Molecular genetic studies of acetyl-CoA carboxylase and 3-methylcrotonyl-CoA carboxylase in plants

Similar reverse-genetics approach is applied to investigate the function of two biotin-containing enzymes at the interface between catabolism and anabolism: acetyl-CoA carboxylase (ACCase) and 3-methylcrotonyl-CoA carboxylase (MCCase).;Plastidic ACCase catalyzes the first committed step of de novo fatty acid biosynthesis. Antisense Arabidopsis expressing antisense RNA of CAC1-A, which encodes the biotin carboxyl carrier protein (BCCP) of plastidic ACCase under the direction of CaMV 35S promoter, have been obtained. CAC1-A antisense plants with reduced BCCP possess different degrees of similar morphological changes. These include slow growth, smaller plant size, and crinkly and variegated-yellow vegetative and cauline leaves eventually leading to premature cell death. The severity of the morphological change is correlated with the magnitude of the reduction of BCCP protein. Reduced BCCP expression does not alter mRNA and protein abundance of the other three subunits of ACCase, nor does it change fatty acid composition, but it reduces fatty acid content in each CAC1-A antisense plant. Reduced BCCP expression also causes cellular and subcellular abnormalities and expression changes in genes involved in transcription and translation, anti-oxidation, pathogen and other stress responses, cell division control, and cellular structure.;MCCase is involved in mitochondrial Leu catabolism and several interconnected metabolic pathways: the mevalonate shunt and isoprenoid catabolism. Antisense Arabidopsis expressing antisense RNA of MCC-A, which encodes the biotin-containing subunit of MCCase under the direction of CaMV 35S promoter, have been generated. MCC-A antisense plants express various levels of MCC-A. In some lines, MCC-A mRNA, MCC-A protein, and MCCase activity are all below detection. No morphological changes have been observed in MCC-A antisense plants. Moreover, MCC-A antisense plants accumulate more 14C-propionyl-CoA, an intermediate metabolite of peroxisomal Leu catabolic pathway, than wild type plants, after feeding plants with 14C-Leu. Searching Arabidopsis cDNA library identifies putative branched-chain amino acid aminotransferase in mitochondria, peroxisomes, chloroplasts and cytoplasm, but not putative branched-chain 2-ketoacid dehydrogenase genes in peroxisomes. Expression patterns of genes of potential interest over time in the dark are investigated using Affymetrix microarray. Our study provides evidence supporting the existence of the peroxisomal Leu catabolic pathway, but the establishment of a detailed pathway needs more investigation

Output Remapping Technique for Soft-Error Rate Reduction in Critical Paths

It is expected that the soft error rate (SER) of combinational logic will increase significantly. Previous solutions to mitigate soft errors in combinational logic suffer from delay penalty or area/power overhead. In this paper, we proposed an output remapping technique to reduce SER of critical paths. Experimental results show up to about 20X increase in Qcritical. So the SER is reduced significantly. This method does not introduce any delay penalty. The area/power overhead is limited as well. The output remapping method is based on our novel glitch width model. The analysis shows that output remapping technique works well along with technology scaling

Computational Understanding of the Selectivities in Metalloenzymes

Metalloenzymes catalyze many different types of biological reactions with high efficiency and remarkable selectivity. The quantum chemical cluster approach and the combined quantum mechanics/molecular mechanics methods have proven very successful in the elucidation of the reaction mechanism and rationalization of selectivities in enzymes. In this review, recent progress in the computational understanding of various selectivities including chemoselectivity, regioselectivity, and stereoselectivity, in metalloenzymes, is discussed

Vitamin D supplementation is beneficial in improving the prognosis of patients with acute respiratory failure in the intensive care unit: a retrospective study based on the MIMIC-IV database

BackgroundVitamin D plays a critical role in the regulation of multiple physiological pathways. Vitamin D deficiency may be a risk factor for life-threatening clinical conditions. Several studies have found that vitamin D supplementation in critically ill patients improves prognosis. The purpose of this study was to determine the association between vitamin D and the prognosis of patients with acute respiratory failure (ARF).MethodsIn this retrospective cohort study, we collected clinical information of ARF patients from the Medical Information Mart for Intensive Care IV (MIMIC-IV) version 2.0 database. The outcome of this study was in-hospital mortality, intensive care unit (ICU) mortality. Patients were divided into the no-vitamin D and vitamin D groups according to whether they received supplementation or not. The correlation between vitamin D and outcome was examined using Kaplan–Meier (KM) survival curves, Cox proportional risk regression models and subgroup analyses. Propensity-score matching (PSM) was used to ensure the robustness of our findings.ResultsThe study finally included 7,994 patients with ARF, comprising 6,926 and 1,068 in the no-vitamin D and vitamin D groups, respectively. The Kaplan–Meier survival curve indicated a significant difference in survival probability between the two groups. After adjustment for a series of confounders, the multivariate Cox proportional hazards models showed that the hazard ratio (95% confidence interval) values for in-hospital and ICU mortality in the no-vitamin D group were 1.67 (1.45, 1.93) and 1.64 (1.36, 1.98), respectively. The results of propensity score-matched (PSM) analysis were consistent with the original population. In the subgroup analysis, Vitamin D supplementation was associated with lower in-hospital mortality in patients with higher clinical scores (SOFA score ≥ 8, OASIS ≥ 38).ConclusionOur study concluded that Vitamin D supplementation may reduce in-hospital and ICU mortality in patients with ARF in the ICU. There may be a beneficial effect on in-hospital mortality in patients with higher clinical scores. Additional randomized controlled trials are needed to follow up to confirm the relationship between vitamin D supplementation and ARF

