there is an endogenous megaplasmid (Csp_135p) of 136 kb, which contains 104 protein-encoding genes and two pseudogenes. Till now, the function of the megaplasmid is unknown; meanwhile the existence of the megaplasmid could cause instability for the strain performance and barrier for efficient genome engineering. In order to investigate the function of the megaplasmid as related to ABE fermentation and other metabolisms, here we successfully eliminated the megaplasmid using the CRISPR-Cas9 system, generating the plasmid-null strain N1-4-C. Results from systematic characterization experiments indicated that the N1-4-C strain exhibited higher plasmid transformation efficiency and better plasmid stability than the N1-4 (HMT) strain. In addition, the N1-4-C strain could produce slightly higher concentration of butanol and ABE, along with more efficient re-assimilation of acids. Since there is a gene encoding a lipase on Csp_135p, we evaluated the capability for ester production of both N1-4-C and N1-4 (HMT) strains. Results demonstrated that the Csp_135p plasmid also contributed to the ester production (such as butyl acetate and butyl butyrate; this is the first report for ester production in C. saccharoperbutylacetonicum ) . This study demonstrated that the CRISPR-Cas9 system can be used as an efficient tool for the curing of endogenous plasmids. The plasmid-null N1-4-C strain can serve as a great platform for the development of more robust strains for biofuel and biochemical production.

Transit-oriented development (TOD) is among the most feasible strategies for relieving urban issues caused by the unbalanced development of transportation and land use. This study proposes a multiobjective TOD land use design framework for the optimization of the land use layout in station catchments. Given the high density and diverse development in Chinese megacities, a planning model that considers nonlinear impacts on ridership, land use efficiency, quality of life, and the environment is constructed. The model applies fine-grained geo-big data to fill gaps in the empirical and statistical data and improve practicability. A genetic multiobjective optimization approach without reliance on objective weighting is used to generate alternative land use schemes. A metro station in Shanghai is applied as a case study. The results indicate that the proposed ridership objective outperforms the commonly used linear function, and the optimization method has superior extreme optima and convergence to baseline models. We also discuss the consistencies and conflicts in the objectives and provide a balanced land use scheme considering local policies. This work provides suggestions for sustainable urban design with coordinated land use and transportation.

Protein homeostasis is critical for maintaining eukaryotic cell function as well as responses to intrinsic and extrinsic stress. The proteasome is a major portion of the proteolytic machinery in mammalian cells and plays an important role in protein homeostasis. Multiple myeloma (MM) is a plasma cell malignancy with high production of immunoglobulins and is especially sensitive to treatments that impact protein catabolism. Therapeutic agents such as proteasome inhibitors have demonstrated significant benefit for myeloma patients in all treatment phases. Here, we demonstrate that the 11S proteasome activator PA28α is upregulated in MM cells and is key for myeloma cell growth and proliferation. PA28α also regulates MM cell sensitivity to proteasome inhibitors. Downregulation of PA28α inhibits both proteasomal load and activity, resulting in a change in protein homeostasis less dependent on the proteasome and leads to cell resistance to proteasome inhibitors. Thus, our findings suggest an important role of PA28α in MM biology, and also provides a new approach for targeting the ubiquitin-proteasome system and ultimately sensitivity to proteasome inhibitors.

Production of renewable chemicals through biological routes is considered as an urgent solution for fossil energy crisis. However, endproduct toxicity inhibits microbial performance and is a key bottleneck for biochemical production. To address this challenge, here we report an example of biosynthesis of high-value and easy-recoverable derivatives to alleviate endproduct toxicity and enhance bioproduction efficiency. By leveraging the natural pathways in solventogenic clostridia for co-producing acyl-CoAs, acids and alcohols as precursors, through rational screening for host strains and enzymes, systematic metabolic engineering— including rational organization of ester-synthesizing enzymes inside of the cell, and elimination of putative prophages, we developed strains that can produce 20.3 g/L butyl acetate and 1.6 g/L butyl butyrate respectively, which were both the unprecedented levels in microbial hosts. Techno-economic analysis indicated a production cost of $986 per metric tonne for butyl acetate production from corn stover comparing to the market price of $1,200-1,400 per metric tonne of butyl acetate, suggesting the economic competitiveness of our developed bioprocess. Our principles of selecting the most appropriate host for specific bioproduction and engineering microbial chassis to produce high-value and easy-separable endproducts are highly applicable to other bioprocesses, and could lead to breakthroughs in biofuel/biochemical production and general bioeconomy.

