Minigene-like inhibition of protein synthesis mediated by hungry codons near the start codon.
Rare AGA or AGG codons close to the initiation codon inhibit protein synthesis by a tRNA-sequestering mechanism as toxic minigenes do. To further understand this mechanism, a parallel analysis of protein synthesis and peptidyl-tRNA accumulation was performed using both a set of lacZ constructs where AGAAGA codons were moved codon by codon from +2, +3 up to +7, +8 positions and a series of 3-8 codon minigenes containing AGAAGA codons before the stop codon. Beta-galactosidase synthesis from the AGAAGA lacZ constructs (in a Pth defective in vitro system without exogenous tRNA) diminished as the AGAAGA codons were closer to AUG codon. Likewise, beta-galactosidase expression from the reporter +7 AGA lacZ gene (plus tRNA, 0.25 microg/microl) waned as the AGAAGAUAA minigene shortened. Pth counteracted both the length-dependent minigene effect on the expression of beta-galactosidase from the +7 AGA lacZ reporter gene and the positional effect from the AGAAGA lacZ constructs. The +2, +3 AGAAGA lacZ construct and the shortest +2, +3 AGAAGAUAA minigene accumulated the highest percentage of peptidyl-tRNA(Arg4). These observations lead us to propose that hungry codons at early positions, albeit with less strength, inhibit protein synthesis by a minigene-like mechanism involving accumulation of peptidyl-tRNA.
Jacinto-Loeza
Eva
E
Departamento de Genética y Biología Molecular, Centro de Investigación y de Estudios Avanzados del IPN, Apartado Postal 14-740, México DF, 07000, México.
Vivanco-Domínguez
Serafín
S
Guarneros
Gabriel
G
Hernández-Sánchez
Javier
J
D002265:Carboxylic Ester Hydrolases / Q000378:metabolism; D003062:Codon; D018387:Codon, Initiator; D005796:Genes; D017930:Genes, Reporter; D016366:Open Reading Frames; D014176:Protein Biosynthesis; D012346:RNA, Transfer, Amino Acyl / Q000378:metabolism
D003062:Codon; D018387:Codon, Initiator; D012346:RNA, Transfer, Amino Acyl; C019037:tRNA, peptidyl-; D002265:Carboxylic Ester Hydrolases; C019674:aminoacyl-tRNA hydrolase