Abstract 3034: air (apoptosis-induced regulator) expression and its sequence localization in human genome
Authors: d'Avenia Morena , Lonoce Angelo , Guryev Victor , Rosati Alessandra , Storlazzi Clelia Tiziana , Macchia Gemma , Falco Antonia , Belisario Maria Antonietta , Rocchi Mariano , Turco Maria Caterina 
Published: June 2012

Abstract
The aim of this study was to characterize a newly identified transcript named air (AX538681) whose genomic sequence is missing in human genome draft. air was identified as a differentially expressed cDNA in TNFα-stimulated Jurkat cells, transfected with a IkBα-hyperexpressing vector, versus control cells (Turco et al, 2007). Indeed air expression appeared to be induced by a pro-apoptotic stimulus and repressed by NF-κB activity.
Air cDNA was isolated by differential screening analysis of an expression library from Jurkat cells; its sequence (3.4kb long, 79% A/T rich) didn't show any similarity with annotated sequences except the homology with an EST (U74659) from human neuroblastoma cells and a 98% identity with traces from chimpanzee WGS database. This evidence suggested that air could belong to an un-sequenced region, whose secondary structure results in cloning instability.
Air sequence is detectable by Southern blot and PCR, providing the evidence of its presence in human genome (in press). air genomic 3’ and 5’ flanking regions (each one ∼2.2kb long) were isolated by vectorette PCR and sequenced to allow the synthesis of a probe (∼8kb long) suitable to perform a FISH assay. Preliminary experiments, performed on a thyroid cancer cell line, showed that air localize on Chr. 22. Moreover, we found that air transcript is longer than 3.4kb, since air 5’ flanking region is detectable by RT-PCR on total RNA (unpublished).
Four main putative ORFs were identified by ORF-Finder tool, but the aminoacidic sequence of three of them did not show any homology with conserved structural motifs while the fourth, localized on the new isolated 5’ transcript end, showed a transmembrane-like motif. Anyway, we don't exclude that air could be a long ncRNA (but it is not the Air described by Nagano et al, 2008), in fact, it is rich in stop codons, it's strongly predisposed to form stem loop secondary structures and its expression coincides with pro-apoptotic events (submitted).
Air role in modulating cell apoptosis was investigated: air silencing in lymphoma, neuroblastoma and AML cells, treated with stressful stimuli, showed a significant (p&lt;0.01) reduction of caspase 3 activation, mitochondrial membrane depolarization, cytochrome c release and appearance of apoptotic cells. At the same time, transfection of human PBMC with an air-hyperexpressing construct resulted in more than 25% appearance of apoptotic cells in respect to controls (Turco et al, 2007). Finally we found that air is also induced in pancreas carcinoma cell lines by ROS-inducing compounds (as PEITC and DEM). Since PEITC inhibits NF-κB activation, air induction by this agent is consistent with the suppressive effect of NF-κB on air expression (in press). We also observed air down-modulation in pancreatic AsPc1 cells treated with CDDP.
These findings identify air as a novel potential tool for enhancing cancer response to therapeutics.
Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 3034. doi:10.1158/1538-7445.AM2011-3034