Presentation on theme: "Figure S1: (A) Table shows number of genes that passed Welchs t-test at different q-values (FDR corrected p-values) and Fold Change cut-offs. (B) Immune."— Presentation transcript:
Figure S1: (A) Table shows number of genes that passed Welchs t-test at different q-values (FDR corrected p-values) and Fold Change cut-offs. (B) Immune system process was identified as the most enriched biological process upon functional enrichment using hypergeometric distribution test with FDR correction (q <0.05), and Fisher Exact Test (p < 0.05) on 1396 RASP regulated transcripts. The immune system process was associated with 177 of 1396 transcripts. Line graph of the177 immune response genes across groups of pre- and post-RASP leukocytes that were also exposed to SEB toxin. Among the 177 transcripts, 26 (red lines) were up-regulated and 151 (blue lines) were down- regulated in post-RASP leukocytes. log fold change RASP + + SEB + + A B
Figure S2: Expression changes of genes important for: (A ) pattern recognition receptors, inflammatory response, chemokinesis and activation of myeloid leukocytes; (B) antigen presentation, T-, B- & NK-cells activations; and (C) Transcription regulations. We used two different total RNA isolation methods and different microarray platforms: Trizol RNA isolation and cDNA microarrays (upper panels - purple bar) ; PAXgene RNA isolation and oligonucleotide arrays (lower panels - blue bar). A) B) C)
Figure S3: Real time PCR array analyses of transcripts involved in chemotaxis, inflammation, and antigen preparation and presentation pathways. Control (pre-RASP); week 5 (about mid-way of the RASP program); week 8 (post-RASP)
A) B) C) D) E) Figure S4: Comparisons of fold changes of transcript levels determined by real time PCR array and cDNA and oligonucleotide microarrays. Shown here are genes significantly associated with: (A) pattern recognition receptors; (B) inflammatory response [to scale the graph, fold changes of -15.2 and -23.8, labeled * and **, respectively, were assigned a values of ~5 and 6, respectively]; (C) antigen preparation and presentation (*fold change: -12.3; assigned value ~ -5 for scaling the graph); (D) transcription factors (* fold change: -12.6; ** fold change: - 12.3; *** fold change: -14; these were adjusted to around -5 for scaling the graph); (E) T-cell activation, differentiation and proliferations. Expression profiles of genes shown in panels A-E were assayed using SABiosciences RT² Profiler (PAHS 406 and PHAS 25) PCR Arrays, cDNA microarrays, and oligonucleotide microarrays. Total RNA samples were isolated using Trizol ® reagents for cDNA microarray analysis, and total RNA samples used for PCR and oligonucleotide arrays were isolated from blood samples collected in PAXgene ® tubes. (Note: Study subject for the PCR arrays were the 10 soldiers who started and completed the RASP).
RASP + + SEB + + Figure S5: Expression pattern of RASP suppressed 151 immune response genes in pre- and post-RASP leukocytes which were also exposed to SEB toxin. These is the same data as that of Figure 4A, without linear transformation against pre-RASP control groups
Figure S6: (A) Regulatory interaction among stress regulated microRNAs (miRs), important transcription factors (NFkB1, NR3C1, SATB1), inflammatory cytokines and antigen presenting molecules; (B) Seven stress-suppressed miRs targeting 48 mRNAs among 288 mRNAs that passed q < 0.001 and 1.5 fold change. Enriched pathways include IL-17A and IL-8 signaling, and NFkB activation pathways. A B fold change
Figure S7: Transcription factors with absolute z-score > 3.0, and their regulatory targets. Targets were identified among stress altered transcripts (288 transcripts, Fig. 1A) that passed Welchs t-test with FDR correction (q < 0.001) and 1.5 fold change cut off. The left panel shows pathways associated with these genes. fold change
Figure S8: Transcription factors targeting transcripts which were differentially regulated among RT-PCR assayed transcripts. Both MYC and NR3C1 were predicted to be activated (according to prediction z-score values, z-score > 2.5). Top functions associated with these targets were apoptosis of leukocytes, hematopoiesis, proliferation of blood cells, and immune response. Top pathways are shown in the table (left panel). fold change
Figure S9: Canonical pathways significantly associated with RASP (battlefield-like stress ) regulated genes that passed Welchs t-test and FDR correction (p <= 0.001) and 1.5 Fold change. Numbers on the right side indicate total # of genes in the pathway.