Global energy demand and environmental concerns have stimulated increasing efforts to produce carbon-neutral fuels directly from renewable resources. Microbially derived aliphatic hydrocarbons, the petroleum-replica fuels, have emerged as promising alternatives to meet this goal. However, engineering metabolic pathways with high productivity and yield requires dynamic redistribution of cellular resources and optimal control of pathway expression. Here we report a genetically encoded metabolic switch that enables dynamic regulation of fatty acids (FA) biosynthesis in Escherichia coli. The engineered strains were able to dynamically compensate the critical enzymes involved in the supply and consumption of malonyl-CoA and efficiently redirect carbon flux toward FA biosynthesis. Implementation of this metabolic control resulted in an oscillatory malonyl-CoA pattern and a balanced metabolism between cell growth and product formation, yielding 15.7-and 2.1-fold improvement in FA titer compared with the wild-type strain and the strain carrying the uncontrolled metabolic pathway. This study provides a new paradigm in metabolic engineering to control and optimize metabolic pathways facilitating the high-yield production of other malonyl-CoA-derived compounds.biofuels | dynamic metabolic control | transcriptional regulation A grand challenge in synthetic biology is to move the design of biomolecular circuits from purely genetic constructs toward systems that integrate different levels of cellular complexity, including regulatory networks and metabolic pathways (1). Despite the fact that a large volume of regulatory architectures and motifs has been discovered (2, 3), little has been accomplished in pathway engineering to improve cellular productivity and yield by exploiting dynamic pathway regulation and metabolic control (4). One essential part in implementing synthetic metabolic control in pathway engineering is to engineer novel metabolite sensors with desired input-output relationships. For example, have designed and applied a regulatory circuit that can sense the glycolytic pathway hallmark metabolite acetylphosphate to control the lycopene biosynthetic pathway (5) and generate oscillatory gene expression (6) as well as achieve artificial cell-cell communication (7). Dahl et al. (8) have used stressresponse promoters to improve farnesyl pyrophosphate production, and Tsao et al. (9) have rewired the Escherichia coli native quorumsensing regulon for autonomous induction of recombinant proteins.Traditional metabolic engineering is largely focused on the overexpression of rate-limiting steps (10

Microbial factories have been engineered to produce lipids from carbohydrate feedstocks for production of biofuels and oleochemicals. However, even the best yields obtained to date are insufficient for commercial lipid production. To maximize the capture of electrons generated from substrate catabolism and thus increase substrate-to-product yields, we engineered 13 strains of Yarrowia lipolytica with synthetic pathways converting glycolytic NADH into the lipid biosynthetic precursors NADPH or acetyl-CoA. A quantitative model was established and identified the yield of the lipid pathway as a crucial determinant of overall process yield. The best engineered strain achieved a productivity of 1.2 g/L/h and a process yield of 0.27 g-fatty acid methyl esters/g-glucose, which constitutes a 25% improvement over previously engineered yeast strains. Oxygen requirements of our highest producer were reduced owing to decreased NADH oxidization by aerobic respiration. We show that redox engineering could enable commercialization of microbial carbohydrate-based lipid production.

No abstract

IMPORTANCE Thyroid cancer is the most pervasive endocrine cancer worldwide. Studies examining the association between thyroid cancer and country, sex, age, sociodemographic index (SDI), and other factors are lacking. OBJECTIVE To examine the thyroid cancer burden and variation trends at the global, regional, and national levels using data on sex, age, and SDI. DESIGN, SETTING, AND PARTICIPANTSIn this cross-sectional study, epidemiologic data were gathered using the Global Health Data Exchange query tool, covering persons of all ages with thyroid cancer in 195 countries and 21 regions from January 1, 1990, to December 31, 2017; data analysis was completed on October 1, 2019. All participants met the Global Burden of Disease Study inclusion criteria. MAIN OUTCOMES AND MEASURES Outcomes included incidence, deaths, and disability-adjusted life-years (DALYs) of thyroid cancer. Measures were stratified by sex, region, country, age, and SDI.The estimated annual percentage changes (EAPCs) and age-standardized rates were calculated to evaluate the temporal trends. RESULTSIncreases of thyroid cancer were noted in incident cases (169%), deaths (87%), and DALYs (75%). Age-standardized incidence rate (ASIR) showed an upward trend over time, with an EAPC of 1.59 (95% CI, 1.51-1.67); decreases were noted in EAPCs of age-standardized death rate (−0.15; 95% CI, −0.19 to −0.12) and age-standardized DALY rate (−0.11; 95% CI, −0.15 to −0.08). Almost half (41.73% for incidence, 50.92% for deaths, and 54.39% for DALYs) of the thyroid cancer burden was noted in Southern and Eastern Asia. In addition, females accounted for most of the thyroid cancer burden (70.22% for incidence, 58.39% for deaths, and 58.68% for DALYs) and increased by years in this population, although the ASIR of males with thyroid cancer (EAPC, 2.18; 95% CI, 2.07-2.28) increased faster than that of females (EAPC, 1.38; 95% CI, 1.30-1.46). A third (34%) of patients with thyroid cancer resided in countries with a high SDI, and most patients were aged 50 to 69 years, which was older than the age in other quintiles (high SDI quintile compared with all other quintiles, P<.05). The most common age at onset of thyroid cancer worldwide was 15 to 49 years in female individuals compared with 50 to 69 years in male individuals (P<.05). Death from thyroid cancer was concentrated in participants aged 70 years or older and increased by years (average annual percentage change, 0.10; 95% CI, 0.01-0.21; P<.05). Furthermore, people in lower SDI quintiles developed thyroid cancer and died from it earlier than those in other quintiles (high and high-middle SDI vs low and low-middle SDI, P<.05). CONCLUSIONS AND RELEVANCEData from this study suggest considerable heterogeneity in the epidemiologic patterns of thyroid cancer across sex, age, SDI, region, and country, providing Key Points Question What were the epidemiologic patterns and variation in the trends of thyroid cancer worldwide from 1990 to 2017? Findings In this cross-sectional study covering data on incidence, deaths, and disabi...

