Exosome complex

The exosome complex (or PM/Scl complex, often just called the exosome) is a multi-protein complex, capable of degrading various types of RNAs. Exosome complexes can be found in both eukaryotic cells and archaea, while in bacteria a simpler complex called the "degradosome" carries out similar functions.

The core of the complex has a six-membered ring structure, to which other proteins are attached. In eukaryotic cells, it is present in the cytoplasm, nucleus and especially the nucleolus, although different proteins interact with the complex in these compartments, in order to regulate the RNA degradation activity of the complex to substrates specific for these cell compartments. Substrates of the exosome include messenger RNA, ribosomal RNA, and many species of small RNAs. The exosome has an exoribonucleolytic function, meaning it degrades RNA starting at one side (the so-called 3' end in this case), rather than cleaving the RNA at specific sites.

Although no causative relation between the complex and any disease is known, several proteins in the complex are the target of autoantibodies in patients with specific autoimmune diseases (especially the PM/Scl overlap syndrome) and some antimetabolitic chemotherapies for cancer function by blocking the activity of the complex.

Discovery

The exosome was first discovered as an RNase in 1997 in the budding yeast "Saccharomyces cerevisiae", an often used model organism.cite journal
author=Mitchell "et al."
title=The Exosome: A Conserved Eukaryotic RNA Processing Complex Containing Multiple 3′→5′ Exoribonucleases
journal=Cell
year=1997
volume=91
issue=4
pages=457–466
pmid=9390555
doi=10.1016/S0092-8674(00)80432-8] Not long after, in 1999, it was realized that the exosome was in fact the yeast equivalent of an already described complex in human cells, the so-called "PM/Scl complex", which had been identified as an autoantigen in patients with certain autoimmune diseases years earlier (see below). [cite journal
author=Allmang "et al."
title=The yeast exosome and human PM-Scl are related complexes of 3' --> 5' exonucleases
journal=Genes and Development
year=1999
volume=13
issue=16
pages=2148–58
pmid=10465791
doi=10.1101/gad.13.16.2148] Purification of this "PM/Scl complex" allowed the identification of more human exosome proteins and eventually the characterization of all components in the complex. [cite journal
author=Brouwer "et al."
title=Three novel components of the human exosome
year=2001
journal=Journal of Biological Chemistry
volume=276
pages=6177–84
pmid=11110791
doi=10.1074/jbc.M007603200] [cite journal
author=Chen "et al."
title=AU binding proteins recruit the exosome to degrade ARE-containing mRNAs
year=2001
journal=Cell
volume=107
pages=451–64
pmid=11719186
doi=10.1016/S0092-8674(01)00578-5] In 2001, the increasing amount of genome data that had become available allowed the prediction of exosome proteins in archaea, although it would take another 2 years before the first exosome complex from an archaeal organism was first purified. [cite journal
author=Koonin "et al."
title=Prediction of the archaeal exosome and its connections with the proteasome and the translation and transcription machineries by a comparative-genomic approach
journal=Genome Research
year=2001
volume=11
issue=2
pages=240–52
pmid=11157787
doi=10.1101/gr.162001] [cite journal
author=Evguenieva-Hackenberg "et al."
title=An exosome-like complex in Sulfolobus solfataricus
journal=EMBO Reports
year=2003
volume=4
issue=9
pages=889–93
pmid=12947419
doi=10.1038/sj.embor.embor929 ]

tructure

Core proteins

The core of the complex has a ring structure consisting of six proteins that all belong to the same class of RNases, the RNase PH-like proteins.cite journal
author=Schilders "et al."
title=Cell and molecular biology of the exosome: how to make or break an RNA
journal=International review of cytology
year=2006
volume=251
pages=159–208
pmid=16939780
doi=10.1016/S0074-7696(06)51005-8] In archaea there are two different PH-like proteins (called Rrp41 and Rrp42), each present three times in an alternating order. Eukaryotic exosome complexes have six different proteins that form the ring structure. [cite journal
author=Lorentzen "et al."
title=The archaeal exosome core is a hexameric ring structure with three catalytic subunits
journal=Nature Structural & Molecular Biology
year=2005
volume=12
pages=575–81
pmid=15951817
doi=10.1038/nsmb952] cite journal
author=Shen "et al."
title=A view to a kill: structure of the RNA exosome
journal=Cell
year=2006
volume=127
pages=1093–5
pmid=17174886
doi=10.1016/j.cell.2006.11.035] Of these six eukaryotic proteins, three resemble the archaeal Rrp41 protein and the other three proteins are more similar to the archaeal Rrp42 protein. [cite journal
author=Raijmakers "et al."
title=Protein-protein interactions between human exosome components support the assembly of RNase PH-type subunits into a six-membered PNPase-like ring
journal=Journal of Molecular Biology
year=2002
volume=323
pages=653–63
pmid=12419256
doi=10.1016/S0022-2836(02)00947-6]

