Upstream Open Reading Frames and Human Genetic Disease


In many eukaryotic messenger ribonucleic acids (mRNAs) one or more short upstream open reading frames (uORFs) precede the initiation codon of the main coding region. For example, in human cells, uORFs are present in about half of the transcripts. Emerging ribosome profiling and peptidomics analyses have recently shown that these uORFs are translated into polypeptides that seem to serve important biological functions. In addition, very interesting examples have shown that these uORFs are cis‐acting RNA elements that can impact gene expression by repressing translation of the downstream main ORF under control conditions and derepressing it under certain pathophysiological stresses. Furthermore, evidence from genetic and bioinformatic studies implicate disturbed uORF‐mediated translational control in the aetiology of human diseases. Identifying more cases and understanding the aberrant mechanisms of uORF‐mediated translational control, as well as discovering the biological function of the uORF‐encoded polypeptides, is fundamental to advance in diagnosis, prognosis and treatment of many human disorders.

Key Concepts:

  • Upstream open reading frames (uORFs) are cis‐acting RNA elements involved in translational regulation, which precede the initiation codon of the main coding region.

  • For a uORF to function as a translational regulatory element, its initiation codon must be recognised, at least at certain times, by the scanning 40S ribosomal subunit and associated translation initiation factors.

  • uORFs can impact gene expression by repressing translation of the downstream main ORF under control conditions, and derepressing it under certain pathophysiological stresses.

  • The impact the uORFs can have on translation depends on variables, such as (1) the distance between the 5′ cap and the uORF, (2) the context in which the uORF AUG (or non‐AUG) is located, (3) the length of the uORF, (4) the sequence and secondary structure of the uORF, (5) the number of uORFs per transcript, (6) the position of the uORF termination codon and (7) the length of the intercistronic sequence(s).

  • uORF‐encoded polypeptides might serve functional roles in cells.

  • Polymorphisms or mutations that introduce/eliminate uORFs or modify the uORF‐encoded peptide can cause human disease.

  • Understanding the mechanisms through which the uORFs regulate gene expression may lead to innovation in diagnosis, prognosis and treatment of many human disorders.

Keywords: upstream open reading frame (uORF); protein synthesis; polypeptide; translational control; gene expression regulation; human genetic disease

Figure 1.

Mechanisms by which uORFs can affect gene expression. (a) The leaky scanning mechanism is dependent on the efficiency of the uAUG (or non‐uAUG) recognition; sometimes the ribosome can translate the uORF, but other timess the scanning machinery bypasses the uAUG, recognising the downstream AUG and translating the main ORF. (b) When a scanning ribosome recognises and translates a functional uORF, there is synthesis of a small peptide; if translation termination of the uORF is efficient, both 60S and 40S ribosomal subunits might dissociate from the transcript and the main ORF is not translated. (c) A uORF can repress translation of the main ORF in a nucleotide or peptide‐dependent manner; in this last case, the uORF‐encoded peptide interacts with the translating machinery and promotes ribosome blockage. Also, the uORF nucleotide sequence can have a role on its translation efficiency, for instance by encoding rare codons that cause the ribosome to stall. (d) After translation termination of the uORF, the 40S ribosomal subunit can remain associated with the transcript, resume scanning and recognise the downstream main AUG – a mechanism designated as translation reinitiation. (e) In response to stress conditions, the presence of one ORF in a transcript can promote an increase of the corresponding protein levels; the higher levels of phosphorylated eIF2α contribute to increase leaky scanning of the uORF and translation of the main ORF is favored. (f) The 5′‐leader sequence containing a uORF, or the uORF, can present a strong secondary structure that impedes the translation of the main ORF by blocking the ribosome. (g) Some uORF‐encoded peptides can affect translation efficiency of the main ORF by trans‐acting in another molecule of transcript. (h) Some uORF‐encoded peptides can play a function in mechanisms other than translational control, for example in DNA repair. (i) The termination codon of a uORF can be recognised as premature and nonsense‐mediated mRNA decay (NMD) is triggered through a mechanism involving the UPF1 protein and ribonucleases. (j) The impact that the uORFs can have on translation depends on (1) distance between the 5′ cap and the uORF (distance to the cap), (2) context in which the uORF AUG (or non‐AUG) is located (AUG context), (3) length of the uORF, (4) number of uORFs per transcript, (5) sequence and secondary structure of the uORF, (6) presence of non‐AUG start codon(s) (7) length of the intercistronic sequence(s) and/or distance between uORFs and (8) position of the uORF termination codon, upstream or downstream of the main initiation codon.