Metabolic engineering entails target modification of cell metabolism to maximize the production of a specific compound. For empowering combinatorial optimization in strain engineering, tools and algorithms are needed to efficiently sample the multidimensional gene expression space and locate the desirable overproduction phenotype. We addressed this challenge by employing design of experiment (DoE) models to quantitatively correlate gene expression with strain performance. By fractionally sampling the gene expression landscape, we statistically screened the dominant enzyme targets that determine metabolic pathway efficiency. An empirical quadratic regression model was subsequently used to identify the optimal gene expression patterns of the investigated pathway. As a proof of concept, our approach yielded the natural product violacein at 525.4 mg/L in shake flasks, a 3.2-fold increase from the baseline strain. Violacein production was further increased to 1.31 g/L in a controlled benchtop bioreactor. We found that formulating discretized gene expression levels into logarithmic variables (Linlog transformation) was essential for implementing this DoE-based optimization procedure. The reported methodology can aid multivariate combinatorial pathway engineering and may be generalized as a standard procedure for accelerating strain engineering and improving metabolic pathway efficiency.

Knee osteoarthritis is the most common degenerative disease of the joints caused by articular cartilage injury, degradation of the joint edge and subchondral bone hyperplasia. Various treatments are used to alleviate the symptoms of patients with knee osteoarthritis, including analgesics and intra-articular injections. Platelet-rich plasma (PRP) is an autologous and multifunctional platelet concentrate of the blood, which stimulates the cartilage healing process and improves the damage caused by articular disease. Hyaluronic acid (HA) is an effective treatment for patients with knee osteoarthritis. In the current study, the effectiveness of PRP and HA combination therapy administered via intra-articular injections for patients with knee osteoarthritis was analyzed. A total of 360 patients with knee osteoarthritis were randomized into four different treatment groups as follows: Double-blind treatment with PRP (2-14 ml); double-blind treatment with HA (0.1-0.3 mg); combination therapy of PRP and HA; and placebo groups. Following treatment, all patients were evaluated using the Western Ontario and McMaster Universities Arthritis Index (WOMAC) and Common Toxicity Criteria. The most common treatment-emergent adverse events were hypertension and proteinuria. The current study demonstrated that PRP and HA treatment significantly improved arthralgia, and PRP treatment was determined to be significantly more effective than HA treatment using the WOMAC pain score (P<0.05). PRP and HA combination treatment significantly improved arthralgia, reduced humoral and cellular immune responses and promoted angiogenesis, which improved the patients' histological parameters compared with PRP or HA treatment alone. These results suggested that PRP and HA combination treatment may be a potential treatment option for patients with knee osteoarthritis in the future.

Malonyl-CoA is the rate-limiting precursor involved in the chain elongation reaction of a range of value-added pharmaceuticals and biofuels. Development of malonyl-CoA responsive sensors holds great promise in overcoming critical pathway limitations and optimizing production titers and yields. By incorporating the Bacillus subtilis trans-regulatory protein FapR and the cis-regulatory element fapO, we constructed a hybrid promoter-regulatory system that responds to a broad range of intracellular malonyl-CoA concentrations (from 0.1 to 1.1 nmol/mgDW) in Escherichia coli. Elimination of regulatory protein and nonspecific DNA cross-communication leads to a sensor construct that exhibits malonyl-CoA-dependent linear phase kinetics with increased dynamic response range. The sensors reported in this study could potentially control and optimize carbon flux leading to robust biosynthetic pathways for the production of malonyl-CoA-derived compounds.

Objective. To investigate the differences in gene expression profiles of adult articular cartilage from patients with Kashin-Beck disease (KBD) versus those with primary knee osteoarthritis (OA).Methods. The messenger RNA expression profiles of articular cartilage from patients with KBD, diagnosed according to the clinical criteria for KBD in China, were compared with those of cartilage from patients with OA, diagnosed according to the Western Ontario and McMaster Universities OA Index. Total RNA was isolated separately from 4 pairs of the KBD and OA cartilage samples, and the expression profiles were evaluated by Agilent 4؋44k Whole Human Genome density oligonucleotide microarray analysis. The microarray data for selected transcripts were confirmed by quantitative real-time reverse transcriptionpolymerase chain reaction (RT-PCR) amplification.Results. For 1.2 ؋ 10 4 transcripts, corresponding to 58.4% of the expressed transcripts, 2-fold changes in differential expression were revealed. Expression levels higher in KBD than in OA samples were observed in a mean ؎ SD 6,439 ؎ 1,041 (14.6 ؎ 2.4%) of the transcripts, and expression levels were lower in KBD than in OA samples in 6,147 ؎ 1,222 (14.2 ؎ 2.8%) of the transcripts. After application of the selection criteria, 1.85% of the differentially expressed genes (P < 0.001 between groups) were detected. These included 233 genes, of which 195 (0.4%) were expressed at higher levels and 38 (0.08%) were expressed at lower levels in KBD than in OA cartilage. Comparisons of the quantitative RT-PCR data supported the validity of our microarray data.Conclusion. Differences between KBD and OA cartilage exhibited a similar pattern among all 4 of the pairs examined, indicating the presence of disease mechanisms, mainly chondrocyte matrix metabolism, cartilage degeneration, and apoptosis induction pathways, which contribute to cartilage destruction in KBD.