DIFFERENTIAL REGULATION OF THE MITOCHONDRIAL ADP ATP TRANSLOCASE GENEIN WOOD FROGS UNDER FREEZING STRESS/

The gene Aat coding for ADP/ATP translocase (AAT) was cloned from live r of the freeze-tolerant wood frog, Rana sylvatica, via differential s creening of a cDNA library from liver of frozen frogs and using probes from control versus frozen frogs. Sequence analysis showed that clone pBfFR07 bearing the AAT cDNA contained a 1318-bp insert with one full -length open reading frame. The deduced amino acid sequence included 3 17 residues, with 81-86% identities to mammalian AAT. A 1750-nt transc ript from the Aat gene was detected using pBfFR07 probe and a putative frog AAT of over 30 kDa was visualized by immunoblotting using a poly clonal antibody raised against chicken AAT. Analysis of liver samples from a time course of freezing showed a maximal 4.5-fold increase in m RNA after 8 h with AAT protein peaking in 24-h frozen frogs. Freezing also induced Aat expression in bladder and lung. In liver, mRNA expres sion also responded positively to anoxia stress but not to experimenta l dehydration of the animals. These results suggest that AAT induction during freezing may be stimulated by the ischemia that develops when plasma freezes; changes in AAT may contribute to stabilizing energetic s in mitochondrial versus cytosolic pools over freeze/thaw cycles. (C) 1997 Elsevier Science B.V.