Alleviating public traffic congestion is an efficient and effective way to improve the travel time reliability and quality of public transport services. The existing public network optimization models usually ignored the essential impact of public traffic congestion on the performance of public transport service. To address this problem, this study proposes a data-based methodology to estimate the traffic congestion of road segments between bus stops (RSBs). The proposed methodology involves two steps: 1) Extracting three traffic indicators of the RSBs from smart card data and bus trajectory data, 2) The self-organizing map (SOM) is used to cluster and effectively recognize traffic patterns embedded in the RSBs. Furthermore, a congestion index for ranking the SOM clusters is developed to determine the congested RSBs. A case study using real-world datasets from a public transport system validates the proposed methodology. Based on the congested RSBs, an exploratory example of public transport network optimization is discussed and evaluated using a genetic algorithm. The clustering results showed that the SOM could suitably reflect the traffic characteristics and estimate traffic congestion of the RSBs. The results obtained in this study are expected to demonstrate the usefulness of the proposed methodology in sustainable public transport improvements.

Additional file 1: Table S1. Primers used in this study. Table S2. Comparative pair-alignment data of amino acid sequences of the Clostridium saccharoperbutylacetonicum RRNPP-type transcriptional regulators. Table S3. Comparative pair-alignment data of amino acid sequences of the signaling-peptide precursors of the five Clostridium saccharoperbutylacetonicum RRNPP-type quorum sensing systems. Figure S1. Schematic of two rounds of PCR to obtain the DNA fragment containing 20-nt gRNA sequence for constructing the CRISPR–Cas9 plasmid for gene deletion. Figure S2. Confirmation of gene deletion by colony PCR. Figure S3. Transcriptional analyses of qssR1, qssR2, qssR3, qssR4 and qssR5 in wild-type N1-4 (HMT) and relevant mutant strains using qRT-PCR. Figure S4. Transcriptional analyses of qssR1, qssR2, qssR3, qssR4 and qssR5 in wild-type N1-4 (HMT) and relevant mutant strains using qRT-PCR. Figure S5. Transcriptional analyses of qssR1, qssR2, qssR3, qssR4 and qssR5 in wild-type N1-4 (HMT) and relevant mutant strains using qRT-PCR. Figure S6. Transcriptional analyses of qssP1, qssP2, qssP3, qssP4 and qssP5 in wild-type N1-4 (HMT) and relevant mutant strains using qRT-PCR. Figure S7. Transcriptional analyses of qssP1, qssP2, qssP3, qssP4 and qssP5 in wild-type N1-4 (HMT) and relevant mutant strains using qRT-PCR. Figure S8. Transcriptional analyses of qssP1, qssP2, qssP3, qssP4 and qssP5 in wild-type N1-4 (HMT) and relevant mutant strains using qRT-PCR. Figure S9. Transcriptional analyses of spo0E-like genes in wild-type N1-4 (HMT) and relevant mutant strains using qRT-PCR.

The spatial relationship of newly grown urban patches to existing urban areas lies at the core of understanding the properties of urban expansion dynamics. Some existing landscape metrics have been used to identify patch expansion types, i.e., infilling, edge-expansion and outlying, capturing the evolution process of urban expansion patterns based on quantifying spatial relationship. However, these existing metrics cannot comprehensively describe the spatial distributions of all new patches relative to old built-up areas, especially for outlying patches, which are the significant elements affecting the urban expansion pattern. We propose a new landscape metric, the Proximity Expansion Index (PEI), to address this problem by incorporating two factors of proximity - distance and boundary sharing rate to old patches. The value of PEI is continuous and has the clear physical meaning for depicting the gradient of the spatial relationship. The landscape expansion types are then redefined by PEI, while the sprawl level of outlying patches is clearly reflected. The variants of PEI are designed as global indices to capture information of the dynamic process of urban expansion from a bottom-up view. We selected Wuhan, a metropolis in central China, as a case area to evaluate PEI based on four periods of remote sensing images (1995, 2000, 2005 and 2010). The results show that the spatial pattern of urban expansion becomes increasingly dispersed, demonstrating that PEI is capable of capturing information of urban expansion evolution. PEI can depict the spatial relationship between new and old patches in a more detailed way by comparing PEI and previous metrics. Using PEI, we can also discover regions of great significance, called outlying seed regions, which have a profound impact on the coalescence of urban morphology.

Social media data have been widely used to enrich human-centric information for situational awareness and disaster assessment. Owing to the granularity of topics detected from disaster-related contents, the effectiveness of social media in reflecting disaster losses is still limited. To address this limitation, this study developed a methodology for assessing disaster losses using social media data, which was composed of data preprocessing, fine-grained topic extraction, and quantitative damage estimation. The proposed methodology was demonstrated in a case study of Typhoons Hato & Pakhar, which caused persistent damage in southern China from August 22 to August 30, 2017. The results highlighted the capability of the proposed methodology in using fine-grained topics to assess disaster losses, e.g., the disaster losses were significantly correlated with infrastructure damage-related topics. The study provided useful insights in disaster damage assessment through the fine-grained topics in social media.

