Mammalian Glutaminase Gls2 Gene Encodes Two Functional Alternative Transcripts by a Surrogate Promoter Usage Mechanism

Glutaminase is expressed in most mammalian tissues and cancer cells, but the regulation of its expression is poorly understood. An essential step to accomplish this goal is the characterization of its species- and cell-specific isoenzyme pattern of expression. Our aim was to identify and characterize transcript variants of the mammalian glutaminase Gls2 gene.We demonstrate for the first time simultaneous expression of two transcript variants from the Gls2 gene in human, rat and mouse. A combination of RT-PCR, primer-extension analysis, bioinformatics, real-time PCR, in vitro transcription and translation and immunoblot analysis was applied to investigate GLS2 transcripts in mammalian tissues. Short (LGA) and long (GAB) transcript forms were isolated in brain and liver tissue of human, rat and mouse. The short LGA transcript arises by a combination of two mechanisms of transcriptional modulation: alternative transcription initiation and alternative promoter. The LGA variant contains both the transcription start site (TSS) and the alternative promoter in the first intron of the Gls2 gene. The full human LGA transcript has two in-frame ATGs in the first exon, which are missing in orthologous rat and mouse transcripts. In vitro transcription and translation of human LGA yielded two polypeptides of the predicted size, but only the canonical full-length protein displayed catalytic activity. Relative abundance of GAB and LGA transcripts showed marked variations depending on species and tissues analyzed.This is the first report demonstrating expression of alternative transcripts of the mammalian Gls2 gene. Transcriptional mechanisms giving rise to GLS2 variants and isolation of novel GLS2 transcripts in human, rat and mouse are presented. Results were also confirmed at the protein level, where catalytic activity was demonstrated for the human LGA protein. Relative abundance of GAB and LGA transcripts was species- and tissue-specific providing evidence of a differential regulation of GLS2 transcripts in mammals

Cost-Effectiveness of Intermittent Preventive Treatment of Malaria in Pregnancy in Southern Mozambique

BACKGROUND: Malaria in pregnancy is a public health problem for endemic countries. Economic evaluations of malaria preventive strategies in pregnancy are needed to guide health policies. METHODS AND FINDINGS: This analysis was carried out in the context of a trial of malaria intermittent preventive treatment in pregnancy with sulphadoxine-pyrimethamine (IPTp-SP), where both intervention groups received an insecticide treated net through the antenatal clinic (ANC) in Mozambique. The cost-effectiveness of IPTp-SP on maternal clinical malaria and neonatal survival was estimated. Correlation and threshold analyses were undertaken to assess the main factors affecting the economic outcomes and the cut-off values beyond which the intervention is no longer cost-effective. In 2007 US$, the incremental cost-effectiveness ratio (ICER) for maternal malaria was 41.46 US$ (95% CI 20.5, 96.7) per disability-adjusted life-year (DALY) averted. The ICER per DALY averted due to the reduction in neonatal mortality was 1.08 US$(95$ (95% CI 0.42, 3.21) per DALY averted. Efficacy was the main factor affecting the economic evaluation of IPTp-SP. The intervention remained cost-effective with an increase in drug cost per dose up to 11 times in the case of maternal malaria and 183 times in the case of neonatal mortality. CONCLUSIONS: IPTp-SP was highly cost-effective for both prevention of maternal malaria and reduction of neonatal mortality in Mozambique. These findings are likely to hold for other settings where IPTp-SP is implemented through ANC visits. The intervention remained cost-effective even with a significant increase in drug and other intervention costs. Improvements in the protective efficacy of the intervention would increase its cost-effectiveness. Provision of IPTp with a more effective, although more expensive drug than SP may still remain a cost-effective public health measure to prevent malaria in pregnancy. TRIAL REGISTRATION: ClinicalTrials.gov NCT00209781

Bidirectional promoters generate pervasive transcription in yeast

Genome-wide pervasive transcription has been reported in many eukaryotic organisms, revealing a highly interleaved transcriptome organization that involves hundreds of previously unknown non-coding RNAs. These recently identified transcripts either exist stably in cells (stable unannotated transcripts, SUTs) or are rapidly degraded by the RNA surveillance pathway (cryptic unstable transcripts, CUTs). One characteristic of pervasive transcription is the extensive overlap of SUTs and CUTs with previously annotated features, which prompts questions regarding how these transcripts are generated, and whether they exert function. Single-gene studies have shown that transcription of SUTs and CUTs can be functional, through mechanisms involving the generated RNAs or their generation itself. So far, a complete transcriptome architecture including SUTs and CUTs has not been described in any organism. Knowledge about the position and genome-wide arrangement of these transcripts will be instrumental in understanding their function. Here we provide a comprehensive analysis of these transcripts in the context of multiple conditions, a mutant of the exosome machinery and different strain backgrounds of Saccharomyces cerevisiae. We show that both SUTs and CUTs display distinct patterns of distribution at specific locations. Most of the newly identified transcripts initiate from nucleosome-free regions (NFRs) associated with the promoters of other transcripts (mostly protein-coding genes), or from NFRs at the 3' ends of protein-coding genes. Likewise, about half of all coding transcripts initiate from NFRs associated with promoters of other transcripts. These data change our view of how a genome is transcribed, indicating that bidirectionality is an inherent feature of promoters. Such an arrangement of divergent and overlapping transcripts may provide a mechanism for local spreading of regulatory signals-that is, coupling the transcriptional regulation of neighbouring genes by means of transcriptional interference or histone modification