4.1. Gene Expression Profiling and RNA Sequence Analyses

Public expression profiling datasets used in this study were generated by HG U133 Plus 2.0 gene chips from Affymetrix (High Wycombe, UK) and obtained from Gene Expression Omnibus (GEO, www.ncbi.nlm.nih.gov). We exploited dataset {"type":"entrez-geo","attrs":{"text":"GSE24759","term_id":"24759"}}GSE24759 for analyses of developing and mature myeloid cells including dendritic cells setting a cut-off for positive/negative gene activity at 7.0 [84]. We used the online tool GEOR which extracts differentially expressed genes of two selected sample groups, generating volcano-plots for visualization and lists of significantly expressed genes for subsequent analysis. In addition, we analyzed RNA-seq based gene expression data from the Human Protein Atlas (www.proteinatlas.org) [85]. Dataset {"type":"entrez-geo","attrs":{"text":"GSE89565","term_id":"89565"}}GSE89565 was analyzed for gene activities in BPDCN and AML patients and dataset {"type":"entrez-geo","attrs":{"text":"GSE16395","term_id":"16395"}}GSE16395 was used to examine normal Langerhans cells and aberrant LHCs [24,86]. Recently, we sequenced transcriptomes from 100 leukemia/lymphoma cell lines including MUTZ-3 [27]. This LL-100 dataset is available from the European Nucleotide Archive (ENA; www.ebi.ac.uk/ena) via PRJEB30312. Comparative expression analysis of MUTZ-3 and 22 control cell lines was processed, analyzed, and normalized as described [27]. Genes were filtered for ≥50 normalized expression, average values for the control cell lines calculated, and the log2 fold changes to MUTZ-3 determined. For visualization of expression values, we used the public R-package shinyngs. Chromatin immuno-precipitation (ChIP)-sequencing (seq) data were obtained from GEO-datasets {"type":"entrez-geo","attrs":{"text":"GSE32465","term_id":"32465"}}GSE32465 (CEBPB, GATA2, SPI1) and {"type":"entrez-geo","attrs":{"text":"GSE96274","term_id":"96274"}}GSE96274 (ETV6) and analyzed using associated online tools and the Integrative Genomics Viewer (www.software.broadinstitute.org), respectively.

Expression profiling data from DC-differentiated MUTZ-3 cells were obtained from ArrayExpress (www.ebi.ac.uk/arrayexpress) via E-MEXP-3787 [29]. Expression profiling data from untreated and siRNA-treated MUTZ-3 cells were generated at the Genome Analytics Facility (Helmholtz Centre for Infection Research, Braunschweig, Germany). These expression profiling data were generated using HG U133 Plus 2.0 gene chips (Affymetrix) and are available from ArrayExpress via E-MTAB-10277. After RMA-background correction and quantile normalization of the spot intensities, the profiling data were expressed as ratios of sample means and subsequently log2 transformed. Data processing was performed via R/Bioconductor using public limma and affy packages.