Nuclear translation: what is the evidence?

Abstract

Recently, several reports have been published in support of the idea that protein synthesis occurs in both the nucleus and the cytoplasm. This proposal has generated a great deal of excitement because, if true, it would mean that our thinking about the compartmentalization of cell functions would have to be re-evaluated. The significance and broad implications of this phenomenon require that the experimental evidence used to support it be carefully evaluated. Here, we critique the published evidence in support of, or in opposition to, the question of whether translation occurs in the nucleus. Arguments in support of nuclear translation focus on three issues: (1) the presence of translation factors and ribosomal components in the nucleus, and their recruitment to sites of transcription; (2) amino acid incorporation in isolated nuclei and in nuclei under conditions that should not permit protein import; and (3) the fact that nuclear translation would account for observations that are otherwise difficult to explain. Arguments against nuclear translation emphasize the absence (or low abundance) from nuclei of many translation factors; the likely inactivity of nascent ribosomes; and the loss of translation activity as nuclei are purified from contaminating cytoplasm. In our opinion, all of the experiments on nuclear translation published to date lack critical controls and, therefore, are not compelling; also, traditional mechanisms can explain the observations for which nuclear translation has been invoked. Thus, while we cannot rule out nuclear translation, in the absence of better supporting data we are reluctant to believe it occurs.

FIGURE 1.

Nonsense mediated decay of mRNAs. The monitoring of mRNAs for premature termination codons (PTCs) is mediated by ribosomes during the “pioneering round of translation.” Several proteins associate with sequences near exon/intron borders of the spliced mRNA, forming the exon junction complex (EJC). Some EJC proteins are removed during export whereas others are removed during translation and thus serve as indicators of whether a particular region of the mRNA has been translated. (A) If no EJC proteins are on the mRNA when the pioneering ribosome encounters a chain termination codon, this codon is read as a normal termination signal, at the 3′ end of the coding region. (B) If the ribosome has not traversed all exon/intron borders, EJC protein(s) remain on the mRNA and serve as an indication that any termination codon encountered is a PTC. As a consequence, nonsense-mediated decay (NMD) is initiated, resulting in destruction of the mRNA. The pioneering round of translation is shown here as happening in the cytoplasm (or the perinuclear cytoplasm), and with the released proteins being imported into the nucleus.

FIGURE 1.

Nonsense mediated decay of mRNAs. The monitoring of mRNAs for premature termination codons (PTCs) is mediated by ribosomes during the “pioneering round of translation.” Several proteins associate with sequences near exon/intron borders of the spliced mRNA, forming the exon junction complex (EJC). Some EJC proteins are removed during export whereas others are removed during translation and thus serve as indicators of whether a particular region of the mRNA has been translated. (A) If no EJC proteins are on the mRNA when the pioneering ribosome encounters a chain termination codon, this codon is read as a normal termination signal, at the 3′ end of the coding region. (B) If the ribosome has not traversed all exon/intron borders, EJC protein(s) remain on the mRNA and serve as an indication that any termination codon encountered is a PTC. As a consequence, nonsense-mediated decay (NMD) is initiated, resulting in destruction of the mRNA. The pioneering round of translation is shown here as happening in the cytoplasm (or the perinuclear cytoplasm), and with the released proteins being imported into the nucleus.

FIGURE 2.

