Trace Amine-Associated Receptors in Monocytes: A Constant Low-Level Expression Jenna K. Caines, Sherri L. Christian, Mark D. Berry Department of Biochemistry, Memorial University of Newfoundland, St. John’s NL Trace amine-associated receptors (TAARs) Is TAAR1 differentially expressed in response to Pro- and Are any TAARs differentially expressed between § G protein-coupled receptors anti-inflammatory stimuli? monocyte and macrophage lineages? § Established throughout the body in vertebrates Table 1: Datasets with n ≥ 3 examining human and mice macrophages Table 2: Datasets with n ≥ 3 containing several monocyte and § Humans have 6 functional isoforms: TAAR1, TAAR2, TAAR5, TAAR6, TAAR8, and TAAR9 treated with pro- and anti-inflammatory stimuli macrophage lineages from mice TAARs and the immune system Dataset Treatment Time TAAR1 p-value Dataset Cell type Number of lineages Treatment § Found in leukocyte populations1 GSE53986 Untreated (control) 0h examined INFγ 24h 0.8 GSE15907 Lung Macrophage 2 NA 1 § TAAR1 suspected of regulating immune response GSE60290 Untreated (control) 0h Peritoneal Macrophage 6 NA § Potential target for pharmacological treatment of immune disorders2 INFγ 18h 0.3 Spleen Macrophage NA GSE43075 Untreated (control) 0h Blood Monocyte 2 NA Hypothesis LPS 4h 0.45 Mesenteric LN monocyte 1 NA GSE121646 Untreated (control) 0h Oral Salmonella § TAAR1 will show differential expression upon activation and LPS 8h 0.7 Small intestine macrophage 2 Typhimurium(72 h) GSE19315 Untreated (control) 0h between leukocyte populations Intestine Macrophage 1 NA LPS 6h 0.85 Methods: microarray analysis Bone marrow Macrophage 1 NA Brain Macrophage (microglia) 1 NA § Collect published microarray data from NCBI GEO and Immgen § Neither LPS or INFγ has significant effect on TAAR1 Bone marrow Monocyte 2 NA Process to evaluate gene expression by microarray expression GSE37448 Lung Macrophage 2 NA Normal cell Treated cell Blood Monocyte 1 NA § TAAR1 expression is not activation-dependent Lymph node (skin draining) Cell samples Not significant: Monocyte 1 NA Not present in cells Fat (viseral) Macrophage 1 NA Peritoneal Macrophage 2 Gata6 knockout Present in both cells Cell culture Liver Macrophage 2 NA Significant: Relative TAAR1 expression in various monocyte and In normal cells only macrophage lineages generated by Immgen mRNA RNA isolation mRNA MF.BM In treated cells only MF.RP.Sp Table 3: P-values for differential expression of TAARs in monocyte Reverse transcription MF.Lu cDNA cDNA MF.103-11b+.Lu and macrophage lineages contrasted with lung macrophages and labeling MF.II-480hi.PC Green Fluorescent Red Fluorescent MF.II+480lo.PC probes MF.Microglia.CNS Gene TAAR1 TAAR2 TAAR5 TAAR8 probes MF.Thio5.II+480int.PC Dataset MF.Thio5.II-480int.PC MF.Thio5.II-480hi.PC GSE15907 0.3 0.05 0.25 0.3 MF.Thio5.II+480lo.PC Hybridize MF.103-11b+.SI cDNA probes to MF.11cloSer.SI GSE37448 0.85 0.8 0.3 0.25 Scan Mo.6C+II-.BM oligo sequences Mo.6C-II-.BM Mo.6C+II-.BI on microarray Mo.6C+II+.BI Mo.6C-II-.BI Mo.6C-II+.BI § Only TAAR2 shows significant difference between lineages Mo.6C-Iiint.BI Mo.6C+II-.LN § RMA normalize the data and extract a list of differentially GN.BM GN.BI GN.Arth.BM expressed genes using R studio GN.Arth.SynF GN.UrAc.PC Anexprs example of RMA normalizationexprs GN.Thio.PC Mo.6C+_II.BI.v2 14 Mo.6C+II-.BIv2 References: 14 MF.Lu.v2 12 MF.11c-11b+.Lu.v2 1 12 Gainetdinov et al. (2018). Trace Amines and Their Receptors. Pharmacological MF.480int.Lv.v2 10 MF.480hi.Lv.v2 10 8 MF.AT.v2 reviews, 70(3), 549–620. EO.AT.v2 8 6 GN.BI.v2 2 Eo.BL.v2 Christian, S. L., & Berry, M. D. (2018). Trace Amine-Associated Receptors as 4 6 2 4 Novel Therapeutic Targets for Immunomodulatory Disorders. Frontiers in MF.11C−11B+.LU_1.CEL MO.6+2+.SLN_1.CEL MF.11C−11B+.LU_1.CEL MO.6+2+.SLN_1.CEL § Examine for significant differential expression of TAARs § TAAR1 is present and shows a constant low-level expression pharmacology, 9, 680..