Antigenic peptides: Reviving nuclear translation
2
NATURE CHEMICAL BIOLOGY | VOL 9 | DECEMBER 2013 | www.nature.com/naturechemicalbiology 759 news & views to be effective. Nevertheless, the synthetic allergen approach is amenable to modeling additional complexity, with haptens replaced by peptide epitope mimics and more than one low-affinity antibody. Achilles had just one weak spot that led to his downfall; an allergic response might prove to be more resilient in response to an attack at a single point. ■ Brian J. Sutton is in the Randall Division of Cell & Molecular Biophysics and the Medical Research Council & Asthma UK Centre in Allergic Mechanisms of Asthma, King’s College London, London, UK. e-mail: [email protected] References 1. Handlogten, M.W., Kiziltepe, T., Serezani, A.P., Kaplan, M.H. & Bilgicer, B. Nat. Chem. Biol. 9, 789–795 (2013). 2. Gould, H.J. & Sutton, B.J. Nat. Rev. Immunol. 8, 205–217 (2008). 3. Hunt, J. et al. J. Biol. Chem. 287, 17459–17470 (2012). 4. Metzger, H., Eglite, S., Haleem-Smith, H., Reischl, I. & Torigoe, C. Mol. Immunol. 38, 1207–1211 (2002). 5. Posner, R.G. et al. Org. Lett. 9, 3551–3554 (2007). 6. Sil, D., Lee, J.B., Luo, D., Holowka, D. & Baird, B. ACS Chem. Biol. 2, 674–684 (2007). 7. Andrews, N.L. et al. Immunity 31, 469–479 (2009). 8. Rajagopalan, K. et al. Proc. Natl. Acad. Sci. USA 93, 6019–6024 (1996). 9. Handlogten, M.W., Kiziltepe, T., Moustakas, D.T. & Bilgicer, B. Chem. Biol. 18, 1179–1188 (2011). 10. Christensen, L.H., Holm, J., Lund, G., Riise, E. & Lund, K. J. Allergy Clin. Immunol. 122, 298–304 (2008). Competing financial interests e author declares no competing financial interests. ANTIGENIC PEPTIDES Reviving nuclear translation Noncanonical translation of prespliced mRNA provides physiological meaning to nuclear translation in generating antigenic peptide substrates for the endogenous major histocompatibility complex class I pathway. Petra Van Damme & Gerben Menschaert I n 1974, the cell surface presentation of antigenic peptide-bound major histocompatibility complex class I molecules (pMHC class I) was found to mediate CD8 + T-cell immunity. It was initially believed that antigenic peptides were raised by proteasomal protein degradation. Recent evidence, however, has suggested that unconventional sources of peptide epitopes generated by a distinct mRNA translation process contribute to the MHC I ligandome 1,2 . For example, mRNA- derived full-length protein production was shown to lag far behind production of antigenic peptides 1 , pointing to pioneer translation products generated during early stages of mRNA maturation or otherwise defective ribosomal products recently shown to include translation products from alternative or defective mRNAs (that is, cryptic epitopes originating for alternate reading frames, introns, nontranslated regions, downstream initiation and stop codon readthrough) as providing alternative sources of biological relevant antigens for MHC class I molecules 1–3 . ese studies have reoriented the focus toward protein synthesis rather than protein degradation as the source of antigenic peptide substrates. New results from Apcher et al. 4 now infer a spatiotemporal dimension to MHC class I antigenic peptide production in which substrates are produced within the nucleus before mRNA matures (Fig. 1). e concept of nuclear translation, described for the first time by Allfrey et al. 5 in the early 1950s, has been heavily debated, though with few converts in the intervening years. Despite some favorable evidence for nuclear translation, the lack of controls and use of impure nuclei or damaged cells meant that this evidence had little influence on the prevailing view of the cytoplasm being the scene of restricted translation. Recently, however, the concept of nuclear translation was revived in a study by David et al. 6 , which demonstrated substantial nucleolar enriched translation in intact cells by means of ribopuromycylation, creating stably associated complexes of ribosomes with puromycylated nascent chains. Interestingly, this study also showed that nuclear translation was specifically inhibited upon influenza infection. One explanation for this result is that nuclear translation functions in the quality control of mRNA transcripts by processes such as nonsense-mediated