A model for coupling nuclear splicing to cytoplasmic translation. The presence of PTCs in specific exons of particular mRNAs can alter the splicing of the pre-mRNA; like NMD, this nonsense-associated alternative splicing (NAS) uses translation to detect the PTC. If the pioneering round of translation occurs in the perinuclear cytoplasm rather than in the nucleus, a signal indicating the presence of the PTC must be transmitted from the ribosome back to the nuclear splicing machinery. (A) This model proposes that a splicing factor (denoted X) binds to an splicing enhancer sequence (ESE) in a nonoptimal exon of a pre-mRNA, allowing for inclusion of the exon in the mRNA, and that this factor remains bound to the ESE as the mRNA exits the nucleus. In the cytoplasm, the ESE binding factor is transferred to the pioneering ribosome and released only when this ribosome reaches the termination codon at the 3′ end of the coding region. Release of the factor allows it to recycle back to the nucleus, to support further rounds of splicing. (B) If the pioneering ribosome encounters a PTC (a termination codon to the 5′ side of an EJC), it is unable to release the factor, thereby retarding return of the factor to the nucleus. Recognition of the PTC also results in NMD; the presence of the factor on the pioneering ribosome may even enhance NMD, resulting in “Super NMD”, as is observed with TCR-β mRNA. (C) A reduced level of the ESE binding factor would result in skipping of the ESE-containing exon during splicing. This cytoplasmic feedback model predicts that in cells expressing both the wild-type and PTC-containing alleles of the same gene, splicing of both types of pre-mRNAs would be affected.

FIGURE 2.

A model for coupling nuclear splicing to cytoplasmic translation. The presence of PTCs in specific exons of particular mRNAs can alter the splicing of the pre-mRNA; like NMD, this nonsense-associated alternative splicing (NAS) uses translation to detect the PTC. If the pioneering round of translation occurs in the perinuclear cytoplasm rather than in the nucleus, a signal indicating the presence of the PTC must be transmitted from the ribosome back to the nuclear splicing machinery. (A) This model proposes that a splicing factor (denoted X) binds to an splicing enhancer sequence (ESE) in a nonoptimal exon of a pre-mRNA, allowing for inclusion of the exon in the mRNA, and that this factor remains bound to the ESE as the mRNA exits the nucleus. In the cytoplasm, the ESE binding factor is transferred to the pioneering ribosome and released only when this ribosome reaches the termination codon at the 3′ end of the coding region. Release of the factor allows it to recycle back to the nucleus, to support further rounds of splicing. (B) If the pioneering ribosome encounters a PTC (a termination codon to the 5′ side of an EJC), it is unable to release the factor, thereby retarding return of the factor to the nucleus. Recognition of the PTC also results in NMD; the presence of the factor on the pioneering ribosome may even enhance NMD, resulting in “Super NMD”, as is observed with TCR-β mRNA. (C) A reduced level of the ESE binding factor would result in skipping of the ESE-containing exon during splicing. This cytoplasmic feedback model predicts that in cells expressing both the wild-type and PTC-containing alleles of the same gene, splicing of both types of pre-mRNAs would be affected.

FIGURE 2.

A model for coupling nuclear splicing to cytoplasmic translation. The presence of PTCs in specific exons of particular mRNAs can alter the splicing of the pre-mRNA; like NMD, this nonsense-associated alternative splicing (NAS) uses translation to detect the PTC. If the pioneering round of translation occurs in the perinuclear cytoplasm rather than in the nucleus, a signal indicating the presence of the PTC must be transmitted from the ribosome back to the nuclear splicing machinery. (A) This model proposes that a splicing factor (denoted X) binds to an splicing enhancer sequence (ESE) in a nonoptimal exon of a pre-mRNA, allowing for inclusion of the exon in the mRNA, and that this factor remains bound to the ESE as the mRNA exits the nucleus. In the cytoplasm, the ESE binding factor is transferred to the pioneering ribosome and released only when this ribosome reaches the termination codon at the 3′ end of the coding region. Release of the factor allows it to recycle back to the nucleus, to support further rounds of splicing. (B) If the pioneering ribosome encounters a PTC (a termination codon to the 5′ side of an EJC), it is unable to release the factor, thereby retarding return of the factor to the nucleus. Recognition of the PTC also results in NMD; the presence of the factor on the pioneering ribosome may even enhance NMD, resulting in “Super NMD”, as is observed with TCR-β mRNA. (C) A reduced level of the ESE binding factor would result in skipping of the ESE-containing exon during splicing. This cytoplasmic feedback model predicts that in cells expressing both the wild-type and PTC-containing alleles of the same gene, splicing of both types of pre-mRNAs would be affected.