The multiphosphorylation domain of the phosphorylase kinase alpha M and alpha L subunits is a hotspot of differential mRNA processing and of molecular evolution.
We have cloned and sequenced human cDNAs encoding the complete phosphorylase kinase alpha subunit muscle isoform (alpha M). The predicted polypeptide is highly similar to the sequence known from rabbit muscle but lacks a major part of its multiphosphorylation domain, including the main phosphorylation site for cAMP-dependent protein kinase (PKA). Analysis of this region by reverse-transcribed polymerase chain reaction (RT-PCR) in several human and rabbit tissues demonstrates that it is subject to elaborate differential mRNA splicing. Amino acids 1012-1024 of the full-length rabbit sequence, including the major PKA phosphorylation site, and amino acids 1025-1041, which harbor at least one endogenous phosphorylation site, can be deleted from the predicted polypeptide individually or in combination. Molecules lacking one or both of these segments constitute a major part of the alpha M subunit population in many rabbit tissues and constitute the vast majority in all human tissues analyzed. Similar, tissue-dependent differential splicing events could be detected by RT-PCR in the human alpha subunit isoform from liver (alpha L). The expression of the differentially spliced alpha M subtypes differs markedly between corresponding human and rabbit tissues. Sequence divergence in this region is particularly high, not only between the muscle and liver isoforms, but also between alpha M sequences from four different animal species. Moreover, a duplication of the exon encoding the main PKA phosphorylation site was discovered in the mouse. Thus, the multiphosphorylation domain of the phosphorylase kinase alpha subunit isoforms is subject to pronounced structural variation not only between different tissues of one organism via differential splicing, but also in the course of evolution.
Wüllrich
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A
Institut für Physiologische Chemie, Ruhr-Universität Bochum, Federal Republic of Germany.
Hamacher
C
C
Schneider
A
A
Kilimann
M W
MW
D017398:Alternative Splicing; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D005075:Biological Evolution; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics; D005091:Exons; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D010750:Phosphoproteins / Q000235:genetics; D010764:Phosphorylase Kinase / Q000235:genetics*; D010766:Phosphorylation; D012323:RNA Processing, Post-Transcriptional; D012333:RNA, Messenger / Q000378:metabolism; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D013045:Species Specificity
D018076:DNA, Complementary; D010750:Phosphoproteins; D012333:RNA, Messenger; D010764:Phosphorylase Kinase