decay (NMD), a surveillance pathway that reduces errors in gene expression by eliminating mRNA transcripts containing premature stop codons (PTCs), which generate defective ribosomal products as a consequence. Another attractive hypothesis is that certain viruses have evolved mechanisms to prevent antigenic peptide formation during the pioneer round of mRNA translation 1 . Apcher et al. 4 address this controversial topic from a different angle by tracking the translational origin of MHC class I peptides. Employing the high sensitivity of detection of MHC class I–presented peptides by specific T cells, the authors introduced antigenic peptide-encoding sequences (or antigenic epitopes), known to be presented on specific MHC class I molecules, in the exonic or intronic sequences of β-globin, with or without a PTC to provoke NMD. Irrespective of the identity of the antigenic peptide, the intronic or exonic location of its mRNA or whether the mRNA was targeted for the NMD pathway, MHC class I–specific peptide presentation could be observed in all cases where the epitope was in frame with the main open reading frame (ORF). This combined data lead to the conclusion that ribosomal activity on prespliced mRNA produces pioneering translation products and that these are an important source of antigenic peptide substrates for the MHC class I pathway. The authors further provide evidence that epitope expression correlates with nuclear retention of its spliced or unspliced mRNA origin, as nuclear mRNA export decreased whereas nuclear retention increased antigen presentation. Finally, ribopuromycylation and proximal ligation assays demonstrated the presence of nascent intron-derived antigenic peptides in close proximity with ribosomal proteins within the nuclear compartment. Overall, these data elegantly demonstrate a role for nuclear translation in generating antigenic peptides from constructs while providing a logical explanation for the need for compartmentalization in limiting competition for MHC I accessibility. e results from Apcher et al. 4 will definitely revive interest in nuclear translation by adding translation products from prespliced mRNAs to the list of antigenic peptide sources. However, one caveat to this work is that cellular stress or non-natural situationssuch as the overexpression system usedmight alter npg © 2013 Nature America, Inc. All rights reserved.
Transcript of Antigenic peptides: Reviving nuclear translation
nature chemical biology | VOL 9 | DECEMBER 2013 | www.nature.com/naturechemicalbiology 759
news & views
to be effective. Nevertheless, the synthetic allergen approach is amenable to modeling additional complexity, with haptens replaced by peptide epitope mimics and more than one low-affinity antibody. Achilles had just one weak spot that led to his downfall; an allergic response might prove to be more resilient in response to an attack at a single point. ■
Brian J. Sutton is in the Randall Division of Cell & Molecular Biophysics and the Medical
Research Council & Asthma UK Centre in Allergic Mechanisms of Asthma, King’s College London, London, UK. e-mail: [email protected]
References1. Handlogten, M.W., Kiziltepe, T., Serezani, A.P., Kaplan, M.H. &
Bilgicer, B. Nat. Chem. Biol. 9, 789–795 (2013).2. Gould, H.J. & Sutton, B.J. Nat. Rev. Immunol. 8, 205–217
(2008).3. Hunt, J. et al. J. Biol. Chem. 287, 17459–17470 (2012).4. Metzger, H., Eglite, S., Haleem-Smith, H., Reischl, I. &
Torigoe, C. Mol. Immunol. 38, 1207–1211 (2002).
5. Posner, R.G. et al. Org. Lett. 9, 3551–3554 (2007).6. Sil, D., Lee, J.B., Luo, D., Holowka, D. & Baird, B. ACS Chem.
Biol. 2, 674–684 (2007).7. Andrews, N.L. et al. Immunity 31, 469–479 (2009).8. Rajagopalan, K. et al. Proc. Natl. Acad. Sci. USA 93,
6019–6024 (1996).9. Handlogten, M.W., Kiziltepe, T., Moustakas, D.T. &
Bilgicer, B. Chem. Biol. 18, 1179–1188 (2011).10. Christensen, L.H., Holm, J., Lund, G., Riise, E. & Lund, K.
J. Allergy Clin. Immunol. 122, 298–304 (2008).
Competing financial interestsThe author declares no competing financial interests.
ANTIGENIC PEPTIDES
Reviving nuclear translationNoncanonical translation of prespliced mRNA provides physiological meaning to nuclear translation in generating antigenic peptide substrates for the endogenous major histocompatibility complex class I pathway.