Figure 2.

uORF‐mediated translational deregulation and human disease. Polymorphisms, mutations, alternative processing or other alterations in the transcript that can create, disrupt or modify a uORF (a, b and c, respectively) may affect translational efficiency of the main ORF, as well as the individual phenotype, and culminate in a pathological condition.



Andrews SJ and Rothnagel JA (2014) Emerging evidence for functional peptides encoded by short open reading frames. Nature Reviews Genetics 15(3): 193–204.

Barbosa C, Peixeiro I and Romão L (2013) Gene expression regulation by upstream open reading frames and human disease. PLoS Genetics 9(8): e1003529.

Barbosa C and Romão L (2014) Translation of the human erythropoietin transcript is regulated by an upstream open reading frame in response to hypoxia. RNA 20(5): 594–608.

Beffagna G, Occhi G, Nava A et al. (2005) Regulatory mutations in transforming growth factor‐beta 3 gene cause arrhythmogenic right ventricular cardiomyopathy type 1. Cardiovascular Research 65(2): 366–373.

Bersano A, Ballabio E, Bresolin N et al. (2008) Genetic polymorphisms for the study of multifactorial stroke. Human Mutation 29(6): 776–795.

Bisio A, Nasti S, Jordan JJ et al. (2010) Functional analysis of CDKN2A/p16INK4a 5′‐UTR variants predisposing to melanoma. Human Molecular Genetics 19(8): 1479–1491.

Braverman N, Chen L, Lin P et al. (2002) Mutation analysis of PEX7 in 60 probands with rhizomelic chondrodysplasia punctata and functional correlations of genotype with phenotype. Human Mutation 20(4): 284–297.

Brown CY, Mize GJ, Pineda M, George DL and Morris DR (1999) Role of two upstream open reading frames in the translational control of oncogene mdm2. Oncogene 18(41): 5631–5637.

Calkhoven CF, Muller C and Leutz A (2000) Translational control of C/EBPalpha and C/EBPbeta isoform expression. Genes and Development 14(15): 1920–1932.

Calvo SE, Pagliarini DJ and Mootha VK (2009) Upstream open reading frames cause widespread reduction of protein expression and are polymorphic among humans. Proceedings of the National Academy of Sciences of the USA 106(18): 7507–7512.

Cazzola M and Skoda RC (2000) Translational pathophysiology: a novel molecular mechanism of human disease. Blood 95(11): 3280–3288.

Davies WL, Vandenberg JI, Sayeed RA and Trezise AE (2004) Post‐transcriptional regulation of the cystic fibrosis gene in cardiac development and hypertrophy. Biochemical and Biophysical Research Communications 319(2): 410–418.

Ghilardi N and Skoda RC (1999) A single‐base deletion in the thrombopoietin (TPO) gene causes familial essential thrombocythemia through a mechanism of more efficient translation of TPO mRNA. Blood 94(4): 1480–1482.

Ghilardi N, Wiestner A, Kikuchi M, Ohsaka A and Skoda RC (1999) Hereditary thrombocythaemia in a Japanese family is caused by a novel point mutation in the thrombopoietin gene. British Journal of Haematology 107(2): 310–316.

Huopio H, Jääskeläinen J, Komulainen J et al. (2002) Acute insulin response tests for the differential diagnosis of congenital hyperinsulinism. Journal of Clinical Endocrinology and Metabolism 87(10): 4502–4507.

Ingolia NT, Lareau LF and Weissman JS (2011) Ribosome profiling of mouse embryonic stem cells reveals the complexity and dynamics of mammalian proteomes. Cell 147(4): 789–802.

Jousse C, Bruhat A, Carraro V et al. (2001) Inhibition of CHOP translation by a peptide encoded by an open reading frame localized in the chop5′UTR. Nucleic Acids Research 29(21): 4341–4351.

Kanaji T, Okamura T, Osaki K et al. (1998) A common genetic polymorphism (46 C to T substitution) in the 5′‐untranslated region of the coagulation factor XII gene is associated with low translation efficiency and decrease in plasma factor XII level. Blood 91(6): 2010–2014.

Kondo S, Schutte BC, Richardson RJ et al. (2002) Mutations in IRF6 cause Van der Woude and popliteal pterygium syndromes. Nature Genetics 32(2): 285–289.

Kondo T, Okabe M, Sanada M et al. (1998) Familial essential thrombocythemia associated with one‐base deletion in the 5′‐untranslated region of the thrombopoietin gene. Blood 92(4): 1091–1096.

Kozak M (1986) Point mutations define a sequence flanking the AUG initiator codon that modulates translation by eukaryotic ribosomes. Cell 44(2): 283–292.

Kozak M (1989) Circumstances and mechanisms of inhibition of translation by secondary structure in eucaryotic mRNAs. Molecular Cell Biology 9(11): 5134–5142.

