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利用者:Ashot~jawiki/核局在化シグナル

核局在化シグナル（かくきょくざいかしぐなる、英：Nuclear localization signal、NLS）とは、核輸送により細胞質から細胞核へ輸送されるタンパク質がもつアミノ酸配列である。このシグナルは1つもしくは複数の、タンパク質表面に露出した、正電荷をもつアミノ酸であるリジンやアルギニンを含む。異なる核局在タンパク質でも、同じNLSをもつと考えられている。これは核にあるタンパク質を細胞外へ輸送する核外排出シグナルとは反対の働きをもつ。

核局在化シグナルの種類[編集]

Classical NLS[編集]

Classical NLSはmonopartiteかbipartiteのいずれかに分類することができる。初めて見つかったNLSはSV40 large T抗原 (en : SV40 large T-antigen) の配列PKKKRKVである（monopartite）[1] 。nucleoplasminのNLSであるKR[PAATKKAGQA]KKKKは普遍的なbipartiteなシグナルの原型である。この普遍的なbipartiteなシグナルは、約10アミノ酸のスペーサーで隔てられた、塩基性アミノ酸のクラスターで構成される[2] 。この2つのシグナルはimportin αにより認識される。Importin αはbipartite NLSそのものを含むのだが、そのNLSはimportin βにより認識される。後者のimportinこそが真の核移行のメディエーターと考えることができる。

Chelsky et al. proposed the consensus sequence K-K/R-X-K/R for monopartite NLSs.[2] A Chelsky sequence may, therefore, be part of the downstream basic cluster of a bipartite NLS. Makkerh et al. carried out comparative mutagenesis on the nuclear localization signals of SV40 T-Antigen (monopartite), C-myc (monopartite) and nucleoplasmin (bipartite), and showed amino acid features common to all three. Notably the role of neutral and acidic amino acids was shown for the first time in contributing to the efficiency of the NLS.[3]

Non-classical NLS[編集]

There are many other types of NLS, such as the acidic M9 domain of hnRNP A1, the sequence KIPIK in yeast transcription repressor Matα2, and the complex signals of U snRNPs. Most of these NLSs appear to be recognized directly by specific receptors of the importin β family without the intervention of an importin α-like protein.[4]

A signal that appears to be specific for the massively produced and transported ribosomal proteins,[5][6] seems to come with a specialized set of importin β-like nuclear import receptors.[7]

Recently a class of NLSs known as PY-NLSs has been proposed, originally by Lee et al.[8] This PY-NLS motif, so named because of the proline-tyrosine amino acid pairing in it, allows the protein to bind to Importin β2 (also known as transportin or karyopherin β2), which then translocates the cargo protein into the nucleus. The structural basis for the binding of the PY-NLS contained in Importin β2 has been determined and an inhibitor of import designed.[9]

NLSの発見[編集]

細胞のDNAを隔てる核膜の存在は、真核生物の定義である。核膜は、DNA複製やRNAへの転写といった核内でおこるイベントを細胞質で起こるタンパク質への翻訳から隔てるものである。核内で必要とされるタンパク質は、何かしらの機構でそこへ輸送されなければならない。核タンパク質が核に局在することができるのを最初に証明した実験はJohn Gurdonによるもので、アフリカツメガエルの卵の細胞質に注入した精製核タンパク質が核に蓄積することを示した。これらの実験はThese experiments were part of a series which subsequently led to studies of nuclear reprogramming, directly relevant to stem cell research.

The presence of several million pore complexes in the oocyte nuclear membrane and the fact that they appeared to admit many different molecules (insulin, bovine serum albumin, gold nanoparticles) led to the view that the pores are open channels and nuclear proteins freely enter the nucleus through the pore and must accumulate by binding to DNA or some other nuclear component. In other words there was thought to be no specific transport mechanism.