Petra Van Damme & Gerben Menschaert
In 1974, the cell surface presentation of antigenic peptide-bound major histocompatibility complex class I
molecules (pMHC class I) was found to mediate CD8+ T-cell immunity. It was initially believed that antigenic peptides were raised by proteasomal protein degradation. Recent evidence, however, has suggested that unconventional sources of peptide epitopes generated by a distinct mRNA translation process contribute to the MHC I ligandome1,2. For example, mRNA-derived full-length protein production was shown to lag far behind production of antigenic peptides1, pointing to pioneer translation products generated during early stages of mRNA maturation or otherwise defective ribosomal products recently shown to include translation products from alternative or defective mRNAs (that is, cryptic epitopes originating for alternate reading frames, introns, nontranslated regions, downstream initiation and stop codon readthrough) as providing alternative sources of biological relevant antigens for MHC class I molecules1–3. These studies have reoriented the focus toward protein synthesis rather than protein degradation as the source of antigenic peptide substrates. New results from Apcher et al.4 now infer a spatiotemporal dimension to MHC class I antigenic peptide production in which substrates are produced within the nucleus before mRNA matures (Fig. 1).
The concept of nuclear translation, described for the first time by Allfrey et al.5 in the early 1950s, has been heavily debated, though with few converts in the intervening years. Despite some favorable evidence for
nuclear translation, the lack of controls and use of impure nuclei or damaged cells meant that this evidence had little influence on the prevailing view of the cytoplasm being the scene of restricted translation. Recently, however, the concept of nuclear translation was revived in a study by David et al.6, which demonstrated substantial nucleolar enriched translation in intact cells by means of ribopuromycylation, creating stably associated complexes of ribosomes with puromycylated nascent chains. Interestingly, this study also showed that nuclear translation was specifically inhibited upon influenza infection. One explanation for this result is that nuclear translation functions in the quality control of mRNA transcripts by processes such as nonsense-mediated decay (NMD), a surveillance pathway that reduces errors in gene expression by eliminating mRNA transcripts containing premature stop codons (PTCs), which generate defective ribosomal products as a consequence. Another attractive hypothesis is that certain viruses have evolved mechanisms to prevent antigenic peptide formation during the pioneer round of mRNA translation1.
Apcher et al.4 address this controversial topic from a different angle by tracking the translational origin of MHC class I peptides. Employing the high sensitivity of detection of MHC class I–presented peptides by specific T cells, the authors introduced antigenic peptide-encoding sequences (or antigenic epitopes), known to be presented on specific MHC class I molecules, in the exonic or intronic sequences of β-globin, with or without
a PTC to provoke NMD. Irrespective of the identity of the antigenic peptide, the intronic or exonic location of its mRNA or whether the mRNA was targeted for the NMD pathway, MHC class I–specific peptide presentation could be observed in all cases where the epitope was in frame with the main open reading frame (ORF). This combined data lead to the conclusion that ribosomal activity on prespliced mRNA produces pioneering translation products and that these are an important source of antigenic peptide substrates for the MHC class I pathway. The authors further provide evidence that epitope expression correlates with nuclear retention of its spliced or unspliced mRNA origin, as nuclear mRNA export decreased whereas nuclear retention increased antigen presentation. Finally, ribopuromycylation and proximal ligation assays demonstrated the presence of nascent intron-derived antigenic peptides in close proximity with ribosomal proteins within the nuclear compartment. Overall, these data elegantly demonstrate a role for nuclear translation in generating antigenic peptides from constructs while providing a logical explanation for the need for compartmentalization in limiting competition for MHC I accessibility.
The results from Apcher et al.4 will definitely revive interest in nuclear translation by adding translation products from prespliced mRNAs to the list of antigenic peptide sources. However, one caveat to this work is that cellular stress or non-natural situationssuch as the overexpression system usedmight alter
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760 nature chemical biology | VOL 9 | DECEMBER 2013 | www.nature.com/naturechemicalbiology
news & views
translational behavior in addition to protein folding, assembly, processing and trafficking. As such, it remains to be discovered whether similar events occur in a natural protein context. Further, as nuclear ER extensions have shown to deeply invaginate nuclei7, additional evidence that these extensions or other microenvironments are not involved are crucial to establish the nucleus as an alternative translation scene.
