Molecular structures and evolution of mouse isozyme genes functioning in the malate-aspartate shuttle

Area of Science:

• Molecular Biology
• Evolutionary Biology
• Biochemistry

Background:

• The malate-aspartate shuttle is crucial for cellular energy metabolism.
• Understanding the transcription of mammalian isozyme genes involved in this shuttle is key to comprehending cellular function.
• Isozymes, such as aspartate aminotransferase (AspAT) and malate dehydrogenase (MDH), exist in both mitochondrial (m) and cytosolic (c) forms.

Purpose of the Study:

• To investigate the molecular mechanisms of transcription for mammalian isozyme genes in the malate-aspartate shuttle.
• To elucidate the structural and evolutionary relationships of these isozymes.
• To characterize the gene organization, cDNA, and genomic DNA of mouse cAspAT, mAspAT, cMDH, and mMDH.

Main Methods:

• Isolation and characterization of cDNAs and genomic DNAs for mouse isozymes.
• Deduction of amino acid sequences and homology analysis.
• Comparative analysis of gene organization, including intron-exon structure and 5' flanking sequences.

Main Results:

• Significant homology was found between mouse cAspAT and mAspAT (47%), and between mouse cMDH and mMDH (23%).
• Surprisingly, inter-species homology (mouse MDH to bacterial MDH) exceeded intra-species homology between mouse mMDH and cMDH.
• Introns were present before the divergence of mitochondrial and cytosolic isozyme genes. Promoter regions lacked TATA/CAAT boxes but contained G+C-rich sequences and multiple transcription start sites. Conserved regions were identified in the 5' flanking sequences of mAspAT, cAspAT, mMDH, and cMDH.

Conclusions:

• The common ancestral MDH gene duplicated before eukaryotic cell emergence, with mammalian and bacterial MDH genes evolving from distinct duplicates.
• Introns predate the divergence of mitochondrial and cytosolic isozyme genes.
• Shared regulatory elements in the 5' flanking regions suggest coordinated transcriptional control for these isozymes.