EndoU is a novel regulator of AICD during peripheral B cell selection

Poe, Jonathan C.; Kountikov, Evgueni I.; Lykken, Jacquelyn M.; Natarajan, Abirami; Marchuk, Douglas A.; Tedder, Thomas F.

The 93.3-98.3 Mbp region encodes 17 pseudogenes and 38 known genes, including 10 olfactory receptor genes not expressed in hematopoietic cells. Therefore, the differential expression of transcripts from these genes was compared using BCR-stimulated CD22 /[inbr] and CD22 /[B6] B cells that remained similarly viable for 18 h. A single gene was differentially expressed between mouse lines, with 4.4-fold higher (Fig. 3 A) . B cells from WT [B6] mice expressed EndoU at lower levels than B cells from WT 129 (WT [129] ) mice before and after BCR ligation ( Fig. 3 To ensure that EndoU protein levels correlated with gene overexpression in CD22 /[B6] B cells, an affinity-purified rabbit polyclonal Ab specific for recombinant EndoU was generated. Western blot analysis of whole cell lysates confirmed increased EndoU expression in both resting and BCRstimulated CD22 /[B6] B cells relative to CD22 /[inbr] B cells ( Fig. 3 C) . EndoU overexpression in CD22 /[B6] mice was B cell specific, as EndoU levels were modest in activated spleen T cells from CD22 /[B6] and CD22 /[inbr] mice. Remarkably, EndoU was also overexpressed in B cells from Ig Tg sHEL mice both before and after BCR ligation but was nearly undetectable in B cells from Ig Tg littermate mice ( Fig. 3 D) . Thus, B cell EndoU overexpression required both a homozygous B6:B6 EndoU locus in CD22 /[B6] and Ig Tg sHEL mice that correlated with their characteristic AICD phenotypes.