The effect of protein context on nuclear location signal function

Roberts, Bruce; Richardson, William D.; Smith, Alan E.

doi:10.1016/0092-8674(87)90500-9

Cited by 206 publications

(124 citation statements)

References 26 publications

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“…Epitope tags have proven reliable for the analysis of nuclear targeting since a given NLS can function at a variety of positions in a protein (Roberts et al, 1987), however insertion of tags could potentially disrupt normal protein targeting. To circumvent this potential problem we characterized BRCA1 subcellular localization with both N-terminal and C-terminal tags as well as non-tagged proteins.…”

Section: Discussionmentioning

confidence: 99%

Differential subcellular localization, expression and biological toxicity of BRCA1 and the splice variant BRCA1-Δ11b

et al. 1997

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The mechanism of BRCA1 tumor suppression in human breast and ovarian cells is the focus of intense investigation. In this report, full length BRCA1 (230 kDa) introduced into cells with CMV promoter constructs was nuclear when transgene expression was low whereas high expression resulted in cytoplasmic accumulation, aberrant nuclear and cell morphology. A nuclear localization signal (NLS) was mapped to BRCA1 amino acid positions 262 ± 570. We describe a splice variant, BRCA1-D11b, missing the majority of exon 11 including the NLS. Exogenous BRCA1-D11b (110 kDa) was cytoplasmic and, unlike the full-length protein, overexpression of the protein encoded by the variant did not appear to be toxic. RNA probe titrations and RT ± PCR demonstrated that BRCA1 and D11b transcripts are coexpressed in a wide variety of cells and tissues. Interestingly, BRCA1-D11b message was greatly reduced or absent in several breast and ovarian tumor lines relative to exon 11 transcripts and a D9,10 splice variant. Taken together our results suggest that fulllength BRCA1 and BRCA1-D11b may have distinct roles in cell growth regulation and tumorigenesis.

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Section: Discussionmentioning

confidence: 99%

Differential subcellular localization, expression and biological toxicity of BRCA1 and the splice variant BRCA1-Δ11b

et al. 1997

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“…The two identical NLSs KRRR of S6 represent a relatively strong signal because when this tetrapeptide is fused to 13-galactosidase it directs the hybrid protein exclusively into the nucleus (Figure 3). Efficiency of nuclear import can be influenced by the protein context of the NLS (Roberts et al, 1987;Nelson and Silver, 1989), by distant sequences (Gao and Knipe, 1992) …”

Section: Differential Import Efficiencies Of the Nlss Of S6mentioning

confidence: 99%

Nuclear and nucleolar targeting of human ribosomal protein S6.

Schmidt

Lipsius

Kruppa

1995

MBoC

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Chimeric proteins were constructed to define the nuclear localization signals (NLSs) of human ribosomal protein S6. The complete cDNA sequence, different cDNA fragments and oligonucleotides of the human ribosomal proteins S6, respectively, were joined to the 5' end of the entire LacZ gene of Escherichia coli by using recombinant techniques. The hybrid genes were transfected into L cells, transiently expressed, and the intracellular location of the fusion proteins was determined by their ,B-galactosidase activity. Three NLSs were identified in the C-terminal half of the S6 protein. Deletion mutagenesis demonstrated that a single NLS is sufficient for targeting the corresponding S6-,Bgalactosidase chimera into the nucleus. Removal of all three putative NLSs completely blocked the nuclear import of the resulting S6-,3-galactosidase fusion protein, which instead became evenly distributed in the cytoplasm. Chimeras containing deletion mutants of S6 with at least one single NLS or unmodified S6 accumulated in the nucleolus. Analysis of several constructs reveals the existence of a specific domain that is essential but not sufficient for nucleolar accumulation of S6.

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“…Reiterated NLSs, of similar but not identical sequence, are present in some proteins, as in the polyoma virus T antigen (Richardson et al, 1986) and the glucocorticoid receptor (Picard & Yamamoto, 1987). In the case of redundant or bipartite signals, mutations at one of them may impair but do not abolish the transport process, suggesting a modular structure of the NLS (Robbins et al, 1991), Although the NLS is sufficient for a protein to be transported into the cell nucleus, it may not be independent of the sequence context in which it is located because the NLS needs to be accessible for interaction with the transport machinery (Roberts et al, 1987), and this accessibility can be modulated by intramolecular (Henkel et al, 1992) or intermolecular masking (for review, see Schmitz et al, 1991).…”

Section: Introductionmentioning

confidence: 99%

Complex structure of the nuclear translocation signal of influenza virus polymerase PA subunit

Nieto

Luna

Bárcena

et al. 1994

Journal of General Virology

101

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The protein regions involved in the nuclear translocation of the influenza virus PA polymerase subunit have been identified by deletion analysis of the protein expressed from a recombinant simian virus 40. Two regions seem to play a role in the process: region 1 (amino acids 124 to 139) and region II (amino acids 186 to 247), A nucleoplasmin-like nuclear translocation signal (NLS) has been identified in region I and an additional NLS appears to be present in region II, although no consensus targeting sequence can be detected. Alteration in any of the regions identified by short deletions completely prevented nuclear transport, whereas elimination of the regions I or I! by large amino-or carboxy-terminal deletions did not prevent nuclear targeting of the truncated protein. In addition, a point mutation at position 154 completely eliminated nuclear transport. A fl-galactosidase fusion protein containing the 280 amino acid terminal region of the PA protein was partially transported to the nucleus and mutant PA proteins with a cytoplasmic phenotype could not be rescued by superinfection with influenza virus. These results suggest that the PA protein contains a functional nuclear targeting region which is required in influenza virus infection, with two independent NLSs, one in region I and the other in region II.

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The effect of protein context on nuclear location signal function

Cited by 206 publications

References 26 publications

Differential subcellular localization, expression and biological toxicity of BRCA1 and the splice variant BRCA1-Δ11b

Differential subcellular localization, expression and biological toxicity of BRCA1 and the splice variant BRCA1-Δ11b

Nuclear and nucleolar targeting of human ribosomal protein S6.

Complex structure of the nuclear translocation signal of influenza virus polymerase PA subunit

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