Krude H, Biebermann H, Luck W et al. (1998) Severe early‐onset obesity, adrenal insufficiency and red hair pigmentation caused by POMC mutations in humans. Nature Genetics 19(2): 155–157.

Liu L, Dilworth D, Gao L et al. (1999) Mutation of the CDKN2A 5′ UTR creates an aberrant initiation codon and predisposes to melanoma. Nature Genetics 21(1): 128–132.

Lovett PS and Rogers EJ (1996) Ribosome regulation by the nascent peptide. Microbiological Reviews 60(2): 366–385.

Lukowski SW, Bombieri C and Trezise AEO (2011) Disrupted posttranscriptional regulation of the cystic fibrosis transmembrane conductance regulator (CFTR) by a 5′UTR mutation is associated with a cftr‐related disease. Human Mutation 32(10): E2266–E2282.

Mendell JT, Sharifi NA, Meyers JL, Martinez-Murillo F and Dietz HC (2004) Nonsense surveillance regulates expression of diverse classes of mammalian transcripts and mutes genomic noise. Nature Genetics 36(10): 1073–1078.

Menschaert G, Van Criekinge W, Notelaers T et al. (2013) Deep proteome coverage based on ribosome profiling aids mass spectrometry‐based protein and peptide discovery and provides evidence of alternative translation products and near‐cognate translation initiation events. Molecular and Cellular Proteomics 12(7): 1780–1790.

Mihailovich M, Thermann R, Grohovaz F, Hentze MW and Zacchetti D (2007) Complex translational regulation of BACE1 involves upstream AUGs and stimulatory elements within the 5′ untranslated region. Nucleic Acids Research 35(9): 2975–2985.

Nguyen HL, Yang X and Omiecinski CJ (2013) Expression of a novel mRNA transcript for human microsomal epoxide hydrolase (EPHX1) is regulated by short open reading frames within its 5′‐untranslated region. RNA 19(6): 752–766.

Niesler B, Flohr T, Nöthen MM et al. (2001) Association between the 5′ UTR variant C178T of the serotonin receptor gene HTR3A and bipolar affective disorder. Pharmacogenetics 11(6): 471–475.

Occhi G, Regazzo D, Trivellin G et al. (2013) A novel mutation in the upstream open reading frame of the CDKN1B gene causes a MEN4 phenotype. PLoS Genetics 9(3): e1003350.

O'Connor T, Sadleir KR, Maus E et al. (2008) Phosphorylation of the translation initiation factor eIF2alpha increases BACE1 levels and promotes amyloidogenesis. Neuron 60(6): 988–1009.

Oner R, Agarwal S, Dimovski AJ et al. (1991) The G→A mutation at position +22 3′ to the Cap site of the beta‐globin gene as a possible cause for a beta‐thalassemia. Hemoglobin 15(1–2): 67–76.

Palam LR, Baird TD and Wek RC (2011) Phosphorylation of eIF2 facilitates ribosomal bypass of an inhibitory upstream ORF to enhance CHOP translation. Journal of Biological Chemistry 286(13): 10939–10949.

Pasaje CFA, Bae JS, Park B‐L et al. (2012) WDR46 is a genetic risk factor for aspirin‐exacerbated respiratory disease in a Korean population. Allergy, Asthma and Immunology Research 4(4): 199–205.

Pichon X, Wilson LA, Stoneley M et al. (2012) RNA binding protein/RNA element interactions and the control of translation. Current Protein and Peptide Science 13(4): 294–304.

Poulat F, Desclozeaux M, Tuffery S et al. (1998) Mutation in the 5′ noncoding region of the SRY gene in an XY sex‐reversed patient. Human Mutation (suppl. 1): S192–S194.

Poyry TAA, Kaminski A and Jackson RJ (2004) What determines whether mammalian ribosomes resume scanning after translation of a short upstream open reading frame? Genes and Development 18(1): 62–75.

Rideau A, Mangeat B, Matthes T, Trono D and Beris P (2007) Molecular mechanism of hepcidin deficiency in a patient with juvenile hemochromatosis. Haematologica 92(1): 127–128.

Rogers GW Jr, Edelman GM and Mauro VP (2004) Differential utilization of upstream AUGs in the β‐secretase mRNA suggests that a shunting mechanism regulates translation. Proceedings of the National Academy of Sciences of the USA 101(9): 2794–2799.

Roy B, Vaughn JN, Kim B‐H et al. (2010) The h subunit of eIF3 promotes reinitiation competence during translation of mRNAs harboring upstream open reading frames. RNA 16(4): 748–761.

Sachs MS and Geballe AP (2006) Downstream control of upstream open reading frames. Genes and Development 20(8): 915–921.

