Novel Promoters for Human NAIP infected cells [26,27,28]; a role consistent with its inclusion in the Nod-Like Receptor (NLR) superfamily of cytosolic pattern recognition sensors [29]. Here the flexibility associated with NAIP regulation in human is further demonstrated, by showing that 59 truncated transcripts arise from two unique Alu SINEs. The resulting ORF is translated in a number of cell lines and primary tissues, and yields a protein possessing only the signature NLR domains. Since Alus are over- represented in gene-rich regions and present transcription factor binding motifs, their role in establishing transcriptional networks is of great interest, as previously suggested [13,30]. These findings indicate, for the first time, that Alu insertions can serve directly as gene promoters and derive novel transcripts and protein isoforms. The existence of NAIP protein isoforms, as described here, should therefore be considered in future experiments addressing its IAP and/or NLR functions. Results Human NAIP is a multicopy gene Copy number variation (CNV) exists in the region of human chromosome 5q13.2 encoding NAIP and other genes [31,32,33], as it does among inbred mouse strains [25]. In the reference human genome at least five copies are annotated [34] (Figure 1a), and while only one of these is full length, NAIPfull, the others are assumed to be pseudogenes since two are 59- and two are 39- deleted, NAIP1 & 2 and YNAIP1 & 2, respectively (Figure 1a, b). Exon content of the NAIP paralogues was verified using dot plots (Figure S1). While assessing their transcription using a variety of RT-PCR primers sets, we found that 39 transcript levels of NAIP are greater than 59 transcript levels in most tissues. In general, NAIP 59 and 39 transcripts showed the smallest differences in the macrophage-rich lung, spleen (Figure 1c), and blood (Figure S2). Figure 1. Expression of predicted NAIP copies in the sequenced human genome. A) General landscape of chromosome 5q13.2, including the NAIP (black arrows), GUSBP1 (grey arrows), and surrounding genes (white arrows). B) Exon architecture of the annotated NAIP copies, verified by dot plots (Figure S1). Slanted lines delimit deletions relative to NAIPfull. Diagrams are not drawn to scale. C) qRT-PCR with primers indicated by small arrowheads in panel B to determine the overall levels of NAIP 59 (light bars) vs 39 (dark bars) transcription. Values are normalized to b-actin levels in each tissue, and shown relative to kidney 59. Each bar represents the mean of at least five independent experiments 6 SD. doi:10.1371/journal.pone.0005761.g001 PLoS ONE | www.plosone.org 2 June 2009 | Volume 4 | Issue 6 | e5761