Molecular characterization of ribulose-1,5-bisphosphate carboxylase/oxygenase activase in rice leaves

Abstract

A full-length cDNA (designated rcaII) encoding the Rubisco activase (RCA) of rice (Oryza sativa L.) has been cloned from a cDNA library constructed with mRNA from green leaves. Sequence analysis resulted in a reading frame of 432 amino acids with a calculated molecular mass of 47.9 kDa and an estimated isoelectric point of 5.97. The deduced amino acid sequence showed 74-89% identity with other Rubisco activases from higher plants. Two highly conserved motifs were identified. Southern blot analysis suggested the presence of a single rca gene in the rice genome. The accumulation of leaf rca mRNA was found to be regulated by an oscillating circadian rhythm, in rice plants grown in a light-dark photoperiod. To purify the rice RCA protein, total soluble protein from rice green leaves was fractionated by ammonium sulfate precipitation, followed by preparative gel electrophoresis. Two polypeptides, designated RCAI and RCAII, were isolated by two-dimensional gel electrophoresis and further confirmed by N-terminal sequencing. The polyclonal antibodies prepared against rice RCAI and RCAII were found to cross-react with two RCA polypeptides present in leaf extracts of spinach and tobacco. Furthermore, two different 3' ends of rca mRNA were detected by reverse transcription-polymerase chain reaction analysis. These cDNA fragments and the related genomic DNA fragment were cloned and sequenced. The sequence of rcaI is almost identical to the corresponding sequence of rcaII, except for its having 33 additional amino acids at the C-terminal portion. It can be concluded that a novel alternative splicing mechanism for a common rca mRNA precursor near the 3' end exists in rice plants.

Photosynthetic Characteristics of Antisense Transgenic Rice Expressing Reduced Levels of Rubisco Activase

“…The expression of genes encoding Rubisco activase (Rca) of rice is high in green organs, especially in leaf blades and leaf sheaths, but almost undetectable in roots. In the leaf blades of rice, Rca expression shows a circadian rhythm the same as observed in other plants (To et al, 1999).…”

“…Rice has two Rubisco activase isoforms with different amino acid sequences at C termini, and their mRNAs are transcribed from a single gene by alternative splicing (To et al, 1999). The expression of genes encoding Rubisco activase (Rca) of rice is high in green organs, especially in leaf blades and leaf sheaths, but almost undetectable in roots.…”

Enhanced Thermostability of Arabidopsis Rubisco Activase Improves Photosynthesis and Growth Rates under Moderate Heat Stress

“…In several plant species, such as Arabidopsis, spinach (Spinacia oleracea), and rice, the short and long isoforms are generated by alternative splicing of a single pre-mRNA (Werneke et al, 1989;To et al, 1999), while in maize, a single pre-mRNA encodes two polypeptides through limited proteolysis of the long isoform at its N-terminal region (Vargas-Suarez et al, 2004). In cotton (Gossypium hirsutum), the short and long isoforms are encoded by separate genes (Salvucci et al, 2003).…”

Characterization of Rubisco Activase Genes in Maize: An α-Isoform Gene Functions alongside a β-Isoform Gene

“…Genomic analyses have identified one RCA gene in spinach (Spinacia oleracea), Arabidopsis, rice, and wheat (Triticum aestivum; Werneke et al, 1988;To et al, 1999;Law and Crafts-Brandner, 2001), two RCA genes in barley (Hordeum vulgare) and cotton (Gossypium hirsutum; Rundle and Zielinski, 1991;Salvucci et al, 2003), and more than three RCA genes in tobacco (Qian and Rodermel, 1993) and soybean (Glycine max; Yin et al, 2010). In some species, such as spinach, Arabidopsis, and rice, alternative splicing of RCA transcripts results in two isoforms of RCA (Werneke et al, 1989;Rundle and Zielinski, 1991;To et al, 1999), whereas in other species such as cotton and soybean, the two RCA isoforms are encoded by different genes (Salvucci et al, 2003;Yin et al, 2010).…”