This view was shown to be incorrect by Dingwall and Laskey in 1982. Using a protein called Nucleoplasmin, the archetypal ‘molecular chaperone’, they identified a domain in the protein which acted as a signal for nuclear entry.[10] This work stimulated research in the area and two years later the first NLS was identified in SV40 Large T-antigen (or SV40, for short). However a functional NLS could not be identified in another nuclear protein simply on the basis of similarity to the SV40 NLS. In fact only a small percentage of cellular (non-viral) nuclear proteins contained a sequence similar to the SV40 NLS. A detailed examination of Nucleoplasmin identified a sequence with two elements made up of basic amino acids separated by a spacer arm. One of these elements was similar to the SV40 NLS but was not able to direct a protein to the cell nucleus when attached to a non-nuclear reporter protein. Both elements are required.[11] This kind of NLS has become known as a bipartite classical NLS. The bipartite NLS is now known to represent the major class of NLS found in cellular nuclear proteins[要出典] and structural analysis has revealed how the signal is recognized by a receptor (importin α) protein[12] (the structural basis of some monopartite NLSs is also known[13]). Many of the molecular details of nuclear protein import are now known. This was made possible by the demonstration that nuclear protein import is a two step process; the nuclear protein binds to the nuclear pore complex in a process which does not require energy. This is followed by an energy dependent translocation of the nuclear protein through the channel of the pore complex.[14][15] By establishing the presence of two distinct steps in the process the possibility of identifying the factors involved was established and led on to the identification of the importin family of NLS receptors and the GTPase Ran.

Mechanism of nuclear import[編集]

Proteins gain entry into the nucleus through the nuclear envelope. The nuclear envelope consists of concentric membranes, the outer and the inner membrane. These are the gateways to the nucleus. The envelope consist of pores or large nuclear complexes.

A protein translated with a NLS will bind strongly to importin (aka karyopherin), and together, the complex will move through the nuclear pore. At this point, Ran-GTP will bind to the importin-protein complex, and its binding will cause the importin to lose affinity for the protein. The protein is released, and now the Ran-GTP/importin complex will move back out of the nucleus through the nuclear pore. A GTPase activating protein (GAP) in the cytoplasm hydrolyzes the Ran-GTP to GDP, and this causes a conformational change in Ran, ultimately reducing its affinity for importin. Importin is released and Ran-GDP is recycled back to the nucleus where a Guanine nucleotide exchange factor (GEF) exchanges its GDP back for GTP.

See also[編集]

References[編集]

