THE CD40 TRAF FAMILY MEMBER INTERACTING MOTIF CARRIES THE INFORMATIONTO RESCUE WEHI-231 CELLS FROM ANTI-IGM-INDUCED GROWTH ARREST

Engagement of the antigen receptor on WEHI 231 murine B lymphoma cells leads to growth arrest and induction of apoptosis. Concomitant signal ing through CD40 sustains proliferation and rescues the cells from apo ptosis. At the molecular level, CD40 has been shown to activate nuclea r factor kappa B (NF-kappa B) and stress-activated protein kinase (SAP K). The aim of our present study was to define the stretch of the CD40 cytoplasmic tail responsible For mediating these effects in WEHI 231 cells. Using recombinant retroviruses with the enhanced green fluoresc ent protein as selection marker we transduced WEHI 231 cells with chim eric molecules consisting of the extracellular and transmembrane regio n of human CD40 or rat CD4 and selected portions of the murine CD40 ta il. Chimeric molecules with cytoplasmic fragments encompassing the ''C D40 tumor necrosis factor-associated factor family member interacting motif'' (TIM) were able to sustain growth and to uphold NF-kappa B act ivity as efficiently as the whole intracellular region of CD40. While the potential of the motif relative to the whole cytoplasmic tail was independent of the heterologous part of the chimeras it was strongly i nfluenced by its distance to the membrane. Placing the 17-amino acid s tretch of the motif too close to the membrane, i.e, only two or four a mino acids apart, destroyed its capacity to mitigate the anti-IgM effe ct. Activation of SAPK through the chimeric molecules always correlate d with their ability to activate NF-kappa B activity and to rescue the cells from apoptosis induced by antigen receptor ligation. Our data i ndicate that CD40-TIM carries most if not all of the information neede d to deliver the signals responsible for sustaining growth in anti-IgM -stimulated WEHI 231 cells.