A chick skeletal-muscle alpha-actinin gene gives rise to two alternatively spliced isoforms which differ in the EF-hand Ca(2+)-binding domain.
A chick non-muscle alpha-actinin cDNA probe encoding the EF-hand region of molecule was used to screen a lambda gt10 chick brain cDNA library from 14-day embryos. A partial 2.1-kb alpha-actinin cDNA was isolated (8W cDNA) which encoded a protein identical to chick skeletal-muscle alpha-actinin, except in the C-terminal part of the first EF hand. In the variant, the 22 residues found in the skeletal-muscle isoform were replaced by a stretch of 26 unique residues. Analysis of the structure of the skeletal-muscle alpha-actinin gene showed that the region of divergence was encoded by two exons which are alternatively spliced. Quantitative reverse transcriptase/polymerase chain reaction (RT/PCR) was used to investigate the levels of the alpha-actinin transcripts in various tissues. The skeletal-muscle alpha-actinin variant was expressed at low levels in brain, liver and spleen, but could not be detected in skeletal muscle. Surprisingly, skeletal-muscle alpha-actinin mRNA was also expressed in brain, liver and spleen. The RT/PCR products were authenticated by using diagnostic restriction enzyme sites and by sequencing. The splice variant derived from the skeletal-muscle alpha-actinin gene was also detected in a variety of cDNA libraries from both adult and embryonic tissues by PCR. Although a transcript encoding this alpha-actinin splice variant is expressed in non-muscle tissues, neither of the two EF-hands would be predicted to be functional, making it unlikely to be a typical non-muscle isoform which are calcium-sensitive with respect to binding actin. The two vertebrate non-muscle alpha-actinins sequenced to date also have a spacer of five amino acids between the two EF hands, whereas in the variant, the spacer is just four residues in length. Further analysis will be required before this alpha-actinin isoform, which we refer to as SKv, can be classified as muscle or non-muscle alpha-actinin. We propose a new nomenclature to describe the various alpha-actinin genes and their transcripts.
Parr
T
T
Department of Biochemistry, University of Leicester, England.
Waites
G T
GT
Patel
B
B
Millake
D B
DB
Critchley
D R
DR
D000185:Actinin / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D017398:Alternative Splicing; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D001921:Brain / Q000502:physiology*; D002118:Calcium / Q000378:metabolism*; D002642:Chick Embryo; D002645:Chickens; D015342:DNA Probes; D005091:Exons; D014644:Genetic Variation; D006801:Humans; D008099:Liver / Q000502:physiology; D008969:Molecular Sequence Data; D009132:Muscles / Q000502:physiology*; D009838:Oligodeoxyribonucleotides; D016133:Polymerase Chain Reaction / Q000379:methods; D011487:Protein Conformation; D017386:Sequence Homology, Amino Acid; D013154:Spleen / Q000502:physiology; D014158:Transcription, Genetic
D015342:DNA Probes; D009838:Oligodeoxyribonucleotides; D000185:Actinin; D002118:Calcium