  1. ^ Kalderon D, Roberts BL, Richardson WD, Smith AE (1984). “A short amino acid sequence able to specify nuclear location”. Cell 39 (3 Pt 2): 499–509. doi:10.1016/0092-8674(84)90457-4. PMID 6096007. 
  2. ^ a b Dingwall C, Robbins J, Dilworth SM, Roberts B, Richardson WD (Sep 1988). “The nucleoplasmin nuclear location sequence is larger and more complex than that of SV-40 large T antigen”. J Cell Biol. 107 (3): 841–9. doi:10.1083/jcb.107.3.841. PMC 2115281. PMID 3417784. http://www.jcb.org/cgi/pmidlookup?view=long&pmid=3417784. 
  3. ^ Makkerh JP, Dingwall C, Laskey RA (August 1996). “Comparative mutagenesis of nuclear localisation signals reveals the importance of neutral and acidic amino acids”. Curr Biol. 6 (8): 1025–7. doi:10.1016/S0960-9822(02)00648-6. PMID 8805337. http://linkinghub.elsevier.com/retrieve/pii/S0960-9822(02)00648-6. 
  4. ^ Mattaj IW, Englmeier L (1998). “Nucleocytoplasmic transport: the soluble phase”. Annu Rev Biochem. 67 (1): 265–306. doi:10.1146/annurev.biochem.67.1.265. PMID 9759490. 
  5. ^ Timmers AC, Stuger R, Schaap PJ, van 't Riet J, Raué HA (June 1999). “Nuclear and nucleolar localisation of Saccharomyces cerevisiae ribosomal proteins S22 and S25”. FEBS Lett. 452 (3): 335–40. doi:10.1016/S0014-5793(99)00669-9. PMID 10386617. http://linkinghub.elsevier.com/retrieve/pii/S0014-5793(99)00669-9. 
  6. ^ Garrett RA, Douthwate SR, Matheson AT, Moore PB, Noller HF (2000). The Ribosome: Structure, Function, Antibiotics, and Cellular Interactions. ASM Press. ISBN 978-1-55581-184-6. http://estore.asm.org/viewItemDetails.asp?ItemID=277 
  7. ^ Rout MP, Blobel G, Aitchison JD (May 1997). “A distinct nuclear import pathway used by ribosomal proteins”. Cell 89 (5): 715–25. doi:10.1016/S0092-8674(00)80254-8. PMID 9182759. http://linkinghub.elsevier.com/retrieve/pii/S0092-8674(00)80254-8. 
  8. ^ Lee BJ, Cansizoglu AE, Süel KE, Louis TH, Zhang Z, Chook YM (August 2006). “Rules for nuclear localisation sequence recognition by karyopherin beta 2”. Cell 126 (3): 543–58. doi:10.1016/j.cell.2006.05.049. PMID 16901787. http://linkinghub.elsevier.com/retrieve/pii/S0092-8674(06)00910-X. 
  9. ^ Cansizoglu AE, Lee BJ, Zhang ZC, Fontoura BM, Chook YM (May 2007). “Structure-based design of a pathway-specific nuclear import inhibitor”. Nature Structural & Molecular Biology 14 (5): 452–4. doi:10.1038/nsmb1229. PMID 17435768. 
  10. ^ Dingwall C, Sharnick SV, Laskey RA (September 1982). “A polypeptide domain that specifies migration of nucleoplasmin into the nucleus”. Cell 30 (2): 449–58. doi:10.1016/0092-8674(82)90242-2. PMID 6814762. http://linkinghub.elsevier.com/retrieve/pii/0092-8674(82)90242-2. 
  11. ^ Dingwall C, Laskey RA (December 1991). “Nuclear targeting sequences--a consensus?”. Trends in Biochemical Sciences 16 (12): 478–81. doi:10.1016/0968-0004(91)90184-W. PMID 1664152. 
  12. ^ Conti E, Kuriyan J (March 2000). “Crystallographic analysis of the specific yet versatile recognition of distinct nuclear localisation signals by karyopherin alpha”. Structure (London, England : 1993) 8 (3): 329–38. PMID 10745017. http://linkinghub.elsevier.com/retrieve/pii/st8315. 
  13. ^ Conti E, Uy M, Leighton L, Blobel G, Kuriyan J (July 1998). “Crystallographic analysis of the recognition of a nuclear localisation signal by the nuclear import factor karyopherin alpha”. Cell 94 (2): 193–204. doi:10.1016/S0092-8674(00)81419-1. PMID 9695948. http://linkinghub.elsevier.com/retrieve/pii/S0092-8674(00)81419-1. 
  14. ^ Dingwall C, Robbins J, Dilworth SM, Roberts B, Richardson WD (September 1988). “The nucleoplasmin nuclear location sequence is larger and more complex than that of SV-40 large T antigen”. The Journal of Cell Biology 107 (3): 841–9. doi:10.1083/jcb.107.3.841. PMC 2115281. PMID 3417784. http://www.jcb.org/cgi/pmidlookup?view=long&pmid=3417784. 
  15. ^ Newmeyer DD, Forbes DJ (March 1988). “Nuclear import can be separated into distinct steps in vitro: nuclear pore binding and translocation”. Cell 52 (5): 641–53. doi:10.1016/0092-8674(88)90402-3. PMID 3345567. http://linkinghub.elsevier.com/retrieve/pii/0092-8674(88)90402-3. 

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