Genome-wide screening using CRISPR coupled with nuclease Cas9 (CRISPR/Cas9) is a powerful technology for the systematic evaluation of gene function. Statistically principled analysis is needed for the accurate identification of gene hits and associated pathways. Here, we describe how to perform computational analysis of CRISPR screens using the MAGeCKFlute pipeline. MAGeCKFlute combines the MAGeCK and MAGeCK-VISPR algorithms and incorporates additional downstream analysis functionalities. MAGeCKFlute is distinguished from other currently available tools by being a comprehensive pipeline that contains a series of functions for analyzing CRISPR screen data. This protocol explains how to use MAGeCKFlute to perform quality control, normalization, batch effect removal, copy number bias correction, gene hit identification, and downstream functional enrichment analysis for CRISPR screens. We also describe gene identification and data analysis in CRISPR screens involving drug treatment. Completing the entire MAGeCKFlute pipeline requires approximately two hours on a desktop computer running Linux or Mac OS and with R support. The MAGeCKFlute package is available at http://www.bioconductor.org/packages/release/bioc/html/MAGeCKFlute.html.

The homeodomain leucine zipper (HD-Zip) genes encode transcription factors that have diverse functions in plant development and have often been implicated in stress adaptation. The HD-Zip genes are the most abundant group of homeobox (HB) genes in plants and do not occur in other eukaryotes. This paper describes the complete annotation of the HD-Zip families I, II and III from rice and compares these gene families with Arabidopsis in a phylogeny reconstruction. Orthologous pairs of rice and Arabidopsis HD-Zip genes were predicted based on neighbour joining and maximum parsimony (MP) trees with support of conserved intron-exon organization. Additionally, a number of HD-Zip genes appeared to be unique to rice. Searching of EST and cDNA databases and expression analysis using RT-PCR showed that 30 out of 31 predicted rice HD-Zip genes are expressed. Most HD-Zip genes were broadly expressed in mature plants and seedlings, but others showed more organ specific patterns. Like in Arabidopsis and other dicots, a subset of the rice HD-Zip I and II genes was found to be regulated by drought stress. We identified both drought-induced and drought-repressed HD-Zip genes and demonstrate that these genes are differentially regulated in drought-sensitive versus drought-tolerant rice cultivars. The drought-repressed HD-Zip family I gene, Oshox4, was selected for promoter-GUS analysis, showing that drought-responsiveness of Oshox4 is controlled by the promoter and that Oshox4 expression is predominantly vascular-specific. Loss-of-function analysis of Oshox4 revealed no specific phenotype, but overexpression analysis suggested a role for Oshox4 in elongation and maturation processes.

Abstract. C-reactive protein (CRP), the prototype marker of inflammation, and cardiac hypertrophy are important prognostic indicators in dialysis patients. Residual renal function (RRF) has also been shown to influence survival of peritoneal dialysis (PD) patients. This study examined the relations between inflammation, RRF, and left ventricular hypertrophy (LVH) and determined whether inflammation, RRF, and LVH combine adversely to predict the outcomes of PD patients. A prospective observational study was performed in 231 chronic PD patients. Left ventricular mass index (LVMi), residual glomerular filtration rate (GFR), CRP, hemoglobin, serum albumin, and BP were determined at study baseline and related to outcomes. On univariate analysis, age (P ϭ 0.002), dialysis duration (P ϭ 0.004), coronary artery disease (P Ͻ 0.001), pulse pressure (P Ͻ 0.001), hemoglobin (P Ͻ 0.001), serum albumin (P ϭ 0.032), log-CRP (P Ͻ 0.001), and GFR (P Ͻ 0.001) were significantly associated with log-LVMi. Log-CRP was positively correlated with pulse pressure (R ϭ 0.218, P ϭ 0.001) and negatively correlated with GFR (R ϭ Ϫ0.272, P Ͻ 0.001). Multivariate analysis showed that log-CRP (P ϭ 0.008) and RRF (P ϭ 0.003) remained associated with log-LVMi independent of hemoglobin, serum albumin, arterial pulse pressure, and coronary artery disease. After follow-up for 30 Ϯ 14 mo, 34.2% patients had died. CRP, RRF, and LVMi each were significantly predictive of all-cause mortality and cardiovascular death. Kaplan-Meier analysis showed a significant increase in all-cause (P Ͻ 0.0001) and cardiovascular mortality (P Ͻ 0.0001) as the number of risk factors, namely CRP Ն50th percentile, no RRF, and LVMiՆ 50th percentile increased with the 2-yr all-cause mortality and cardiovascular death reaching as high as 61% and 46%, respectively, for patients who had all three risk factors. Compared with patients with none of the three risk factors, those with all three risk factors had an adjusted hazards ratio of 6.94 (P Ͻ 0.001) and 5.43 (P ϭ 0.001) for all-cause mortality and cardiovascular mortality, respectively. In conclusion, inflammation, RRF, and LVH are interrelated and combine adversely to increase mortality and cardiovascular death risk of PD patients.Left ventricular hypertrophy (LVH) is a major cardiovascular complication and an important predictor for mortality in end stage renal disease (ESRD) patients (1,2). Other than hypertension, anemia, and hypoalbuminemia, well-known factors for LVH in ESRD (3), the worsening of LVH with decline in renal function in predialysis patients (4) as well as the regression of LVH after successful kidney transplantation (5) supports a link between renal function and LVH. Our group recently demonstrated a significant cross-sectional relationship between residual renal function (RRF) and LVH in nondiabetic ESRD patients who were receiving peritoneal dialysis (PD) therapy (6). The exact mechanisms for this association are unclear, but our data suggested that uremia, worsening anemia, and hypoalbuminemia with decl...

