The DIVa Maturase Binding Site in the Yeast Group II Intron aI2 Is Essential for Intron Homing but Not for In Vivo Splicing

AUTOR(ES)

Huang, Hon-Ren

FONTE

American Society for Microbiology

RESUMO

Splicing of the Saccharomyces cerevisiae mitochondrial DNA group II intron aI2 depends on the intron-encoded 62-kDa reverse transcriptase-maturase protein (p62). In wild-type strains, p62 remains associated with the excised intron lariat RNA in ribonucleoprotein (RNP) particles that are essential for intron homing. Studies of a bacterial group II intron showed that the DIVa substructure of intron domain IV is a high-affinity binding site for its maturase. Here we first present in vitro evidence extending that conclusion to aI2. Then, experiments with aI2 DIVa mutant strains show that the binding of p62 to DIVa is not essential for aI2 splicing in vivo but is essential for homing. Because aI2 splicing in the DIVa mutant strains remains maturase dependent, splicing must rely on other RNA-protein contacts. The p62 that accumulates in the mutant strains has reverse transcriptase activity, but fractionation experiments at high and low salt concentrations show that it associates more weakly than the wild-type protein with endogenous mitochondrial RNAs, and that phenotype probably explains the homing defect. Replacing the DIVa of aI2 with that of the closely related intron aI1 improves in vivo splicing but not homing, indicating that DIVa contributes to the specificity of the maturase-RNA interaction needed for homing.

Documentos Relacionados

• Structure-function relationships in a self-splicing group II intron: a large part of domain II of the mitochondrial intron aI5 is not essential for self-splicing.
• Splicing defective mutants of the COXI gene of yeast mitochondrial DNA: initial definition of the maturase domain of the group II intron aI2.
• Mechanism of maturase-promoted group II intron splicing
• Mitochondrial Mg2+ homeostasis is critical for group II intron splicing in vivo
• A region of group I introns that contains universally conserved residues but is not essential for self-splicing.