To improve the utilization of sucrose for 2,3-butanediol (2,3-BD) production, four combinations of heterologous energy-conserving sucrose utilization pathways were introduced into Bacillus subtilis Δtet strain (a derivative from B. subtilis 168) and B. subtilis FJ-1 strain (a new isolate in our lab). Results demonstrated that the combination of cscB (encoding sucrose permease) from Escherichia coli and gtfA (encoding sucrose phosphorylase) from Streptococcus mutans showed the most remarkable enhancement for the 2,3-BD production. With sucrose and sugar cane juice as substrate, respectively, the Δtet-CEG strain (B. subtilis Δtet strain containing the energy-conserving sucrose utilization pathway as referred above) showed 36.5% and 24.7% increase in 2,3-BD production than the control Δtet strain, and the FJ-1-CEG strain also produced 23.8% and 44.5% more 2,3-BD than the control FJ-1 strain. The metabolic engineering strategy demonstrated in this study can be extensively applied to other microorganisms for r...

14-3-3 proteins are a family of master regulators of intracellular signaling, yet their impact on proteasome function is unknown. We demonstrate that 14-3-3ζ binds the 11S proteasome activator, limiting proteasome assembly and cellular capacity for protein degradation. To define the functional impact of 14-3-3ζ proteasomal binding in myeloma cells, silencing and overexpression experiments are performed. We find that downregulation of 14-3-3ζ impairs myeloma cell growth and confers resistance to clinically used proteasome inhibitors. In a large cohort of newly diagnosed myeloma patients, elevated expression of 14-3-3ζ is associated with high risk myeloma genetic subtypes and worse prognosis overall. Our work demonstrates the important role of 14-3-3ζ in regulating proteasome function, myeloma cell growth and sensitivity to therapeutics, and suggests regulation of 14-3-3ζ as a new approach in myeloma therapy.