The puromycin-tagging method6, here applied to detect nuclear nascent peptides
holding antigenic MHC class I epitopes, can further be exploited to provide genome- and proteome-wide evidence for nuclear translation events. In combination with the enrichment of pure nuclei, two recent technologies termed ribosome profiling8 and puromycin-associated nascent chain proteomics (PUNCH-P)9 may prove to be of indispensible value. Ribosome profiling, based on deep sequencing of ribosome-protected mRNA fragments, enables a genome-wide
delineation of ORFs, including small ORFs (sORFs). Whereas MS-based antigenic peptide mapping has typically relied on databases holding canonical protein products, ribosome profiling additionally enables investigation of the cryptic peptide repertoire as puromycin or alternative antibiotic treatments8,10 lead to ribosome accumulation at both cognate and near-cognate translation start sites, facilitating their discovery. Interestingly, highly conserved intronic sORFs have recently been detected11. Although the majority of such ORFs most likely point to missed splice annotations, some conserved ORFs might even hint at nuclear translation serving another biological purpose.
For the first time, a physiological role for nuclear translation is being put forward by coupling it to antigen presentation. This way, nuclear translation might ensure a dedicated immune surveillance mechanism in guaranteeing the efficient formation of viral pMHC1 complexes and the presentation of a comprehensive repertoire of self epitopes by portraying antigenic peptides indicative of differential mRNA splicing. Further proof is necessary to extend these initial findings to unravel endogenous antigen presentation before this concept can pave the way for improving current immune therapies. ■
Petra Van Damme is at the Department of Medical Protein Research, VIB, Ghent, Belgium, and the Department of Biochemistry, Ghent University, Ghent, Belgium. Gerben Menschaert is at the Laboratory for Bioinformatics and Computational Genomics, Department of Mathematical Modeling, Statistics and Bioinformatics, Faculty of Bioscience Engineering, Ghent University, Ghent, Belgium. e-mail: [email protected]
References1. Apcher, S. et al. Proc. Natl. Acad. Sci. USA 108, 11572–11577
(2011).2. Starck, S.R. & Shastri, N. Cell Mol. Life Sci. 68, 1471–1479
(2011).3. Yewdell, J.W. Trends Immunol. 32, 548–558 (2011).4. Apcher, S. et al. Proc. Natl. Acad. Sci. USA doi:10.1073/
pnas.1309956110 (30 September 2013).5. Allfrey, V.G. & Mirsky, A.E. Nature 176, 1042–1049
(1955).6. David, A. et al. J. Cell Biol. 197, 45–57 (2012).7. Fricker, M., Hollinshead, M., White, N. & Vaux, D. J. Cell
Biol. 136, 531–544 (1997).8. Ingolia, N.T., Lareau, L.F. & Weissman, J.S. Cell 147, 789–802
(2011).9. Aviner, R., Geiger, T. & Elroy-Stein, O. Genes Dev. 27,
1834–1844 (2013).10. Fritsch, C. et al. Genome Res. 22, 2208–2218 (2012).11. Crappé, J. et al. BMC Genomics 14, 648 (2013).
Competing financial interestsThe authors declare no competing financial interests.
80S
E P A
Nucleus Transcription
Splicing
Export
5’ cap 3’
AAAA 3’
3’E P A
MHC class I
Cytoplasm
ER
TAP
80S
5’ cap
EJC
E P A 5’ cap
80S
AAAA 3’
PTC
5’ cap AAAA 3’
Degradation
Golgi
5’
Figure 1 | Model for antigenic MHC class I peptide substrate generation. Full-length endogenous or exogenous protein degradation (purple elements, cytoplasm) only contributes mildly to the MHC class I ligandome. A substantial portion of MHC class I substrates is produced during the pioneer rounds of mRNA translation (preceding NMD) via a noncanonical translation mechanism in the nucleus (red elements) or perinuclear cytoplasm (blue elements) and using noncanonical mRNA as templates (that is, prespliced or defective mRNA), enabling more direct presentation to the MHC class I pathway. Nuclear RNA-processing events marking mRNA for efficient cytoplasmic translation include 5′ cap formation, 3′ polyadenylation and pre-mRNA splicing (exons and introns are indicated as rectangles and lines, respectively). Splicing enhances translation via deposition of the exon-junction complex (EJC) (yellow and purple elements). Some EJC proteins are removed during mRNA export (purple elements), and others are removed during translation (yellow elements). mRNA inspection for the presence of PTCs (red hexagon) occurs during the pioneer round of mRNA translation (inset). If the ribosome has not traversed all of the EJC-marked exon-intron boundaries, this serves as a signal that any termination codon encountered is a PTC. Consequently, NMD is initiated. The red intronic square is representative of an intronic antigenic epitope giving rise to an MHC class I substrate upon translation of its nuclear prespliced mRNA. TAP, transporter associated with antigen-processing complex.
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