Analysis of CRM1-Dependent Nuclear Export in Permeabilized Cells

Kehlenbach, Ralph H.; Port, Sarah A.

doi:10.1007/978-1-4939-3530-7_30

Cited by 2 publications

(1 citation statement)

References 26 publications

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“…These two approaches are often combined, for example when using biochemical assays to validate predictions made from structural data [11,16,34,35]. On the other hand, cellular assays, based on the localization of reporter proteins, have been widely used to identify novel NESs or transport factors, and to test the effect of potential CRM1 inhibitors [22,39,40], but they are not routinely used to test assumptions made from structural and in vitro data. In fact, a possible correlation between the affinity of NESs for CRM1 and their export activity has only recently been evaluated [31].…”

Section: Discussionmentioning

confidence: 99%

Using a Simple Cellular Assay to Map NES Motifs in Cancer-Related Proteins, Gain Insight into CRM1-Mediated NES Export, and Search for NES-Harboring Micropeptides

Sendino

Omaetxebarria

Rodríguez

2020

IJMS

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The nuclear export receptor CRM1 (XPO1) recognizes and binds specific sequence motifs termed nuclear export signals (NESs) in cargo proteins. About 200 NES motifs have been identified, but over a thousand human proteins are potential CRM1 cargos, and most of their NESs remain to be identified. On the other hand, the interaction of NES peptides with the “NES-binding groove” of CRM1 was studied in detail using structural and biochemical analyses, but a better understanding of CRM1 function requires further investigation of how the results from these in vitro studies translate into actual NES export in a cellular context. Here we show that a simple cellular assay, based on a recently described reporter (SRVB/A), can be applied to identify novel potential NESs motifs, and to obtain relevant information on different aspects of CRM1-mediated NES export. Using cellular assays, we first map 19 new sequence motifs with nuclear export activity in 14 cancer-related proteins that are potential CRM1 cargos. Next, we investigate the effect of mutations in individual NES-binding groove residues, providing further insight into CRM1-mediated NES export. Finally, we extend the search for CRM1-dependent NESs to a recently uncovered, but potentially vast, set of small proteins called micropeptides. By doing so, we report the first NES-harboring human micropeptides.

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Section: Discussionmentioning

confidence: 99%

Using a Simple Cellular Assay to Map NES Motifs in Cancer-Related Proteins, Gain Insight into CRM1-Mediated NES Export, and Search for NES-Harboring Micropeptides

Sendino

Omaetxebarria

Rodríguez

2020

IJMS

View full text Add to dashboard Cite

show abstract

Using a simple cellular assay to map NES motifs in cancer-related proteins, gain insight into CRM1-mediated NES export, and search for NES-harbouring micropeptides

Sendino

Omaetxebarria

Rodriguez

2020

Preprint

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The nuclear export receptor CRM1 (XPO1) recognizes and binds specific sequence motifs termed nuclear export signals (NESs) in cargo proteins. About 200 NES motifs have been identified, but over a thousand human proteins are potential CRM1 cargos, and most of their NESs remain to be identified. On the other hand, the interaction of NES peptides with the 'NES-binding groove' of CRM1 has been studied in detail using structural and biochemical analyses, but a better understanding of CRM1 function requires further investigation of how the results from these in vitro studies translate into actual NES export in a cellular context. Here we show that a simple cellular assay, based on a recently described reporter (SRVB/A), can be applied to identify novel potential NESs motifs, and to obtain relevant information on different aspects of CRM1-mediated NES export. Using cellular assays, we first map 19 new sequence motifs with nuclear export activity in 14 cancer-related proteins that are potential CRM1 cargos. Next, we investigate the effect of mutations in individual NES-binding groove residues, providing further insight into CRM1-mediated NES export. Finally, we extend the search for CRM1-dependent NESs to a recently uncovered, but potentially vast, set of small proteins called micropeptides. By doing so, we report the first NES-harbouring human micropeptides.

show abstract

Analysis of CRM1-Dependent Nuclear Export in Permeabilized Cells

Cited by 2 publications

References 26 publications

Using a Simple Cellular Assay to Map NES Motifs in Cancer-Related Proteins, Gain Insight into CRM1-Mediated NES Export, and Search for NES-Harboring Micropeptides

Using a Simple Cellular Assay to Map NES Motifs in Cancer-Related Proteins, Gain Insight into CRM1-Mediated NES Export, and Search for NES-Harboring Micropeptides

Using a simple cellular assay to map NES motifs in cancer-related proteins, gain insight into CRM1-mediated NES export, and search for NES-harbouring micropeptides

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