Introduction Extensive genomic characterization of multiple myeloma has identified subtypes with prognostic and therapeutic implications. In contrast, less is known about the myeloma epigenome. One challenge that has hindered epigenetic studies are assays amenable to biobanked specimens. Here, we sought to determine whether ATAC-seq and RNA-seq of myeloma cells from cryopreserved bone marrow aspirates recapitulated those from fresh samples and used this approach to investigate enhancers of myeloma oncogenes. Methods Consent and collection of specimens followed approved Institutional Review Board protocols. Mononuclear cells were enrichment by Ficoll gradient centrifugation and were either cryopreserved in 10% DMSO and RPMI media with 10% FBS or used to isolate viable CD138+CD38+ myeloma cells. RNA-seq used the mRNA HyperPrep kit (Kapa Biosystems) with RNA from 50,000 cells. ATAC-seq used the Tn5 transposase (Illumina) on 20,000 cells. Sequencing was performed on an HiSeq 4000 (Illumina). Sequencing data were quality and adapter trimmed using Trim Galore! And mapped to the GRCh37 genome using STAR (RNA-seq) or bowtie2 (ATAC-seq). MACS2 was used to determine chromatin accessible regions and R was used for downstream analyses. H3K27ac ChIP-seq from Jin et al. (Blood, 2018) were downloaded from the European nucleotide archive (PRJEB25605). RNA-seq from CoMMpass (NCT01454297) were downloaded from dbGaP phs000748.v7.p4. Enhancer RNAs were interrogated in intergenic regions excluding 500 bp upstream of TSSs and 5 kb downstream of transcription termination sites to avoid contamination from exonic mRNAs or intronic pre-mRNAs. Results We compared RNA-seq and ATAC-seq data from myeloma cells isolated from fresh bone marrow aspirates to those cryopreserved for up to 6 months from the same aspirate. RNA-seq and ATAC-seq data from fresh and frozen samples from the same aspirate were highly correlated with each other but distinct from other samples as depicted by principal component analysis (Fig. A,B). Inspection of CCND1 showed high levels of RNA in two patients and this was consistent in both fresh and frozen specimens as well as with FISH results indicating a t(11;14) translocation in these samples (Fig. C). Similarly, fresh and frozen specimens from the same patient showed consistent expression for CCND2 and MYC and these data corresponded with chromatin accessibility found near these genes (Fig. D, see regions shaded in gray). Based on these results we expanded our analysis to include 8 biobanked specimens, which identified 91,632 regions of chromatin accessibility that were enriched around plasma cell lineage genes such as IRF4, CD38, SLAMF7, and IGH. Chromatin accessibility often predicted proximal gene expression and this was especially pronounced for regions enriched for histone 3 lysine 27 acetylation (H3K27ac) - a mark of enhancers. Active enhancers are sometimes demarcated by enhancer RNAs (eRNAs) observable in RNA-seq data, thus we queried intergenic regions marked by chromatin accessibility and H3K27ac for eRNAs using RNA-seq data on 768 myeloma specimens from the CoMMpass study. This identified transcription at 4,729 of 13,452 potential regions. eRNA expression was highly correlated with proximal gene expression. To illustrate this point, we performed t-SNE clustering based on mRNA and eRNA expression and color-coded each sample by myeloma gene expression subtype (Fig. E). Interestingly, this identified several regions highly correlated with oncogene expression between myeloma subtypes. For example, an enhancer ~154 kb upstream of CCND2 was uniquely transcribed in the MAF subtype (Fig. F) and this was highly correlated with CCND2 expression (Fig. G). Conclusions Cryopreservation of myeloma bone marrow aspirates allows isolation and analysis of biobanked samples that produce RNA-seq and ATAC-seq data that are highly congruent with those obtained from fresh samples and this represents a strategy for retrospective genomic and epigenomic studies. Chromatin accessibility analysis identified distinct enhancer elements regulating oncogenes in myeloma subtypes providing mechanistic insight into myeloma pathology. Figure 1 Disclosures Lin: Amgen: Current Employment, Current equity holder in publicly-traded company. Hofmeister:Bristol Myers Squibb: Honoraria, Research Funding; Janssen: Honoraria, Research Funding; Nektar: Honoraria, Research Funding; Sanofi: Honoraria, Research Funding; Oncopeptides: Honoraria; Oncolytics Biotech: Research Funding; Imbrium: Honoraria; Karyopharm: Honoraria, Research Funding. Nooka:Celgene: Consultancy, Honoraria, Research Funding; Sanofi: Consultancy, Honoraria; Adaptive Technologies: Consultancy, Honoraria; Spectrum Pharmaceuticals: Consultancy; Takeda: Consultancy, Honoraria, Research Funding; Janssen: Consultancy, Honoraria, Research Funding; Amgen: Consultancy, Honoraria, Research Funding; Bristol-Myers Squibb: Consultancy, Honoraria, Research Funding; GlaxoSmithKline: Consultancy, Honoraria, Other: Personal Fees: Travel/accomodations/expenses, Research Funding; Karyopharm Therapeutics, Adaptive technologies: Consultancy, Honoraria, Research Funding; Oncopeptides: Consultancy, Honoraria. Lonial:GSK: Consultancy, Honoraria, Other: Personal fees; BMS: Consultancy, Honoraria, Other: Personal fees, Research Funding; Takeda: Consultancy, Other: Personal fees, Research Funding; Novartis: Consultancy, Honoraria, Other: Personal fees; Janssen: Consultancy, Honoraria, Other: Personal fees, Research Funding; Merck: Consultancy, Honoraria, Other: Personal fees; JUNO Therapeutics: Consultancy; TG Therapeutics: Membership on an entity's Board of Directors or advisory committees; Millennium: Consultancy, Honoraria; Onyx: Honoraria; Genentech: Consultancy; Karyopharm: Consultancy; Amgen: Consultancy, Honoraria, Other: Personal fees; Sanofi: Consultancy; Abbvie: Consultancy. Boise:AstraZeneca: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Genetech: Membership on an entity's Board of Directors or advisory committees.

Background Clostridium saccharoperbutylacetonicum N1-4 (HMT) is a strictly anaerobic, spore-forming Gram-positive bacterium capable of hyper-butanol production through the well-known acetone–butanol–ethanol fermentation process. Recently, five putative RRNPP-type QSSs (here designated as QSS1 to QSS5) were predicted in this bacterial strain, each of which comprises a putative RRNPP-type regulator (QssR1 to QssR5) and a cognate signaling peptide precursor (QssP1 to QssP5). In addition, both proteins are encoded by the same operon. The functions of these multiple RRNPP-type QSSs are unknown. Results To elucidate the function of multiple RRNPP-type QSSs as related to cell metabolism and solvent production in N1-4 (HMT), we constructed qssR-deficient mutants ΔR1, ΔR2, ΔR3 and ΔR5 through gene deletion using CRISPR–Cas9 and N1-4-dcas9-R4 (with the QssR4 expression suppressed using CRISPR–dCas9). We also constructed complementation strains by overexpressing the corresponding regulator gene. Based on systematic characterization, results indicate that QSS1, QSS2, QSS3, and QSS5 positively regulate the sol operon expression and thus solvent production, but they likely negatively regulate cell motility. Consequently, QSS4 might not directly regulate solvent production, but positively affect cell migration. In addition, QSS3 and QSS5 appear to positively regulate sporulation efficiency. Conclusions Our study provides the first insights into the roles of multiple RRNPP-type QSSs of C. saccharoperbutylacetonicum for the regulation of solvent production, cell motility, and sporulation. Results of this study expand our knowledge of how multiple paralogous QSSs are involved in the regulation of essential bacterial metabolism pathways.

