Alternative mRNA splicing generates the two ribulosebisphosphate carboxylase/oxygenase activase polypeptides in spinach and Arabidopsis.
Sequence analysis of ribulosebisphosphate carboxylase/oxygenase (rubisco) activase cDNA and genomic clones isolated from spinach and Arabidopsis thaliana indicates that the two polypeptides of rubisco activase arise from alternative splicing of a common pre-mRNA. In spinach, two 5' splice sites are used in processing a single 137-nucleotide intron near the 3' end of the primary transcript. This intron was either removed completely or, alternatively, the first 22 nucleotides of the intervening sequence were retained in the mature rubisco activase mRNA. The 22-nucleotide auxiliary exon contains an in-frame ochre termination codon and leads to the synthesis of a 41-kilodalton polypeptide. Removal of the entire 137-nucleotide intervening sequence results in the synthesis of a larger 45-kilodalton polypeptide. Thus, alternative splicing of the spinach rubisco activase mRNA results in the synthesis of two polypeptides that are identical except for 37 additional amino acids at the C terminus of the 45-kilodalton polypeptide. This conclusion was confirmed by Cleveland peptide mapping and by N-terminal and C-terminal amino acid sequence analyses of both purified polypeptides. This method of producing the two rubisco activase polypeptides may be an evolutionarily conserved feature in higher plants because a nearly identical process occurs in the production of the two rubisco activase polypeptides in Arabidopsis. In Arabidopsis, an alternatively spliced intron resides at precisely the same position as the alternatively spliced intron in spinach and results in the synthesis of 44-kilodalton and 47-kilodalton rubisco activase polypeptides. In contrast to spinach, however, the retained portion of the intervening sequence does not contain an in-frame termination codon. Rather, a shift in reading frame leads to termination of translation of the smaller polypeptide within the coding region of the larger polypeptide.
Werneke
J M
JM
Agricultural Research Service, United States Department of Agriculture, Urbana, Illinois 61801.
Chatfield
J M
JM
Ogren
W L
WL
D000595:Amino Acid Sequence; D001483:Base Sequence; D003001:Cloning, Molecular; D004789:Enzyme Activation / Q000235:genetics*; D008969:Molecular Sequence Data; D010940:Plant Proteins / Q000737:chemistry / Q000235:genetics; D010944:Plants / Q000201:enzymology* / Q000235:genetics; D012326:RNA Splicing / Q000235:genetics*; D012333:RNA, Messenger / Q000378:metabolism*; D012273:Ribulose-Bisphosphate Carboxylase / Q000737:chemistry / Q000235:genetics*
D010940:Plant Proteins; D012333:RNA, Messenger; C054765:rca protein, plant; D012273:Ribulose-Bisphosphate Carboxylase