Abstract
Multiple AMP deaminase (AMP-D) isoforms have been found in vertebrates, and tissue-specific inherited deficiencies of AMP-D have been described in two unrelated clinical syndromes suggesting there may be more than one AMP-D gene in higher eukaryotes. Using a newly isolated cDNA cloned from an adult rat brain library and a previously reported cDNA cloned from adult rat skeletal muscle, two linked AMP-D genes have been identified in rat and man. ampd1 is expressed at high levels in skeletal muscle of the adult rat. ampd2 is the predominant gene expressed in non-muscle tissues and smooth muscle of the adult rat, and it is also the predominant gene expressed in embryonic muscle and undifferentiated myoblasts. Both genes are expressed in cardiac muscle of the adult rat. The peptides encoded by these two genes have distinct immunological properties. The conservation of nucleotide sequence and exon/intron boundaries in these two genes suggests they arose by duplication of a common primordial gene around 150 million years ago.
Highlights
Using a newly isolated cDNA cloned from an adult rat brain library and a previously reported cDNA cloned from adult rat skeletal muscle, two linked AMP deaminase (AMP-D) genes have been identified in rat and man, ampdl is expressed at high levels in skeletal muscle of the adult rat. ampd[2] is the predominant gene expressed in nonmuscle tissues and smooth muscle of the adult rat, and it is the predominant gene expressed in embryonic muscle and undifferentiated myoblasts
Cloning of a Second AMP-D cDNA-Previous RNase mapping experiments identified a 97-base segment of similar sequence in the 2.5 and 3.4-kb AMP-D transcripts expressed in adult skeletal muscle and undifferentiated myoblasts, respectively (5)
These observations suggest this cDNA is derived from an unprocessed transcript for a form of AMP-D that is related to but distinct from the transcript produced in adult rat skeletal muscle
Summary
Co~ve~~~o~~ Library Screens-Rat brain Xgtll cDNA hbraries were obtained from Dr P. The hbrsries described above were screened with the &lymerese chain reaction technique described by Jansen and Ledley (12) Specific for ampd[2] sequences obtained from the partial cDNA isolated by conventional library screening, and unrelated 20-mer oligonucl~tides CTGCA-3’) specific to 3’-coding sequences of the ampd[2] partial cDNA clone isolated by conventional library screening. Analytic Procedures-Genomic DNA was isolated from rat or human cell lines, restricted with endonucleases, electrophoresed in 0.7% agarose gels, and transferred to Nytran filters as described previously (8). AMP-D activity in different tissues or cell extracts was assayed as described previously (4). After multiple rounds of selection, clones of L6 myoblasts were obtained that exhibited a 7-fold increase in AMP-D activity
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