Freezing-induced genes in the wood frog (rana sylvatica):.
Cai, Q., and K. B. Storey.
Institute of Biochemistry and Department of Biology, Carleton
University, 1125 Colonel By Drive, Ottawa, Ontario, Canada K1S
5B6
APStracts 3:0379R, 1996.
Differential screening of a cDNA library produced from liver of the
freeze tolerant wood frog, Rana sylvatica, was used to search for
freezing-induced genes. Five freezing-responsive cDNA clones
representing different genes were isolated when about 80,000 plaques
of a cDNA library, prepared from liver of frozen frogs (24 h at
-2.5C), were screened with 32P-labeled total cDNA probes from control
(5C) versus freezing-exposed frogs. Two clones, pBfFR45 and pBfFR04
are reported here in detail and were found to be homologous with the
genes for the 97 \f "Symbol" and SYMBOL 103 \f "Symbol" subunits of
fibrinogen, respectively. The clone pBfFR45 carried a 2305 bp cDNA
sequence that was of bipartite structure, containing two open reading
frames (ORFs). The first ORF potentially encoded a 332 residue
polypeptide, covering a partial sequence of the N-terminal region of
the SYMBOL 97 \f "Symbol" chain. The second ORF encoded a 247 amino
acid sequence, covering the whole C-terminal region of the SYMBOL 97
\f "Symbol" chain; this was highly homologous to the FASORF of
chicken SYMBOL 97 \f "Symbol"-fibrinogen and the extended SYMBOL 97
\f "Symbol"-chain of the human protein. Under control (5C)
conditions, moderate levels of fibrinogen SYMBOL 97 \f
"Symbol" and SYMBOL 103 \f "Symbol" transcripts
were exclusively found in liver. When frogs were given survivable
freezing exposures, levels of these transcripts in liver were highly
induced. Transcription of these genes was also elevated in gut and
lung during freezing but mRNA levels in these tissues were lower than
in liver. A time course assay confirmed that the transcript levels of
both SYMBOL 97 \f "Symbol" and SYMBOL 103 \f
"Symbol" subunit genes were dramatically elevated within
the early hours of freezing and reached a maximum 3-fold increase
over control levels after 8 h of freezing exposure. Two other
physiological stresses, whole body dehydration and anoxia exposure,
mimic individual elements of freezing stress in wood frogs. Northern
blot hybridization analysis showed that the expression of both the
SYMBOL 97 \f "Symbol" and SYMBOL 103 \f "Symbol"
genes was also upregulated in response to dehydration in vivo (20 %
of total body water lost) but both were completely inhibited by
anoxia exposure.
Received 19 July 1996; accepted in final form 17 October 1996.
APS Manuscript Number R419-6.
Article publication pending Am. J. Physiol. (Regulatory Integrative
Comp. Physiology).
ISSN 1080-4757 Copyright 1996 The American Physiological Society.
Published in APStracts on 5 November 1996