Annexin 5 mediates a peroxide-induced Ca(2+) influx in B cells.
Annexin 5 is a Ca(2+)-binding protein, the function of which is poorly understood. Structural and electrophysiological studies have shown that annexin 5 can mediate Ca(2+) fluxes across phospholipid membranes in vitro [1]. There is, however, no direct evidence for the existence of annexin 5 Ca(2+) channels in living cells. Here, we show that annexin 5 inserts into phospholipid vesicle membranes at neutral pH in the presence of peroxide. We then used targeted gene disruption to explore the role of annexin 5 in peroxide-induced Ca(2+) signaling in DT40 pre-B cells. DT40 clones lacking annexin 5 exhibited normal Ca(2+) responses to both thapsigargin and B-cell receptor stimulation, but lacked the sustained phase of the response to peroxide. This late phase was due to Ca(2+) influx from the extracellular space, demonstrating that annexin 5 mediates a peroxide-induced Ca(2+) influx. Thus, peroxide induces annexin 5 membrane insertion in vitro, and peroxide-induced Ca(2+) entry in vivo in DT40 cells requires annexin 5. Our results are consistent with a role for annexin 5 either as a Ca(2+) channel, or as a signaling intermediate in the peroxide-induced Ca(2+)-influx pathway.
Kubista
H
H
Department of Physiology, University College London, London, WC1E 6BT, UK.
Hawkins
T E
TE
Patel
D R
DR
Haigler
H T
HT
Moss
S E
SE
D000818:Animals; D017306:Annexin A2 / Q000235:genetics / Q000378:metabolism; D017304:Annexin A5 / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D001402:B-Lymphocytes / Q000187:drug effects / Q000378:metabolism*; D002118:Calcium / Q000819:agonists / Q000378:metabolism*; D002460:Cell Line; D002645:Chickens; D004591:Electrophoresis, Polyacrylamide Gel; D017353:Gene Deletion; D006851:Hydrochloric Acid / Q000494:pharmacology; D006861:Hydrogen Peroxide / Q000494:pharmacology; D006863:Hydrogen-Ion Concentration; D007075:Immunoglobulin M / Q000494:pharmacology; D019284:Thapsigargin / Q000494:pharmacology; D013997:Time Factors
D017306:Annexin A2; D017304:Annexin A5; D007075:Immunoglobulin M; D019284:Thapsigargin; D006861:Hydrogen Peroxide; D006851:Hydrochloric Acid; D002118:Calcium