Source of nicotinamide governs its metabolic fate in cultured cells, mice, and humans.

Metabolic routing of nicotinamide (NAM) to NAD+ or 1-methylnicotinamide (MeNAM) has impacts on human health and aging. NAM is imported by cells or liberated from NAD+. The fate of 2H4-NAM in cultured cells, mice, and humans was determined by stable isotope tracing. 2H4-NAM is an NAD+ precursor via the salvage pathway in cultured A549 cells and human PBMCs and in A549 cell xenografts and PBMCs from 2H4-NAM-dosed mice and humans, respectively. 2H4-NAM is a MeNAM precursor in A549 cell cultures and xenografts, but not isolated PBMCs. NAM released from NAD+ is a poor MeNAM precursor. Additional A549 cell tracer studies yielded further mechanistic insight. NAMPT activators promote NAD+ synthesis and consumption. Surprisingly, NAM liberated from NAD+ in NAMPT activator-treated A549 cells is also routed toward MeNAM production. Metabolic fate mapping of the dual NAM sources across the translational spectrum (cells, mice, humans) illuminates a key regulatory node governing NAD+ and MeNAM synthesis.

Inferring metabolic pathway activity levels from RNA-Seq data.

BACKGROUND: Assessing pathway activity levels is a plausible way to quantify metabolic differences between various conditions. This is usually inferred from microarray expression data. Wide availability of NGS technology has triggered a demand for bioinformatics tools capable of analyzing pathway activity directly from RNA-Seq data. In this paper we introduce XPathway, a set of tools that compares pathway activity analyzing mapping of contigs assembled from RNA-Seq reads to KEGG pathways. The XPathway analysis of pathway activity is based on expectation maximization and topological properties of pathway graphs. RESULTS: XPathway tools have been applied to RNA-Seq data from the marine bryozoan Bugula neritina with and without its symbiotic bacterium "Candidatus Endobugula sertula". We successfully identified several metabolic pathways with differential activity levels. The expression of enzymes from the identified pathways has been further validated through quantitative PCR (qPCR). CONCLUSIONS: Our results show that XPathway is able to detect and quantify the metabolic difference in two samples. The software is implemented in C, Python and shell scripting and is capable of running on Linux/Unix platforms. The source code and installation instructions are available at http://alan.cs.gsu.edu/NGS/?q=content/xpathway .

Host differentially expressed genes during association with its defensive endosymbiont.

Mutualism, a beneficial relationship between two species, often requires intimate interaction between the host and symbiont to establish and maintain the partnership. The colonial marine bryozoan Bugula neritina harbors an as yet uncultured endosymbiont, "Candidatus Endobugula sertula," throughout its life stages. The bacterial symbiont is the putative source of bioactive complex polyketide metabolites, the bryostatins, which chemically defend B. neritina larvae from predation. Despite the presence of "Ca. Endobugula sertula" in all life stages of the host, deterrent bryostatins appear to be concentrated in reproductive portions of the host colony, suggesting an interaction between the two partners to coordinate production and distribution of the metabolites within the colony. In this study, we identified host genes that were differentially expressed in control colonies and in colonies cured of the symbiont. Genes that code for products similar to glycosyl hydrolase family 9 and family 20 proteins, actin, and a Rho-GDP dissociation inhibitor were significantly downregulated (more than twice) in antibiotic-cured non-reproductive zooids compared to control symbiotic ones. Differential expression of these genes leads us to hypothesize that the host B. neritina may regulate the distribution of the symbiont within the colony via mechanisms of biofilm degradation and actin rearrangement, and consequently, influences bryostatin localization to bestow symbiont-associated protection to larvae developing in the reproductive zooids.