Choice of a start codon in a single transcript determines DNA ligase 1 isoform production and intracellular targeting in Arabidopsis thaliana

Abstract

DNA ligase 1 (AtLIG1) is the only essential DNA ligase activity in Arabidopsis and is implicated in the important processes of DNA replication, repair and recombination and in transgene insertion during Agrobacterium-mediated plant transformations. The mitochondrial and nuclear forms of DNA ligase 1 in Arabidopsis are translated from a single mRNA species through the control of translation initiation from either the first (M1) or second (M2) in-frame AUG codons respectively. Translation from a third in-frame AUG codon (M3) occurs on transcripts in which M1 and M2 are mutagenized to stop codons. Wild-type AtLIG1-GFP constructs (where GFP stands for green fluorescent protein) can be targeted in planta to both the nucleus and mitochondria. AtLIG1-GFP translation from M1 specifically targets the fusion protein only to mitochondria in planta, whereas translation from M2 or M3 targets the fusion protein only to the nucleus. Interestingly, the AtLIG1-GFP fusion protein in which translation is initiated from M1 contains both an N-terminal mtPS (mitochondrial targeting presequence) and a nuclear localization signal; nonetheless, this protein is only targeted to the mitochondria. This result raises intriguing questions on the translational control mechanisms that regulate how the protein products of a single transcript are targeted to more than one cellular compartment.

Comparative survey of plastid and mitochondrial targeting properties of transcription factors in Arabidopsis and rice

“…In case of the DNA ligase 1, translation initiation from a first in-frame start codon produces a protein that is exclusively targeted to mitochondria, while the use of an alternative second start codon produces a protein that is found only in the nucleus (Sunderland et al 2006). The existence of a chloroplast-localized protein initiated at a potential third AUG that was previously proposed (Sunderland et al 2004) could not be confirmed.…”

Alternative translation start sites and hidden coding potential of eukaryotic mRNAs

“…The nucleotide context around the alternative start codons in the AtLIG1 transcripts affects the translation initiation to ensure a balanced synthesis of both nuclear and mitochondrial AtLIG1 isoforms, probably via a context-dependent leaky scanning. (24) Alternative TISs and leaky scanning are responsible for the synthesis of plastidic and cytoplasmic/ nuclear isoforms of tRNA ligase of Arabidopsis thaliana and Oryza sativa, plastidic and mitochondrial isoforms of spinach protoporphyrinogen oxidase and arabidopsis DNA polymerase. (15,25,26) The mRNA of a human spastin gene contains two alternative translation start sites and produces two protein isoforms, 68 and 60 kDa, respectively.…”

Mitochondrial Biogenesis and Function in Arabidopsis†

“…What became evident with the complete sequencing of both the mitochondrial and nuclear genomes was the added complexity and mixed evolutionary origin of the components involved in expressing the mitochondrial genome. The basic replication machinery involves at least a DNA polymerase (Christensen et al, 2005), a DNA gyrase (Wall et al, 2004) and a DNA ligase (Sunderland et al, 2004(Sunderland et al, , 2006. Plant mitochondrial genomes are organised as multiple circular molecules that undergo a high frequency of inter-and intramolecular recombination (Lonsdale et al, 1984;Mackenzie and McIntosh, 1999).…”