Upstream open reading frame in 5′-untranslated region reduces titin mRNA translational efficiency
| Autor: | Mauricio Valenzuela, Adrian G. Cadar, Angel Lin, Chee Chew Lim, Lin Zhong |
|---|---|
| Rok vydání: | 2014 |
| Předmět: |
Sarcomeres
Untranslated region animal structures Five prime untranslated region Translational efficiency RNA Stability Molecular Sequence Data Biophysics Obscurin Polymorphism Single Nucleotide Biochemistry Article Cell Line Mice Open Reading Frames Sequence Homology Nucleic Acid Upstream open reading frame Animals Humans Myotilin Connectin Myocytes Cardiac RNA Messenger Molecular Biology Base Sequence biology Cell Biology musculoskeletal system Molecular biology Rats Actinin alpha 2 HEK293 Cells Gene Expression Regulation Doxorubicin embryonic structures cardiovascular system biology.protein Titin 5' Untranslated Regions Protein Kinases tissues Half-Life |
| Zdroj: | Biochemical and Biophysical Research Communications. 453:185-191 |
| ISSN: | 0006-291X |
| Popis: | Titin is the largest known protein and a critical determinant of myofibril elasticity and sarcomere structure in striated muscle. Accumulating evidence that mRNA transcripts are post-transcriptionally regulated by specific motifs located in the flanking untranslated regions (UTRs) led us to consider the role of titin 5'-UTR in regulating its translational efficiency. Titin 5'-UTR is highly homologous between human, mouse, and rat, and sequence analysis revealed the presence of a stem-loop and two upstream AUG codons (uAUGs) converging on a shared in frame stop codon. We generated a mouse titin 5'-UTR luciferase reporter construct and targeted the stem-loop and each uAUG for mutation. The wild-type and mutated constructs were transfected into the cardiac HL-1 cell line and primary neonatal rat ventricular myocytes (NRVM). SV40 driven 5'-UTR luciferase activity was significantly suppressed by wild-type titin 5'-UTR (∼ 70% in HL-1 cells and ∼ 60% in NRVM). Mutating both uAUGs was found to alleviate titin 5'-UTR suppression, while eliminating the stem-loop had no effect. Treatment with various growth stimuli: pacing, PMA or neuregulin had no effect on titin 5'-UTR luciferase activity. Doxorubicin stress stimuli reduced titin 5'-UTR suppression, while H2O2 had no effect. A reported single nucleotide polymorphism (SNP) rs13422986 at position -4 of the uAUG2 was introduced and found to further repress titin 5'-UTR luciferase activity. We conclude that the uAUG motifs in titin 5'-UTR serve as translational repressors in the control of titin gene expression, and that mutations/SNPs of the uAUGs or doxorubicin stress could alter titin translational efficiency. |
| Databáze: | OpenAIRE |
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