Nuclear export signal

A nuclear export signal (NES) is a short target peptide containing 4 hydrophobic residues in a protein that targets it for export from the cell nucleus to the cytoplasm through the nuclear pore complex using nuclear transport. It has the opposite effect of a nuclear localization signal, which targets a protein located in the cytoplasm for import to the nucleus. The NES is recognized and bound by exportins.

NESs serve several vital cellular functions. They assist in regulating the position of proteins within the cell. Through this NESs effect transcription and several other nuclear functions that are essential to proper cell function.[1] The export of many types of RNA from the nucleus is required for proper cellular function. The NES determines what type of pathway the varying types of RNA may use to exit the nucleus and perform their function and the NESs may effect the directionality of molecules exiting the nucleus.[2]

Structure

Computer analysis of known NESs found the most common spacing of the hydrophobic residues to be LxxxLxxLxL, where "L" is a hydrophobic residue (often leucine) and "x" is any other amino acid; the spacing of these hydrophobic residues may be explained by examination of known structures that contain an NES, as the critical residues usually lie in the same face of adjacent secondary structures within a protein, which allows them to interact with the exportin.[3] Ribonucleic acid (RNA) is composed of nucleotides, and thus, lacks the nuclear export signal to move out of the nucleus. As a result, most forms of RNA will bind to a protein molecule to form a ribonucleoprotein complex to be exported from the nucleus.

Eukaryotic Linear Motif resource defines the NES motif for exportin within a single entry, TRG_NES_CRM1_1. The single-letter amino acid sequence pattern of NES, in regular expression format, is:[4]

([DEQ].{0,1}[LIM].{2,3}[LIVMF][^P]{2,3}[LMVF].[LMIV].{0,3}[DE])|
([DE].{0,1}[LIM].{2,3}[LIVMF][^P]{2,3}[LMVF].[LMIV].{0,3}[DEQ])

In the above expression, LIMVF are all hydrophobic residues, while DEQ are hydrophilic aspartic acid, glutamic acid, and glutamine. In human language, this is an extension of the "common pattern" that includes hydrophilic residues surrounding it as well as slight variations in the length of xxx and xx fragments seen above.

Mechanism

Nuclear export first begins with the binding of Ran-GTP (a G-protein) to exportin. This causes a shape change in exportin, increasing its affinity for the export cargo. Once the cargo is bound, the Ran-exportin-cargo complex moves out of the nucleus through the nuclear pore. GTPase activating proteins (GAPs) then hydrolyze the Ran-GTP to Ran-GDP, and this causes a shape change and subsequent exportin release. Once no longer bound to Ran, the exportin molecule loses affinity for the nuclear cargo as well, and the complex falls apart. Exportin and Ran-GDP are recycled to the nucleus separately, and guanine exchange factor (GEF) in the nucleus switches the GDP for GTP on Ran.

mRNA Transport

During the processing phase of mRNA export, mRNA is covered with RNA-binding proteins, this yields ribonucleoprotein particles (mRNPs). Through the process, the correct assortment of proteins is necessary for the mRNPs eventual export. The heterodimeric export receptor yeast Mex67-Mtr2/human NXF1-NXT1 is the NES responsible for signalling the completion of the mRNP construction. This signal mediates the mRNPs movement out of the nuclear envelope through the NPC. The assembled mRNPs are directed out of the nucleus and into the cytoplasm by two DEAD-box ATPases. On the nucleus side is yeast Sub2/human UAP56 and on the cytoplasm side is the ATPase yeast Dbp5/human DDX19.[5]

Chemotherapy

The process of nuclear export is responsible for some resistance to chemotherapy drugs. By limiting a cells nuclear export activity it may be possible to reverse this resistance. By inhibiting CRM1, the export receptor, export through the nuclear envelope may be slowed. Survivin is a NES that inhibits cellular apoptosis. It interacts with the mitotic spindles during cellular division. Due to the usually rapid proliferation of tumour cells, survivin is more expressed during the presence of cancer. The level of survivin correlates to how resistance to chemotherapy a cancerous cell is and how likely that cell is to replicate again. By producing antibodies to target the NES survivin, apoptosis of cancerous cells can be increased.[6]

Examples

NES signals were first discovered in the human immunodeficiency virus type 1 (HIV-1) Rev protein and cAMP-dependent protein kinase inhibitor (PKI). The karyopherin receptor CRM1 has been identified as the export receptor for leucine-rich NESs in several organisms and is an evolutionarily conserved protein. The export mediated by CRM1 can be effectively inhibited by the fungicide leptomycin B (LMB), providing excellent experimental verification of this pathway.[7]

Other proteins of various functions have also been experimentally inhibited of the NES signal such as the cyto-skeletal protein actin, which functions include cell motility and growth. The use of LBM as a NES inhibitor proved successful for actin resulting in accumulation of the protein within the nucleus, concluding universal functionality of NES throughout various protein functional groups.[8]

Regulation

Not all NES substrates are constitutively exported from the nucleus, meaning that CRM1-mediated export is a regulated event. Several ways of regulating NES-dependent export have been reported. These include masking/unmasking of NESs, phosphorylation and even disulfide bond formation as a result of oxidation.

