Cold acclimation-induced up-regulation of the ribosomal protein L7 gene in the freeze tolerant wood frog, Rana sylvatica

Natural freezing survival by the wood frog, Rana sylvatica, involves multiple organ-specific changes in gene expression. The present study used differential display PCR to find cold-responsive genes in wood frog skin. A cDNA was retrieved from skin that was in higher amounts in cold- versus warm-acclimated frogs. The cDNA was used to probe a wood frog liver cDNA library and retrieve a long sequence that, after the further application of 5′RACE, was shown to encode the full sequence of the ribosomal large subunit protein 7 (RPL7) (GenBank accession number AF175983). Wood frog RPL7 contained 246 amino acids and shared 90% identity with Xenopus laevis RPL7, 82-83% with chicken and zebrafish homologues, and 79% with mammalian RPL7. Multiple binding domains found in human RPL7 showed differing degrees of conservation in the frog protein. Transcript levels of rpl7 were elevated up to 4-fold in skin of cold-acclimated frogs as compared with warm-acclimated animals. Organ-specific responses by rpl7 transcripts also occurred when frogs were given survivable freezing exposures. Transcripts rose by 1.8-3.3 fold in brain and skeletal muscle during freezing but were unaffected in central organs such as liver and heart. Up-regulation of rpl7 also occurred in brain of anoxia-exposed frogs and RPL7 protein levels increased strongly in heart under both freezing and dehydration stresses. Cold- and freezing-responsive up-regulation of the rpl7 gene and RPL7 protein in selected organs suggests that targeted changes in selected ribosomal proteins may be an integral part of natural freeze tolerance