Programmable gene regulation for metabolic engineering using decoy transcription factor binding sites.

Transcription factor decoy binding sites are short DNA sequences that can titrate a transcription factor away from its natural binding site, therefore regulating gene expression. In this study, we harness synthetic transcription factor decoy systems to regulate gene expression for metabolic pathways in Escherichia coli. We show that transcription factor decoys can effectively regulate expression of native and heterologous genes. Tunability of the decoy can be engineered via changes in copy number or modifications to the DNA decoy site sequence. Using arginine biosynthesis as a showcase, we observed a 16-fold increase in arginine production when we introduced the decoy system to steer metabolic flux towards increased arginine biosynthesis, with negligible growth differences compared to the wild type strain. The decoy-based production strain retains high genetic integrity; in contrast to a gene knock-out approach where mutations were common, we detected no mutations in the production system using the decoy-based strain. We further show that transcription factor decoys are amenable to multiplexed library screening by demonstrating enhanced tolerance to pinene with a combinatorial decoy library. Our study shows that transcription factor decoy binding sites are a powerful and compact tool for metabolic engineering.

Sparse Space Shift Keying Modulation with Enhanced Constellation Mapping.

For reducing the switching frequency between the radio frequency (RF) chain and transmit antennas, a class of new sparse space shift keying modulation (SSSK) schemes are presented. This new class is proposed to simplify hardware implementation, through carefully designing the spatial constellation mapping pattern. Specifically, different from traditional space shift keying modulation (SSK), the proposed SSSK scheme utilizes more time slots to construct a joint design of time and spatial domain SSK modulation, while maintaining the special structure of single RF chain. Since part of the multi-dimension constellations of SSSK concentrate the energy in less time slots, the RF-switching frequency is effectively reduced due to the sparsity introduced in the time domain. Furthermore, through theoretical analysis, we obtain the closed-form expression of the bit error probability for the SSSK scheme, and demonstrate that slight performance gain can be achieved compared to traditional SSK with reduced implementation cost. Moreover, we integrate transmit antenna selection (TAS) to achieve considerable performance gain. Finally, simulation results confirm the effectiveness of the proposed SSSK scheme compared to its traditional counterpart.

Salicylate Increases Fitness Cost Associated with MarA-Mediated Antibiotic Resistance.

Antibiotic resistance is generally associated with a fitness deficit resulting from the burden of producing and maintaining resistance machinery. This additional cost suggests that resistant bacteria will be outcompeted by susceptible bacteria in conditions without antibiotics. However, in practice, this process is slow in part because of regulation that minimizes expression of these genes in the absence of antibiotics. This suggests that if it were possible to turn on their expression, the cost would increase, thereby accelerating removal of resistant strains. Experimental and theoretical studies have shown that environmental chemicals can change the fitness cost associated with resistance and therefore have a significant impact on population dynamics. The multiple antibiotic resistance activator (MarA) is a clinically important regulator in Escherichia coli that activates downstream genes to increase resistance against multiple classes of antibiotics. Salicylate is an inducer of MarA that can be found in the environment and derepresses marA's expression. In this study, we sought to unravel the interplay between salicylate and the fitness cost of MarA-mediated antibiotic resistance. Using salicylate as an inducer of MarA, we found that a wide spectrum of concentrations can increase burden in resistant strains compared to susceptible strains. Induction resulted in rapid exclusion of resistant bacteria from mixed populations of antibiotic-resistant and susceptible cells. A mathematical model captures the process and predicts its effect in various environmental conditions. Our work provides a quantitative understanding of salicylate exposure on the fitness of different MarA variants and suggests that salicylate can lead to selection against MarA-mediated resistant strains. More generally, our findings show that natural inducers may serve to bias population membership and could impact antibiotic resistance and other important phenotypes.

C-band wavelength tunable mode-locking fiber laser based on CD-SMS structure.

A wavelength-tuning passively mode-locked all-fiber laser based on cascaded dual single mode fiber-graded index multimode fiber-single mode fiber (CD-SMS) structure is proposed in this paper. The CD-SMS structure geometry, where two SMSs were fused together, and each of SMS was coined into the paddles of polarization controller (PC). By adjusting the orientation angle of the paddles, magnitude of the nonlinear phase shifts of the light can be controlled to realize mode-locking and filtering. Experimentally, mode locking was obtained based on the SMS structure with pulse duration of 930 fs at the central wavelength 1568 nm and FWHM bandwidth of 2.9 nm. The fundamental repetition rate obtained was 18.22 MHz. By inserting another SMS into the ring cavity, and make-up CD-SMS, the five main peaks of mode-locked pulses were tuned in the range of 1533-1573 nm was achieved by mechanically tuning the orientation angles of paddles of two PCs, the wavelength tuning range covers the whole C-band, reaching 40 nm. However, the tunable range is limited by gain bandwidth of erbium ion. The wavelength-tuning mode locked fiber laser based on CD-SMS provides a way to a wide range of tunable ultrafast lasers.

Bidirectional control of absence seizures by the basal ganglia: a computational evidence.

Absence epilepsy is believed to be associated with the abnormal interactions between the cerebral cortex and thalamus. Besides the direct coupling, anatomical evidence indicates that the cerebral cortex and thalamus also communicate indirectly through an important intermediate bridge-basal ganglia. It has been thus postulated that the basal ganglia might play key roles in the modulation of absence seizures, but the relevant biophysical mechanisms are still not completely established. Using a biophysically based model, we demonstrate here that the typical absence seizure activities can be controlled and modulated by the direct GABAergic projections from the substantia nigra pars reticulata (SNr) to either the thalamic reticular nucleus (TRN) or the specific relay nuclei (SRN) of thalamus, through different biophysical mechanisms. Under certain conditions, these two types of seizure control are observed to coexist in the same network. More importantly, due to the competition between the inhibitory SNr-TRN and SNr-SRN pathways, we find that both decreasing and increasing the activation of SNr neurons from the normal level may considerably suppress the generation of spike-and-slow wave discharges in the coexistence region. Overall, these results highlight the bidirectional functional roles of basal ganglia in controlling and modulating absence seizures, and might provide novel insights into the therapeutic treatments of this brain disorder.