The binding of NES to the export receptor of a protein gives the universal export function of NES an individually specified activation of export to each protein. Studies of specified NES amino acid sequences for particular proteins show the possibility of blocking the NES activation of one protein with an inhibitor for that amino acid sequence while other proteins of the same nucleus remain unaffected.[9]

NESbase

NESbase is a database of proteins with experimentally verified leucine-rich nuclear export signals (NES). The verification is performed by, among others, Technical University of Denmark Center for Biological Sequence Analysis and University of Copenhagen Department of Protein Chemistry. Every entry in its database includes information whether nuclear export signals were sufficient for export or if it was only mediated transport by CRM1 (exportin).[10]

gollark: Okay, so learn that and prove it so [REDACTED].
gollark: It would be useful if it was proved or disproved, and one of the things it would be useful for is Project PURPOSELESS CHAIRMANSHIPS, you see.
gollark: Yes, I know, I'm saying it's necessary for the successful completion of Project PURPOSELESS CHAIRMANSHIPS.
gollark: Yes, we get those frequently.
gollark: Please do, it would really help with Project PURPOSELESS CHAIRMANSHIPS?

References
  1. Fukuda, Makoto; Asano, Shiro; Nakamura, Takahiro; Adachi, Makoto; Yoshida, Minoru; Yanagida, Mitsuhiro; Nishida, Eisuke (November 1997). "CRM1 is responsible for intracellular transport mediated by the nuclear export signal". Nature. 390 (6657): 308–311. Bibcode:1997Natur.390..308F. doi:10.1038/36894. ISSN 0028-0836. PMID 9384386.
  2. Li, Zhengguo; Kearse, Michael G.; Huang, Chuan (2019-01-02). "The nuclear export of circular RNAs is primarily defined by their length". RNA Biology. 16 (1): 1–4. doi:10.1080/15476286.2018.1557498. ISSN 1547-6286. PMC 6380329. PMID 30526278.
  3. la Cour T, Kiemer L, Mølgaard A, Gupta R, Skriver K, Brunak S (June 2004). "Analysis and prediction of leucine-rich nuclear export signals". Protein Eng. Des. Sel. 17 (6): 527–36. doi:10.1093/protein/gzh062. PMID 15314210.
  4. "ELM - Detail for TRG_NES_CRM1_1". elm.eu.org. Retrieved 10 April 2019.
  5. Xie, Yihu; Ren, Yi (November 2019). "Mechanisms of nuclear mRNA export: A structural perspective". Traffic. 20 (11): 829–840. doi:10.1111/tra.12691. ISSN 1398-9219. PMC 7074880. PMID 31513326.
  6. El-Tanani, Mohamed; Dakir, El-Habib; Raynor, Bethany; Morgan, Richard (2016-03-14). "Mechanisms of Nuclear Export in Cancer and Resistance to Chemotherapy". Cancers. 8 (3): 35. doi:10.3390/cancers8030035. ISSN 2072-6694. PMC 4810119. PMID 26985906.
  7. Fukuda, Makoto; Asano, Shiro; Nakamura, Takahiro; Adachi, Makoto; Yoshida, Minoru; Yanagida, Mitsuhiro; Nishida, Eisuke (1997-11-20). "CRM1 is responsible for intracellular transport mediated by the nuclear export signal". Nature. 390 (6657): 308–311. Bibcode:1997Natur.390..308F. doi:10.1038/36894. ISSN 0028-0836. PMID 9384386.
  8. Wada, Atsushi; Fukuda, Makoto; Mishima, Masanori; Nishida, Eisuke (1998-03-16). "Nuclear export of actin: a novel mechanism regulating the subcellular localization of a major cytoskeletal protein". The EMBO Journal. 17 (6): 1635–1641. doi:10.1093/emboj/17.6.1635. ISSN 0261-4189. PMC 1170511. PMID 9501085.
  9. Rowe, Thomas C.; Ostrov, David; Dawson, Jana L.; Pernazza, Danielle; Lawrence, Nicholas J.; Sullivan, Daniel M. (2013-11-15). "Targeting The Nuclear Export Signal In Multiple Myeloma". Blood. 122 (21): 1925. doi:10.1182/blood.V122.21.1925.1925. ISSN 0006-4971.
  10. Tanja la Cour; Ramneek Gupta; Kristoffer Rapacki; Karen Skriver; Flemming M. Poulsen; Søren Brunak (2003). "NESbase version 1.0: a database of nuclear export signals". Nucleic Acids Research. 31 (1): 393–396. doi:10.1093/nar/gkg101. PMC 165548. PMID 12520031.
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