Background:Obesity, physical inactivity and reduced physical fitness may contribute to the rising burden of chronic diseases in China. We investigated these factors in China over a 14-year period using data from the randomized national surveys in 2000, 2005, 2010 and 2014. Methods:We conducted four national surveys among 151,656, 163,386, 154,931and 146,703 Chinese adults aged 20-59 in 2000, 2005, 2010 and 2014, respectively. Body mass index (BMI, kg/m 2 ) was used to evaluate underweight (BMI<18·5), overweight (BMI 23·0-27·5) and obesity (BMI≥27·5). Central obesity was defined as waist circumference >90 cm in men and >85 cm in women. Leisure-time physical activity (LTPA) was evaluated by whether or not the participants had completed the recommended minimum 150-min moderate or 75-min vigorous exercise per week. Indices for assessing physical fitness included forced vital capacity, resting heart rate, handgrip strength, sit and reach, and standing on one leg. Findings:The prevalence of obesity increased from 8·6% in 2000, 10·3% in 2005, and 12·2% in 2010 to 12·9% in 2014 (0·32% per year). Comparable estimates were 37·4%, 39·2%, 40·7% and 41·2% for overweight and 13·9%, 18·3%, 22·1% and 24·9% for central obesity. The corresponding upward trends per year were 0·27% and 0·78%, respectively. The prevalence of overweight, obesity and central obesity increased with age and was higher in men than women.. A simultaneous decrease was observed in the prevalence of underweight (5·4% in 2000 versus 4·6% in 2014, a downward trend of 0·06% per year). More participants met the minimum LTPA recommendation (17·2% in 2000 versus 22·8% in 2014), with the prevalence change per year being 0·33%, 0·50%, 0·37%, 0·06% for underweight, normal-weight, overweight, and obesity, respectively. Physical fitness deterioration was observed for all measures examined except resting heart rate. Interpretation:Despite increased participation in LTPA, we observed an upward trend in overweight/obesity and a decline in physical fitness in Chinese adults. Continued nationwide interventions are needed for promoting physical activity and other healthy lifestyles in China. Funding:National Physical Fitness Surveillance Center, Ministry of Science and Technology of the People's Republic of China. Introduction In recent decades, China has witnessed a rapid rise in the burden of noncommunicable diseases (NCDs). Despite differences in sampling methods and 3 diagnostic criteria, the estimated prevalence grew from 9.7% to 11.6% for type 2 diabetes, and from 15.5% to 50.1% for pre-diabetes between 2008 and 2010. 1 These diseases accounted for an estimated 80% of deaths and 70% of total disease burden in China. 2 While aging of the population is an important contributing factor, such health consequences are also likely attributed to the drastic changes in lifestyles following China's economic boom that result in obesity and physical inactivity. 3 These two factors, despite their strong correlation, have been independently linked to increased risk of ...