e21106 Background: Although immune checkpoint inhibitors (ICIs) have shown remarkable benefit for treatment of advanced non-small lung cancer (NSCLC), only a minority of patients can achieve durable responses and the majority produce an ultra-rapid progressive disease. Methods: Here, we collected the availably published datasets and mined the determinants of response to immunotherapy on pathway levels. 106 NSCLC patients treated with immunotherapy were combined from Rizvi et. al and Hellman et. al studies (whole exon sequencing) as the discovery dataset. Two independent validation datasets were comprised of the MSKCC cohort (targeted sequencing) and data by Anagnostou and colleagues (whole exon sequencing). The Cancer Genome Atlas (TCGA) somatic mutation and gene expression data were applied to explore the immunobiology features. Results: In the first combined cohort, we detected NOTCH pathway altered in 71% patients with durable clinical benefit (DCB) while only 36% among no durable benefit (NDB) (p = 0.005). Compared to NDB group, co-occurrence of NOTCH and at least two DDR (co-DDR) pathway was discovered in DCB group and contributed to a prolonged progression-free survival (PFS) [ 22.1 vs 3.6 months, p < 0.0001, HR, 0.34, 95% confidence interval (CI), 0.2-0.59]. In two independent datasets, co-occurrence of NOTCH+/co-DDR+ was also validated to be a better immunotherapy efficacy [ Cohort 2: 13 vs 6 months, p = 0.034, HR, 0.55, 95% CI, 0.31-0.96 ; Cohort 3: 21 vs 11 months, p = 0.067, HR, 0.45, 95% CI, 0.18-1.1 ]. By analyzing TCGA cohort, we found patients with coexisting NOTCH and co-DDR pathway had a higher TMB, more infiltration of CD4+T cells. Conclusions: Overall, co-occurrence of NOTCH and co-DDR pathway reflect a better immunotherapy efficacy in advanced NSCLC. This genomic predictor show promise in stratifying patients that suit for immunotherapy for future clinical practice.

Research on the realistic and comprehensive identification of citywide spatial patterns of economic segregation is valuable for the sustainable development of cities. The consideration of human activities in segregation research inspires us to develop an alternative method to contribute to this type of research. In our method, we emphasize the combination of collective activity spaces (CASs) and spatial economic data, both of which are obtained from dynamic human activities. We first reveal the realistic use of urban spaces from human mobility patterns to generate multilevel CASs as basic analytical units. Then, we use a type of realistic economic data generated from human activities to measure the segregation level of each CAS. We realize this measurement by tailoring a segregation index, named the Term Frequency-Inverse Document Frequency-Index of Concentration at the Extremes-based (TFIDF-ICE-based) segregation index, for our economic data. Through these methods, we can uncover citywide multilevel spatial patterns of economic segregation realistically and comprehensively. Using Beijing and Wuhan as cases, we demonstrate and discuss the applicability and value of our method.

Numerous urban indicators scale with urban population in a system of cities. However, whether the scaling law also exists in an intra-urban system and whether the scaling law across cities can be applied in the temporal growth of individual cities are not clear. Taking Shanghai, China, as an example, we collected urban population, building areas and other urban indicators from 2005 to 2017 in districts (the secondary administrative unit of Chinese cities). The building area has a robust scaling relationship with urban population among different districts in each year, indicating that the scaling law also exists in the intra-urban system composed by districts. The scaling exponent between building areas and population in the intra-urban system is less than one (sub-linear relationship), which means building areas increase more slowly than population across districts. Temporally, building areas increase faster than population in most individual districts, showing a super-linear relationship. The contrasting relationship between building area and population across districts and over time indicates that the scaling law cannot be applied in the temporal growth of individual districts or cities. In conclusion, the scaling law also exists in the intra-urban system, and the cross-sectional scaling law cannot be applied to individual parts over time. Our further analyses using GDP and other five urban indicators confirm the two findings.

The turning points of housing prices play a significant role in the real estate market and economy. However, because multiple factors impact the market, the prediction of the turning points of housing prices faces significant challenges. To solve this problem, in this study, a historical data-based model that incorporates a multi-population genetic algorithm with elitism into the log-periodic power law model is proposed. This model overcomes the weaknesses of multivariate and univariate methods that it does not require any external factors while achieving excellent interpretations. We applied the model to the case study collected from housing prices in Wuhan, China, from December 2016 to October 2018. To verify its reliability, we compared the results of the proposed model to those of the log-periodic power law model optimized by the standard genetic algorithm and simulated annealing, the results of which indicate that the proposed model performs best in terms of prediction. Efficiently predicting and analyzing the housing prices will help the government promulgate effective policies for regulating the real estate market and protect home buyers.

