Translation attenuation via 3' terminal codon usage in bovine csn1s2 is responsible for the difference in αs2- and ß-casein profile in milk.
RNA Biol
; 12(3): 354-67, 2015.
Article
em En
| MEDLINE
| ID: mdl-25826667
ABSTRACT
The rate of secretion of αs2-casein into bovine milk is approximately 25% of that of ß-casein, yet mammary expression of their respective mRNA transcripts (csn1s2 and csn2) is not different. Our objective was to identify molecular mechanisms that explain the difference in translation efficiency between csn1s2 and csn2. Cell-free translational efficiency of csn2 was 5 times that of csn1s2. Transcripts of csn1s2 distributed into heavier polysomes than csn2 transcripts, indicating an attenuation of elongation and/or termination. Stimulatory and inhibitory effects of the 5' and 3' UTRs on translational efficiency were different with luciferase and casein sequences in the coding regions. Substituting the 5' and 3' UTRs from csn2 into csn1s2 did not improve csn1s2 translation, implicating the coding region itself in the translation difference. Deletion of a 28-codon fragment from the 3' terminus of the csn1s2 coding region, which displays codons with low correlations to cell fitness, increased translation to a par with csn2. We conclude that the usage of the last 28 codons of csn1s2 is the main regulatory element that attenuates its expression and is responsible for the differential translational expression of csn1s2 and csn2.
Palavras-chave
40S, small ribosomal subunit; 60S, large ribosomal subunit; AA, amino acid; ARE, AU-rich element; Apaf-1, apoptosis protease activating factor 1; DLG1, disc large 1 ncosuppressor; FMR1, fragile X mental retardation 1; HRP, horseradish eroxidase; IE, inhibitory element; IRE, iron-responsive element; IRES, nternal ribosome entry site; IRP, iron-regulatory protein; MACT, bovine mammary epithelial cell; PABP, poly(A) binding protein; PAGE, polyacrylamide gel electrophoresis; PCR, polymerase chain reaction; PVDF, polyvinylidene fluoride; RACE, rapid amplification of cDNA ends; RBP, RNA-binding protein; RRL, rabbit reticulocyte lysate; RT, reverse transcription; SDS, sodium dodecyl sulfate; SE, standard error; STR, single-stranded nucleic acid binding protein; TBS-T, Tris-buffered saline containing 0.5%; TfR, transferrin receptor; Tween 20; UTR, untranslated region; aa-tRNA, aminoacyl-tRNA; aaRS, aminoacyl-tRNA synthetase; bovine casein; cDNA, complementary DNA; cell-free translation; coding region; codon usage; eEF, eukaryotic elongation factor; eIF, eukaryotic initiation factor; eRF, eukaryotic termination factor; m7G, 7-methylated uanidine; mRNA, messenger RNA; qPCR, real-time polymerase chain reaction; sAUG, start codon; tRNA, transfer RNA; translational efficiency; uAUG, upstream start codon; uORF, open reading frame; untranslated region; ΔG, free energy
Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Biossíntese de Proteínas
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Códon
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Sequência de Bases
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Caseínas
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Deleção de Sequência
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Leite
Tipo de estudo:
Prognostic_studies
Limite:
Animals
Idioma:
En
Ano de publicação:
2015
Tipo de documento:
Article