The cis -encoded BTH_s13 controls BTH_I1527/SLT gene expression by transcript degradation. (A) BTH_s13 is adjacent to a sRNA prediction from Sipht (red) in the IGR between BTH_I1526 and BTH_I1527/SLT (green). Probe intensities for BTH_s13 expression in B. thailandensis treated with the three conditions described in Figure 3A are depicted. The yellow arrowhead bars map the different BTH_s13 sequences obtained by RACE. The grey boxes labeled 5’ and 3’ denote the probe sites used in the Northern blots shown in C. (B) PCR products were generated by applying RACE to the same cDNA template using primers specific to BTH_s13 and SLT. (C) Total RNA (15 μg) was isolated from Bt CDC272 grown in host medium (RPMI 10% serum) or derived from the extracellular (THP1-EC) or intracellular bacterial fraction (THP1-IC) of THP-1 infected cells. The Northern blots were derived from a single Northern blot consecutively processed with probes to 5’ BTH_s13, 3’ region of the BTH_I1527/SLT gene, and the 5S rRNA as a loading control. A representative of three independent experiments is shown. (D) Total RNA was isolated from Bt CDC272 cultured in LB, LB+Kan or LB+Serum, and from the THP1-EC and THP1-IC fractions of THP-1 infected cells. The relative RNA levels of SLT are presented as fold change of SLT transcript detected with the 3’ primer set and were calculated as the transcript ratio between each treatment relative to growth in LB. The BTH_s13/SLT ratio is derived from the transcript levels detected with primer sets corresponding to the 5’/BTH_s13 and the 3’ region of SLT for each condition. The average and standard deviation from three independent experiments perfomed in duplicate are presented. The “*” denotes statistical significance (p<0.05) for SLT expression in B. thailandensis grown in LB + treatment compared to LB alone or for BTH_s13 expression compared to SLT.