The emergence of social media provides a rich source of posts to facilitate situational awareness and management during crisis events. Since interactions within social media users are frequently used to establish social networks, previous studies have explored how social media contributes to the information diffusion about crisis events. However, most studies have concentrated on the dynamics of the online social network, whereas the information diffused within social space and physical world during different event evolution stages tends to be overlooked. To address this problem, this study introduces a methodology for modeling and analyzing the information diffusion of crisis events based on the retweet patterns of social media users at city level. The proposed methodology consists of two parts: 1) Fisher ordinal clustering method is applied to divide the evolution stages of crisis events based on social media user activities. 2) To explore the information diffusion of crisis events in the physical world, the social-physical network (SPnet) is constructed by extending the retweet network to the geographical space for each evolution stage. The Weibo data related to the Kunming terrorist attack in 2014 are analyzed to demonstrate the methodology. By identifying the geo-location of users (90.8% of the total active users), two types of SPnet are constructed based on the retweet patterns of ordinary users and media users, respectively. The results show that the information diffusion in the SPnet of ordinary users are mainly from Kunming, while in the SPnet of media users, major developed cities are the key nodes for information diffusion. This study provides useful insights for understanding how human activities react to crisis events through the social-physical network at city level.

The spatial structures of cities have changed dramatically with rapid socio-economic development in ways that are not well understood. To support urban structural analysis and rational planning, we propose a framework to identify urban functional regions and quantitatively explore the intensity of the interactions between them, thus increasing the understanding of urban structures. A method for the identification of functional regions via spatial semantics is proposed, which involves two steps: (1) the study area is classified into three types of functional regions using taxi origin/destination (O/D) flows; and (2) the spatial semantics for the three types of functional regions are demonstrated based on point-of-interest (POI) categories. To validate the existence of urban functional regions, we explored the intensity of interactions quantitatively between them. A case study using POI data and taxi trajectory data from Beijing validates the proposed framework. The results show that the proposed framework can be used to identify urban functional regions and promotes an enhanced understanding of urban structures.

A Bacillus amyloliquefaciens strain with enhanced γ-PGA production was constructed by metabolically engineering its γ-PGA synthesis-related metabolic networks: by-products synthesis, γ-PGA degradation, glutamate precursor synthesis, γ-PGA synthesis and autoinducer synthesis. The genes involved in by-products synthesis were firstly deleted from the starting NK-1 strain. The obtained NK-E7 strain with deletions of the epsA-O (responsible for extracellular polysaccharide synthesis), sac (responsible for levan synthesis), lps (responsible for lipopolysaccharide synthesis) and pta (encoding phosphotransacetylase) genes, showed increased γ-PGA purity and slight increase of γ-PGA titer from 3.8 to 4.15 g/L. The γ-PGA degrading genes pgdS (encoding poly-gamma-glutamate depolymerase) and cwlO (encoding cell wall hydrolase) were further deleted. The obtained NK-E10 strain showed further increased γ-PGA production from 4.15 to 9.18 g/L. The autoinducer AI-2 synthetase gene luxS was deleted in NK-E10 strain and the resulting NK-E11 strain showed comparable γ-PGA titer to NK-E10 (from 9.18 to 9.54 g/L). In addition, we overexpressed the pgsBCA genes (encoding γ-PGA synthetase) in NK-E11 strain; however, the overexpression of these genes led to a decrease in γ-PGA production. Finally, the rocG gene (encoding glutamate dehydrogenase) and the glnA gene (glutamine synthetase) were repressed by the expression of synthetic small regulatory RNAs in NK-E11 strain. The rocG-repressed NK-anti-rocG strain exhibited the highest γ-PGA titer (11.04 g/L), which was 2.91-fold higher than that of the NK-1 strain. Fed-batch cultivation of the NK-anti-rocG strain resulted in a final γ-PGA titer of 20.3g/L, which was 5.34-fold higher than that of the NK-1 strain in shaking flasks. This work is the first report of a systematically metabolic engineering approach that significantly enhanced γ-PGA production in a B. amyloliquefaciens strain. The engineering strategies explored here are also useful for engineering cell factories for the production of γ-PGA or of other valuable metabolites.