Prevalence and causes of corneal blindness.

BACKGROUND: The study aimed to assess the prevalence and causes of corneal blindness in a rural northern Chinese population. DESIGN: Cross-sectional study. PARTICIPANTS OR SAMPLES: The cluster random sampling method was used to select the sample. METHODS: This population-based study included 11 787 participants of all ages in rural Heilongjiang Province, China. These participants underwent a detailed interview and eye examination that included the measurement of visual acuity, slit-lamp biomicroscopy and direct ophthalmoscopy. An eye was considered to have corneal blindness if the visual acuity was <9/18 because of corneal diseases. MAIN OUTCOME MEASURES: The main outcome measure was prevalence rates of corneal blindness and low vision. RESULTS: Among the 10 384 people enrolled in the study, the prevalence of corneal blindness is 0.3% (95% confidence interval 0.2-0.4%). The leading cause was keratitis in childhood (40.0%), followed by ocular trauma (33.3%) and keratitis in adulthood (20.0%). Age and illiteracy were found to be associated with an increased prevalence of corneal blindness. CONCLUSION: Blindness because of corneal diseases in rural areas of Northern China is a significant public health problem that needs to be given more attention.

Prevalence of blindness and low vision: a study in the rural Heilongjiang Province of China.

BACKGROUND: The prevalence of blind individuals in the north of China is unknown. The study aimed to investigate the prevalence and causes of blindness and low vision in rural areas in Heilongjiang province of China in 2008-2009. DESIGN: Cross-sectional study. PARTICIPANTS OR SAMPLES: A cluster random sampling method was used to recruit participants of all ages in rural areas of Heilongjiang. METHODS: Trained professionals performed interviews and clinical examinations to measure visual acuity. The relationships between blindness or low vision and age, gender and education level were analysed. MAIN OUTCOME MEASURES: The main outcome measure was prevalence rates of bilateral blindness and bilateral low vision. RESULTS: Of the 11 787 subjects, 10 384 (88.1%) were surveyed. The overall age-adjusted prevalence rates were 0.7% (95% confidence interval: 0.5-0.8%) for bilateral blindness and 1.7% (95% confidence interval: 1.4-1.9%) for bilateral low vision. The prevalence rates of blindness and low vision were higher in the elderly and uneducated population (P < 0.05). The main causes for blindness and low vision were cataracts (44.1 and 46.0%, respectively) and refractive errors (17.7 and 42.5%, respectively). CONCLUSION: Blindness and low vision are highly prevalent among people with cataracts and refractive errors. Eye care planning must focus on treating the avoidable and curable forms of blindness.

[Screening and identification of a single chain antibody fragment].

OBJECTIVE: To screen and identify anti- Aflatoxin B1 single chain antibodies (scFv) from Tomlinson (I) library. METHODS: The phages absorbed on the ELISA plate were eluted by four methods including glycine buffer elution, trypsin elution, AFB1 competitive elution and competitive elution following trypsin treatment. The phage positive clones were transformed into Escherichia coli HB2151 and soluble scFv protein was expressed with the induction of Isopropyl beta-D-1-Thiogalavtopy ranoside (IPTG). The scFv was identified by ELISA and DNA sequence. RESULTS: Comparing the four methods, we found that the most efficient way to get positive clones was competitive elution following trypsin treatment. Obtainning two positive clones that could bind specifically with free AFB1, which their relative affinity were 0.4 microg/mL and 0.7 microg/mL, respectively. DNA sequencing results showed that the scFv belonged to human immunoglobulin variable region. CONCLUSION: The specific human scFv could be obtained with phage antibody library, and our method provided an alternative for screening recombinant antibody against anti-hapten.

Controlling and exploiting cell-to-cell variation in metabolic engineering.

Individual cells within a population can display diverse phenotypes due to differences in their local environment, genetic variation, and stochastic expression of genes. Understanding this cell-to-cell variation is important for metabolic engineering applications because variability can impact production. For instance, recent studies have shown that production can be highly heterogeneous among engineered cells, and strategies that manage this diversity improve yields of biosynthetic products. These results suggest the potential of controlling variation as a novel approach towards improving performance of engineered cells. In this review, we focus on identifying the origins of cell-to-cell variation in metabolic engineering applications and discuss recent developments on strategies that can be employed to diminish, accept, or even exploit cell-to-cell variation.

Bacterial persistence induced by salicylate via reactive oxygen species.

Persisters are phenotypic variants of regular cells that exist in a dormant state with low metabolic activity, allowing them to exhibit high tolerance to antibiotics. Despite increasing recognition of their role in chronic and recalcitrant infections, the mechanisms that induce persister formation are not fully understood. In this study, we find that salicylate can induce persister formation in Escherichia coli via generation of reactive oxygen species (ROS). Salicylate-induced ROS cause a decrease in the membrane potential, reduce metabolism and lead to an increase in persistence. These effects can be recovered by culturing cells in the presence of a ROS quencher or in an anaerobic environment. Our findings reveal that salicylate-induced oxidative stress can lead to persistence, suggesting that ROS, and their subsequent impact on membrane potential and metabolism, may play a broad role in persister formation.