Transcriptomic analysis of B cells from patients with chronic lymphocytic leukemia
CD22 (also known as Siglec-2) belongs to the Siglec family of glycan-recognition proteins and negatively regulates B-cell receptor-mediated signaling. CD22 is down-regulated in the B cells of patients with chronic lymphocytic leukemia (CLL). However, the mechanism regulating the transcription of CD22 is incompletely understood. In this study, we defined the minimal promoter of the human CD22 gene and identified the transcription factors that bind to the CD22 promoter by proteomic analysis. This transcriptome dataset (by RNAseq of peripheral blood B cells from 6 patients with CLL) was used (1) to validate the positive correlation between the expression levels of CD22 transcript (assessed by RNAseq) and CD22 protein (assessed by flow cytometry), and (2) to test the correlations between the expression levels of CD22 transcript and those of transcription factors that bind to minimal CD22 promoter. Overall design: B cells were purified from donor blood with CD19 MicroBeads (Miltenyi Biotec). Total RNA was prepared from 5×10^6 cells with RNeasy Mini Kit (Qiagen) and subjected to library preparation with KAPA mRNA HyperPrep Kit (Roche). The next-generation sequencing (20 million paired-end (2 × 150 bp) sequencing per sample) with NovaSeq 6000 (Illumina) was performed. The raw data was mapped and normalized, and transcripts per million (TPM) were calculated for each gene by HISAT2.