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Plant Molecular Biology

, Volume 37, Issue 2, pp 243–263 | Cite as

Alternative splicing of a precursor-mRNA encoded by the Chlorella sorokiniana NADP-specific glutamate dehydrogenase gene yields mRNAs for precursor proteins of isozyme subunits with different ammonium affinities

  • Philip W. Miller
  • Waltraud I. Dunn
  • Robert R. Schmidt
Article

Abstract

Chlorella sorokiniana has seven ammonium-inducible, chloroplastic NADP-specific glutamate dehydrogenase (NADP-GDH) isozymes composed of varying ratios of α- and β-subunits. Southern blot and allele-specific PCR analyses indicate that the C. sorokiniana genome possesses a single 7178 bp nuclear NADP-GDH gene. cDNA cloning and sequencing, 5′-RACE-PCR analysis, and RNase protection analysis identified two NADP-GDH mRNAs that are identical with the exception of a 42 nt sequence located within the 5′-coding region of the longer mRNA. The 42 nt sequence, termed an auxon because it serves as an exon or intron, appears to undergo alternative splicing from the precursor mRNA by a process that is regulated by both nutritional and environmental signals. Depending upon whether the auxon is included or excluded in a mature mRNA, the gene can be considered to consist of 22 or 23 exons, respectively. The 2074 and 2116 nt mRNAs encode precursor proteins of 56350 and 57850 Da, respectively. The N-termini of the purified mature α- and β-subunits were sequenced, identifying full-length subunits of 53501 and 52342 Da, respectively. The sequences of the subunits are identical except for an 11 amino acid extension at the N-terminus of the α-subunit. The α-subunit has an additional α-helical domain at its N-terminus compared with the β-subunit. By correlating the abundances of the two mRNAs with the levels (and relative turnover rates) of the α- and β-subunit antigens during induction in Chlorella, the larger mRNA is proposed to encode the larger subunit.

alternative splicing Chlorella sorokiniana chloroplastic glutamate dehydrogenase isozymes precursor mRNA 

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Copyright information

© Kluwer Academic Publishers 1998

Authors and Affiliations

  • Philip W. Miller
    • 2
  • Waltraud I. Dunn
    • 1
  • Robert R. Schmidt
    • 1
  1. 1.Department of Microbiology and Cell ScienceUniversity of FloridasGainesvilleUSA
  2. 2.The Monsanto CompanyChesterfieldUSA

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