Activated B lymphocytes express L1 mRNA, but not L1 protein.

<p>(<b>A and B</b>) L1, IAP and MLV mRNA expression. After digestion with DNase, mRNA was reverse-transcribed into cDNA using oligo(dT) primers, followed by L1-, IAP- and MLV-specific quantitative real-time PCR. PCR data were quantified according to the comparative <i>C</i>_T method. The amount of target mRNA was normalized to the endogenous control 18S ribosomal RNA. Values represent mean ± SD; y-axis, fold change in mRNA expression. (A) LPS- plus IL-4-stimulated (LPS) and unstimulated B lymphocytes from the spleens of AID-deficient (AID KO; n = 12 mice) and wild-type (WT; n = 12 mice) BALB/c mice. Student's t test, *p≤0.05 and **p≤0.001. (B) Tissues (pancreas, testis) and activated B cells (spleen LPS) from a wild-type BALB/c mouse and mouse cell lines F9 and MOPC104E; F9, mouse embryonal carcinoma cell line expressing high levels of L1 mRNA; spleen LPS, LPS- plus IL-4-stimulated B lymphocytes, as in panel A; n = 2 independent experiments. (<b>C and D</b>) ORF1 protein expression. Cell lysates were electrophoresed, Western blotted and developed with anti-ORF1 or anti-actin antibody. The position of the molecular mass standard (in kDa) is indicated to the left of the blots; the positions of the exogenous (exo) and endogenous (endo) human (hORF1) and mouse (mORF1) ORF1 bands are indicated to the right of the blots; LPS blasts, lysates of LPS- plus IL-4-activated B lymphocytes from BALB/c mice; + AID and – AID, AID-sufficient and AID-deficient, respectively. (C) anti-mORF1 staining; testis, whole testis lysates; IR, cells gamma irradiated 24 h before lysis with 0, 1 or 50 Gy. (D) anti-hORF1 staining; 2102Ep, lysate from a human embryonal carcinoma cell line expressing high levels of ORF1; Phoenix and LPS blasts, lysates of cells transfected (Phoenix) or transduced (LPS Blasts) with a retroviral human ORF1 (+ exo hORF1) or a GFP-only construct (− exo hORF1).