Located on top of this ring are three proteins that have an S1 RNA binding domain (RBD). Two proteins in addition have a K-homology (KH) domain. In eukaryotes, three different "S1" proteins are bound to the ring, whereas in archaea either one or two different "S1" proteins can be part of the exosome (although there are always three S1 subunits attached to the complex). [cite journal
author=Walter "et al."
title=Characterization of native and reconstituted exosome complexes from the hyperthermophilic archaeon Sulfolobus solfataricus
journal=Molecular Microbiology
year=2006
volume=62
pages=1076–89
pmid=17078816
doi=10.1111/j.1365-2958.2006.05393.x ]

This ring structure is very similar to that of the proteins RNase PH and PNPase. In bacteria, the protein RNase PH, which is involved in tRNA processing, forms a hexameric ring consisting of six identical RNase PH proteins. [cite journal
author=Ishii "et al."
title=Crystal structure of the tRNA processing enzyme RNase PH from Aquifex aeolicus
journal=Journal of Biological Chemistry
volume=278
pages=32397–404
year=2003
pmid=12746447
doi=10.1074/jbc.M300639200 ] In the case of PNPase, which is a phosphorolytic RNA degrading protein found in bacteria and the chloroplasts and mitochondria of some eukaryotic organisms, two RNase PH domains and both an S1 and KH RNA binding domain are part of a single protein, which forms a trimeric complex that adopts a structure almost identical to that of the exosome. [cite journal
author= Symmons "et al."
title=A duplicated fold is the structural basis for polynucleotide phosphorylase catalytic activity, processivity, and regulation
journal=Structure
year=2000
volume=8
pages=1215–26
pmid=11080643
doi=10.1016/S0969-2126(00)00521-9 ] Because of this high similarity in both protein domains and structure, these complexes are thought to be evolutionary related and have a common ancestor. [cite journal
author=Lin-Chao "et al."
title=The PNPase, exosome and RNA helicases as the building components of evolutionarily-conserved RNA degradation machines
journal=Journal of Biomedical Science
year=2007
volume=14
pages=523–32
pmid=
doi=10.1007/s11373-007-9178-y] In bacteria, a separate RNase PH protein exists that is involved in transfer RNA processing, which has been shown to adopt a similar six-membered ring structure, but in this case consisting of 6 identical protein subunits. [cite journal
author= Harlow "et al."
title=Crystal structure of the phosphorolytic exoribonuclease RNase PH from Bacillus subtilis and implications for its quaternary structure and tRNA binding
journal=Protein Science
year=2004
volume=13
pages=668–77
pmid=14767080
doi=10.1110/ps.03477004 ] The RNase PH-like exosome proteins, PNPase and RNase PH all belong to the RNase PH family of RNases and are phosphorolytic exoribonucleases, meaning they use inorganic phosphate to remove nucleotides from the 3' end of RNA molecules.

Associated proteins

Besides these nine core exosome proteins, two other proteins often associate with the complex in eukaryotic organisms. One of these is Rrp44, a hydrolytic RNase, which belongs to the RNase R family of hydrolytic exoribonucleases (nucleases that use water to cleave the nucleotide bonds). In yeast, Rrp44 is associated with "all" exosome complexes and has a crucial role in the activity of the yeast exosome complex. [cite journal
title=The exosome subunit Rrp44 plays a direct role in RNA substrate recognition
author= Schneider "et al."
journal=Molecular Cell
year=2007
volume=27
pages=324–31
pmid=17643380
doi=10.1016/j.molcel.2007.06.006] Remarkably, while a human homologue of the protein exists, no evidence has been found to date that its human homologue is even associated with the human exosome complex. protein family. [cite journal
author=Mian "et al."
title=Comparative sequence analysis of ribonucleases HII, III, II PH and D
journal=Nucleic Acids Research
year=1997
volume=25
pages=3187–3195
pmid=9241229
doi=10.1093/nar/25.16.3187] The protein PM/Scl-100 is most commonly part of exosome complexes in the nucleus of cells, but can form part of the cytoplasmic exosome complex as well. [cite journal
author=Raijmakers "et al."
title=The exosome, a molecular machine for controlled RNA degradation in both nucleus and cytoplasm
journal=European Journal of Cell Biology
year=2004
volume=83
pages=175–83
pmid=15346807
doi=10.1078/0171-9335-00385]

Regulatory proteins

Apart from these two tightly-bound protein subunits, many proteins interact with the exosome complex in both the cytoplasm and nucleus of cells. These loosely-associated proteins may regulate the activity and specificity of the exosome complex. In the cytoplasm, the exosome interacts with AU rich element (ARE) binding proteins (e.g. KRSP and TTP), which can promote or prevent degradation of mRNAs. The nuclear exosome associates with RNA binding proteins (e.g. MPP6 in humans and Rrp47/C1D in humans and yeast) that control ribosomal RNA processing.