Sherry ST, Ward MH, Kholodov M et al. (2001) dbSNP: the NCBI database of genetic variation. Nucleic Acids Research 29(1): 308–311.

Silva AL and Romão L (2009) The mammalian nonsense‐mediated mRNA decay pathway: to decay or not to decay! Which players make the decision? FEBS Letters 583(3): 499–505.

Sivagnanasundaram S, Morris AG, Gaitonde EJ et al. (2000) A cluster of single nucleotide polymorphisms in the 5′‐leader of the human dopamine D3 receptor gene (DRD3) and its relationship to schizophrenia. Neuroscience Letters 279(1): 13–16.

Slavoff SA, Heo J, Budnik BA et al. (2014) A human short ORF‐encoded peptide that stimulates DNA end joining. Journal of Biological Chemistry 289(16): 10950–10957.

Slavoff SA, Mitchell AJ, Schwaid AG, Hanakahi LA and Saghatelian A (2013) Peptidomic discovery of short open reading frame‐encoded peptides in human cells. Nature Chemical Biology 9(1): 59–64.

Somers J, Pöyry T and Willis AE (2013) A perspective on mammalian upstream open reading frame function. International Journal of Biochemistry and Cell Biology 45(8): 1690–1700.

Sonenberg N and Hinnebusch AG (2009) Regulation of translation initiation in eukaryotes: mechanisms and biological targets. Cell 136(4): 731–745.

Sözen MM, Whittall R, Öner C et al. (2005) The molecular basis of familial hypercholesterolaemia in Turkish patients. Atherosclerosis 180(1): 63–71.

Tassin J, Dürr A, Bonnet AM et al. (2000) Levodopa‐responsive dystonia. GTP cyclohydrolase I or parkin mutations? Brain 123(Pt 6): 1112–1121.

Vanderperre B, Lucier JF, Bissonnette C et al. (2013) Direct detection of alternative open reading frames translation products in human significantly expands the proteome. PLoS One 8(8): e70698.

Watatani Y, Ichikawa K, Nakanishi N et al. (2008) Stress‐induced translation of ATF5 mRNA is regulated by the 5′‐untranslated region. Journal of Biological Chemistry 283(5): 2543–2553.

Wen Y, Liu Y, Xu Y et al. (2009) Loss‐of‐function mutations of an inhibitory upstream ORF in the human hairless transcript cause Marie Unna hereditary hypotrichosis. Nature Genetics 41(2): 228–233.

Wethmar K, Begay V, Smink JJ et al. (2010b) C/EBP uORF mice – a genetic model for uORF‐mediated translational control in mammals. Genes and Development 24(1): 15–20.

Wethmar K, Smink JJ and Leutz A (2010a) Upstream open reading frames: molecular switches in (patho)physiology. BioEssays 32(10): 885–893.

Wiestner A, Schlemper RJ, van der Maas AP and Skoda RC (1998) An activating splice donor mutation in the thrombopoietin gene causes hereditary thrombocythaemia. Nature Genetics 18(1): 49–52.

Witt H, Luck W, Hennies HC et al. (2000) Mutations in the gene encoding the serine protease inhibitor, Kazal type 1 are associated with chronic pancreatitis. Nature Genetics 25(1): 213–216.

Yepiskoposyan H, Aeschimann F, Nilsson D, Okoniewski M and Mühlemann O (2011) Autoregulation of the nonsense‐mediated mRNA decay pathway in human cells. RNA 17(12): 2108–2118.

Zhao C, Datta S, Mandal P, Xu S and Hamilton T (2010) Stress‐sensitive regulation of IFRD1 mRNA decay is mediated by an upstream open reading frame. Journal of Biological Chemistry 285(12): 8552–8562.

Zhou W and Song W (2006) Leaky scanning and reinitiation regulate BACE1 gene expression. Molecular Cellular Biology 26(9): 3353–3364.

Further Reading

Gebauer F and Hentze MW (2004) Molecular mechanisms of translational control. Nature Review Molecular Cell Biology 5(10): 827–835.

Pestova TV, Lorsch JR and Hellen CUT (2007) The mechanism of translation initiation in eukaryotes. In: Mathews MB, Sonenberg N and Hershey JWB (eds) Translational Control in Biology and Medicine, pp. 87–128. Cold Spring Harbor, NY: Cold Spring Harbor Laboratory Press.

Contact Editor close
Submit a note to the editor about this article by filling in the form below.

* Required Field

How to Cite close
Barbosa, Cristina, Onofre, Cláudia, and Romão, Luísa(Aug 2014) Upstream Open Reading Frames and Human Genetic Disease. In: eLS. John Wiley & Sons Ltd, Chichester. [doi: 10.1002/9780470015902.a0025714]