Developing and applying a gene functional association network for anti-angiogenic kinase inhibitor activity assessment in an angiogenesis co-culture model

<p>Abstract</p> <p>Background</p> <p>Tumor angiogenesis is a highly regulated process involving intercellular communication as well as the interactions of multiple downstream signal transduction pathways. Disrupting one or even a few angiogenesis pathways is often insufficient to achieve sustained therapeutic benefits due to the complexity of angiogenesis. Targeting multiple angiogenic pathways has been increasingly recognized as a viable strategy. However, translation of the polypharmacology of a given compound to its antiangiogenic efficacy remains a major technical challenge. Developing a global functional association network among angiogenesis-related genes is much needed to facilitate holistic understanding of angiogenesis and to aid the development of more effective anti-angiogenesis therapeutics.</p> <p>Results</p> <p>We constructed a comprehensive gene functional association network or interactome by transcript profiling an in vitro angiogenesis model, in which human umbilical vein endothelial cells (HUVECs) formed capillary structures when co-cultured with normal human dermal fibroblasts (NHDFs). HUVEC competence and NHDF supportiveness of cord formation were found to be highly cell-passage dependent. An enrichment test of Biological Processes (BP) of differentially expressed genes (DEG) revealed that angiogenesis related BP categories significantly changed with cell passages. Built upon 2012 DEGs identified from two microarray studies, the resulting interactome captured 17226 functional gene associations and displayed characteristics of a scale-free network. The interactome includes the involvement of oncogenes and tumor suppressor genes in angiogenesis. We developed a network walking algorithm to extract connectivity information from the interactome and applied it to simulate the level of network perturbation by three multi-targeted anti-angiogenic kinase inhibitors. Simulated network perturbation correlated with observed anti-angiogenesis activity in a cord formation bioassay.</p> <p>Conclusion</p> <p>We established a comprehensive gene functional association network to model in vitro angiogenesis regulation. The present study provided a proof-of-concept pilot of applying network perturbation analysis to drug phenotypic activity assessment.</p

MicroRNAs in Human Pituitary Adenomas

MicroRNAs (miRNAs) are a class of recently identified noncoding RNAs that regulate gene expression at posttranscriptional level. Due to the large number of genes regulated by miRNAs, miRNAs play important roles in many cellular processes. Emerging evidence indicates that miRNAs are dysregulated in pituitary adenomas, a class of intracranial neoplasms which account for 10–15% of diagnosed brain tumors. Deregulated miRNAs and their targets contribute to pituitary adenomas progression and are associated with cell cycle control, apoptosis, invasion, and pharmacological treatment of pituitary adenomas. To provide an overview of miRNAs dysregulation and functions of these miRNAs in pituitary adenoma progression, we summarize the deregulated miRNAs and their targets to shed more light on their potential as therapeutic targets and novel biomarkers

Downregulation of CD147 expression alters cytoskeleton architecture and inhibits gelatinase production and SAPK pathway in human hepatocellular carcinoma cells

<p>Abstract</p> <p>Background</p> <p>CD147 plays a critical role in the invasive and metastatic activity of hepatocellular carcinoma (HCC) cells by stimulating the surrounding fibroblasts to express matrix metalloproteinases (MMPs). Tumor cells adhesion to extracellular matrix (ECM) proteins is the first step to the tumor metastasis. MMPs degrade the ECM to promote tumor metastasis. The aim of this study is to investigate the effects of small interfering RNA (siRNA) against CD147 (si-CD147) on hepatocellular carcinoma cells' (SMMC-7721) architecture and functions.</p> <p>Methods</p> <p>Flow cytometry and western blot assays were employed to detect the transfection efficiency of si-CD147. Confocal microscopy was used to determine the effects of si-CD147 on SMMC-7721 cells' cytoskeleton. Invasion assay, gelatin zymography and cell adhesion assay were employed to investigate the effects of si-CD147 on SMMC-7721 cells' invasion, gelatinase production and cell adhesive abilities. Western blot assay was utilized to detect the effects of si-CD147 on focal adhesion kinase (FAK), vinculiln and mitogen-activated protein kinase (MAPK) expression in SMMC-7721 cells.</p> <p>Results</p> <p>Downregulation of CD147 gene induced the alteration of SMMC-7721 cell cytoskeleton including actin, microtubule and vimentin filaments, and inhibited gelatinase production and expression, cells invasion, FAK and vinculin expression. si-CD147 also blocked SMMC-7721 cells adhesion to collagen IV and phosphorylation level of SAPK/JNKs. SAPK/JNKs inhibitor SP600125 inhibited gelatinase production and expression.</p> <p>Conclusion</p> <p>CD147 is required for normal tumor cell architecture and cell invasion. Downregulation of CD147 affects HCC cell structure and function. Moreover, the alteration of cell behavior may be related to SAPK/JNK Pathway. siRNA against CD147 may be a possible new approach for HCC gene therapy.</p