In addition to single proteins, other protein complexes interact with the exosome. One of those is the cytoplasmic Ski complex, which includes an RNA helicase (Ski2) and is involved in mRNA degradation. [cite journal
author=Wang "et al."
title=Domain interactions within the Ski2/3/8 complex and between the Ski complex and Ski7p
journal=RNA
year=2005
volume=11
pages=1291–302
pmid=16043509
doi=10.1261/rna.2060405 ] In the nucleus, the processing of rRNA and snoRNA by the exosome is mediated by the TRAMP complex, which contains both RNA helicase (Mtr4) and polyadenylation (Trf4) activity. [cite journal
author=LaCava "et al."
title=RNA degradation by the exosome is promoted by a nuclear polyadenylation complex
journal=Cell
year=2005
volume=121
pages=713–24
pmid=15935758
doi=10.1016/j.cell.2005.04.029 ]

Function

Enzymatic function

As stated above, the exosome complex contains many proteins that contain ribonuclease domains. These are all 3'-5' exoribonuclease domains, meaning the enzymes degrade RNA molecules from their 3' end. The complex contains both phosphorolytic exoribonucleases (the RNase PH-like proteins) and, in eukaryotes, also hydrolytic exoribonucleases (the RNase R and RNase D domain proteins). The phosphorolytic enzymes use inorganic phosphate to cleave the phosphodiester bonds - releasing nucleotide disphosphates. The hydrolytic enzymes use water to hydrolyse these bonds - releasing nucleotide monosphosphates).

In archaea, the Rrp41 subunit of the complex is a phosphorolytic exoribonuclease. Three copies of this protein are present in the ring and this enzyme is responsible for the activity of the complex. In eukaryotes, none of the human RNase PH subunits have retained this catalytic activity, meaning the core ring structure of the human exosome has no enzymatically-active protein. [cite journal
author=Liu "et al."
title=Erratum: Reconstitution, activities, and structure of the eukaryotic RNA exosome
journal= Cell
year=2007
volume=131
pages=188–189
pmid=17174896
doi=10.1016/j.cell.2007.09.019 ] In yeast this is compensated by one of the associated hydrolytic enzymes, Rrp44, which is responsible for most of the exosome ribonuclease activitycite journal
author=Dziembowski "et al."
title=A single subunit, Dis3, is essentially responsible for yeast exosome core activity
journal=Nature Structural & Molecular Biology
year=2007
volume=14
pages=15–22
pmid=17173052
doi=10.1038/nsmb1184 ] , but this particular hydrolytic subunit may be restricted to yeast, as no protein homologous to Rrp44 is present in the human (and archaeal) exosome complex. [cite journal
author=Liu "et al."
title=Reconstitution, activities, and structure of the eukaryotic RNA exosome
journal= Cell
year=2006
volume=127
pages=1223–37
pmid=17174896
doi=10.1016/j.cell.2006.10.037 ] [cite journal
author=Lorentzen "et al."
title=Structural basis of 3' end RNA recognition and exoribonucleolytic cleavage by an exosome RNase PH core
journal=Molecular Cell
year=2005
volume=20
pages=473–81
pmid=16285928
doi=10.1016/j.molcel.2005.10.020]

Still, in both human and yeast, another hydrolytic enzyme can be associated with the complex (Rrp6), which contributes to the activity of the yeast exosome and is solely responsible for the activity of the human complex. Although originally the S1 domain proteins were thought to have 3'-5' hydrolytic exoribonuclease activity as well, this existence of this activity has recently been questioned and these proteins might have just a role in binding substrates prior to their degradation by the complex.