Background Poly-γ-glutamic acid (γ-PGA) is a valuable polymer with glutamate as its sole precursor. Enhancement of the intracellular glutamate synthesis is a very important strategy for the improvement of γ-PGA production, especially for those glutamate-independent γ-PGA producing strains. Corynebacterium glutamicum has long been used for industrial glutamate production and it exhibits some unique features for glutamate synthesis; therefore introduction of these metabolic characters into the γ-PGA producing strain might lead to increased intracellular glutamate availability, and thus ultimate γ-PGA production. Results In this study, the unique glutamate synthesis features from C. glutamicum was introduced into the glutamate-independent γ-PGA producing Bacillus amyloliquefaciens NK-1 strain. After introducing the energy-saving NADPH-dependent glutamate dehydrogenase (NADPH-GDH) pathway, the NK-1 (pHT315-gdh) strain showed slightly increase (by 9.1%) in γ-PGA production. Moreover, an optimized metabolic toggle switch for controlling the expression of ɑ-oxoglutarate dehydrogenase complex (ODHC) was introduced into the NK-1 strain, because it was previously shown that the ODHC in C. glutamicum was completely inhibited when glutamate was actively produced. The obtained NK-PO1 (pHT01-xylR) strain showed 66.2% higher γ-PGA production than the NK-1 strain. However, the further combination of these two strategies (introducing both NADPH-GDH pathway and the metabolic toggle switch) did not lead to further increase of γ-PGA production but rather the resultant γ-PGA production was even lower than that in the NK-1 strain. Conclusions We proposed new metabolic engineering strategies to improve the γ-PGA production in B. amyloliquefaciens. The NK-1 (pHT315-gdh) strain with the introduction of NADPH-GDH pathway showed 9.1% improvement in γ-PGA production. The NK-PO1 (pHT01-xylR) strain with the introduction of a metabolic toggle switch for controlling the expression of ODHC showed 66.2% higher γ-PGA production than the NK-1 strain. This work proposed a new strategy for improving the target product in microbial cell factories. Electronic supplementary material The online version of this article (doi:10.1186/s12934-017-0704-y) contains supplementary material, which is available to authorized users.

Background Sucrose is an naturally abundant and easily fermentable feedstock for various biochemical production processes. By now, several sucrose utilization pathways have been identified and characterized. Among them, the pathway consists of sucrose permease and sucrose phosphorylase is an energy-conserving sucrose utilization pathway because it consumes less ATP when comparing to other known pathways. Bacillus amyloliquefaciens NK-1 strain can use sucrose as the feedstock to produce poly-γ-glutamic acid (γ-PGA), a highly valuable biopolymer. The native sucrose utilization pathway in NK-1 strain consists of phosphoenolpyruvate-dependent phosphotransferase system and sucrose-6-P hydrolase and consumes more ATP than the energy-conserving sucrose utilization pathway. Results In this study, the native sucrose utilization pathway in NK-1 was firstly deleted and generated the B. amyloliquefaciens 3Δ strain. Then four combination of heterologous energy-conserving sucrose utilization pathways were constructed and introduced into the 3Δ strain. Results demonstrated that the combination of cscB (encodes sucrose permease) from Escherichia coli and sucP (encodes sucrose phosphorylase) from Bifidobacterium adolescentis showed the highest sucrose metabolic efficiency. The corresponding mutant consumed 49.4% more sucrose and produced 38.5% more γ-PGA than the NK-1 strain under the same fermentation conditions. Conclusions To our best knowledge, this is the first report concerning the enhancement of the target product production by introducing the heterologous energy-conserving sucrose utilization pathways. Such a strategy can be easily extended to other microorganism hosts for reinforced biochemical production using sucrose as substrate. Electronic supplementary material The online version of this article (doi:10.1186/s12934-017-0712-y) contains supplementary material, which is available to authorized users.

Additional file 5: Figure S2. Comparison of Îł-PGA production through fermentation with NK-1, NK-TP and NK-TP (pHT01-xylR) strains.

Additional file 2: Table S2. Genes sequences used in this article.

Additional file 1: Table S1. Primers used in this work.

Additional file 4: Figure S1. Verification of the function of metabolic toggle switch using the bgaB reporter gene.

Additional file 3: Table S2. Intracellular glutamate concentrations among different strains.

Additional file 1: Table S1. Primers used in this work.

Additional file 2. Genes sequences used in this article.

We constructed a metabolically engineered glutamate-independent Bacillus amyloliquefaciens strain with considerable γ-PGA production. It was carried out by double-deletion of the cwlO gene and epsA-O cluster, as well as insertion of the vgb gene in the bacteria chromosome. The final generated strain NK-PV elicited the highest production of γ-PGA (5.12 g l(-1)), which was 63.2% higher than that of the wild-type NK-1 strain (3.14 g l(-1)). The γ-PGA purity also improved in the NK-PV strain of 80.4% compared with 76.8% for the control. Experiments on bacterial biofilm formation experiment showed that NK-1 and NK-c (ΔcwlO) strains can form biofilm; the epsA-O deletion NK-7 and NK-PV strains could only form an incomplete biofilm.

Summary e-Poly-L-lysine (e-PL), showing a wide range of antimicrobial activity, is now industrially produced as a food additive by a fermentation process. A new strain capable of producing e-PL was isolated from a soil sample collected from Gutian, Fujian Province, China. Based on its morphological and biochemical features and phylogenetic similarity with 16S rRNA gene, the strain was identified as Streptomyces albulus and named NK660. The yield of e-PL in 30 l fed-batch fermentation with pH control was 4.2 g l−1 when using glycerol as the carbon source. The structure of e-PL was determined by nuclear magnetic resonance (NMR) and matrix-assisted laser desorption/ionization–time of flight mass spectrometry (MALDI-TOF MS). Previous studies have shown that the antimicrobial activity of e-PL is dependent on its molecular size. In this study, the polymerization degree of the e-PL produced by strain NK660 ranged from 19 to 33 L-lysine monomers, with the main component consisting of 24–30 L-lysine monomers, which implied that the e-PL might have higher antimicrobial activity. Furthermore, the e-PL synthetase gene (pls) was cloned from strain NK660 by genome walking. The pls gene with its native promoter was heterologously expressed in Streptomyces lividans ZX7, and the recombinant strain was capable of synthesizing e-PL. Here, we demonstrated for the first time heterologous expression of the pls gene in S. lividans. The heterologous expression of pls gene in S. lividans will open new avenues for elucidating the molecular mechanisms of e-PL synthesis.

Multiple myeloma (MM) is a commonly occurring hematologic malignancy in the United States with poor prognosis. Among all treatments, proteasome inhibitor (PI) based regimens have been a major breakthrough for patients' outcomes. Available PIs all target 20S proteasome core complex, and the duration of response is limited by toxicity and resistance development. Until now, the underlying mechanism of drug resistance remains unclear. The proteasome is the major proteolytic machinery in protein homeostasis which is pivotal for myeloma cell survival. A functional proteasome consists of 20S proteasome core particle with regulatory particle on one or both ends. There are 3 types of proteasome regulators that could activate a 20S proteasome, PA700 (19S), 11S REG (PA28) and PA200. The 11S REG (PA28) protein family consists of three members, α, β, and γ. PA28 α/β are IFN-γ inducible and with higher expression in antigen presenting cells. Currently, the function of 11S subunit remains largely unknown. Our analysis of plasma cells from MM patients and healthy donors has demonstrated that expression of 11S proteasome is higher in myeloma cells than normal plasma cells and progressively upregulated with disease progression. To further identify the function of 11S proteasome especially PA28α in MM, we generate PA28α knockdown stable MM cell lines. We have found that knockdown of PA28α inhibits MM cell growth and proliferation, also induces myeloma cell resistance to PIs. The mechanism of PI resistance is different from knocking down of 19S or 20S proteasome subunits. Silencing of PA28α inhibits proteasome activity and decreases proteasome work load concurrently, resulting in a favorable proteasome load vs capacity ratio. Altogether, in this report, we describe the function of PA28α in MM cells, also provide novel insights into regulating PIs sensitivity through modulation of the 11S proteasome subunit PA28α. Disclosures Hofmeister: Nektar: Honoraria, Membership on an entity's Board of Directors or advisory committees; Karyopharm: Membership on an entity's Board of Directors or advisory committees; Imbrium: Membership on an entity's Board of Directors or advisory committees; Celgene: Honoraria; Janssen: Membership on an entity's Board of Directors or advisory committees; Oncopeptides: Membership on an entity's Board of Directors or advisory committees. Kaufman:Karyopharm: Membership on an entity's Board of Directors or advisory committees; Pharmacyclics: Membership on an entity's Board of Directors or advisory committees; Janssen: Honoraria; Amgen: Consultancy; Bristol-Myers Squibb: Consultancy; Incyte: Consultancy; Celgene: Consultancy; Winship Cancer Institute of Emory University: Employment; AbbVie: Consultancy; Takeda: Consultancy; TG Therapeutics: Consultancy. Nooka:Amgen: Honoraria, Other: advisory board participation; GSK: Honoraria, Other: advisory board participation; Celgene: Honoraria, Other: advisory board participation; Takeda: Honoraria, Other: advisory board participation; Spectrum pharmaceuticals: Honoraria, Other: advisory board participation; BMS: Honoraria, Other: advisory board participation; Janssen: Honoraria, Other: advisory board participation; Adaptive technologies: Honoraria, Other: advisory board participation. Boise:Genentech Inc.: Membership on an entity's Board of Directors or advisory committees; AstraZeneca: Honoraria, Research Funding. Lonial:Takeda: Consultancy, Research Funding; Amgen: Consultancy; BMS: Consultancy; Janssen: Consultancy, Research Funding; GSK: Consultancy; Karyopharm: Consultancy; Genentech: Consultancy; Celgene Corporation: Consultancy, Research Funding.