ubstrates

The exosome is involved in the degradation and processing of a wide variety of RNA species. In the cytoplasm of cells, it is involved in the turn-over of messenger RNA (mRNA) molecules. The complex can degrade mRNA molecules that have been tagged for degradation because they contain errors, through interactions with proteins from the nonsense mediated decay or non-stop decay pathways. Alternatively, mRNAs are degraded as part of their normal turnover. Several proteins that stabilize or destabilize mRNA molecules through binding to AU-rich elements in the 3' UTR of mRNAs interact with the exosome complex. [cite journal
author= LeJeune "et al."
title=Nonsense-mediated mRNA decay in mammalian cells involves decapping, deadenylating, and exonucleolytic activities
journal=Molecular Cell
year=2003
volume=12
pages=675–87
pmid=14527413
doi=10.1016/S1097-2765(03)00349-6 ] [cite journal
author= Wilson "et al."
title=A genomic screen in yeast reveals novel aspects of nonstop mRNA metabolism
journal=Genetics
year=2007
pmid=17660569
doi=10.1534/genetics.107.073205
volume=177
pages=773 ] [cite journal
author= Lin "et al."
title=Localization of AU-rich element-containing mRNA in cytoplasmic granules containing exosome subunits
journal=Journal of Biological Chemistry
year=2007
volume=282
pages=19958–68
pmid=17470429
doi=10.1074/jbc.M702281200 ] In the nucleus, the exosome is required for the correct processing of several small nuclear RNA molecules.cite journal
author= Allmang "et al."
title=Functions of the exosome in rRNA, snoRNA and snRNA synthesis
journal=EMBO Journal
year=1999
volume=18
pages=5399–410
pmid=10508172
doi=10.1093/emboj/18.19.5399 ] Finally, the nucleolus is the compartment where the majority of the exosome complexes are found. There it plays a role in the processing of the 5.8S ribosomal RNA (the first identified function of the exosome) and of several small nucleolar RNAs. [cite journal
author= Schilders "et al."
title=MPP6 is an exosome-associated RNA-binding protein involved in 5.8S rRNA maturation
journal=Nucleic Acids Research
year=2005
volume=33
pages=6795–804
pmid=16396833
doi=10.1093/nar/gki982 ]

Although most cells have other enzymes that can degrade RNA, either from the 3' or 5' end of the RNA, the exosome complex is essential for cell survival. When the expression of exosome proteins is artificially reduced or stopped, for example by RNA interference, growth stops and the cells eventually die. Both the core proteins of the exosome complex, as well as the two main associated proteins, are essential proteins. [cite journal
author=van Dijk "et al."
title=Human cell growth requires a functional cytoplasmic exosome, which is involved in various mRNA decay pathways
journal=RNA
year=2007
volume=13
pages=1027–35
pmid=17545563
doi=10.1261/rna.575107] Bacteria do not have an exosome complex, however, similar functions are performed by a simpler complex that includes the protein PNPase, called the "degradosome". [cite journal |author=Carpousis AJ
title=The Escherichia coli RNA degradosome: structure, function and relationship in other ribonucleolytic multienzyme complexes
journal=Biochem. Soc. Trans.
volume=30
issue=2
pages=150–5
year=2002
pmid=12035760
url=http://www.biochemsoctrans.org/bst/030/0150/bst0300150.htm
doi=10.1042/BST0300150]

Disease

Autoimmunity

The exosome complex is the target of autoantibodies in patients that suffer from various autoimmune diseases. These autoantibodies are mainly found in people that suffer from the PM/Scl overlap syndrome, an autoimmune disease in which patients have symptoms from both scleroderma and either polymyositis or dermatomyositis. [cite journal
author= J.E. Pope
title=Scleroderma overlap syndromes
journal=Current Opinion in Rheumatology
year=2002
volume=14
pages=704–10
pmid=12410095
doi=10.1097/00002281-200211000-00013 ] Autoantibodies can be detected in the serum of patients by a variety of assays. In the past, the most commonly used methods were double immunodiffusion using calf thymus extracts, immunofluorescence on HEp-2 cells or immunoprecipitation from human cell extracts. In immunoprecipitation assays with sera from anti-exosome positive sera, a distinctive set of proteins is precipitated. Already years before the exosome complex was identified, this pattern was termed the "PM/Scl complex". [cite journal
author=Gelpi "et al."
title=Identification of protein components reactive with anti-PM/Scl autoantibodies
journal=Clinical and Experimental Immunology
volume=81
year=1991
pages=59–64
pmid=2199097] Immunofluorescence using sera from these patients usually shows a typical staining of the nucleolus of cells, which sparked the suggestion that the antigen recognized by autoantibodies might be important in ribosome synthesis. [cite journal
author=Targoff "et al."
title=Nucleolar localization of the PM-Scl antigen
journal=Arthritis & Rheumatism
volume=28
year=1985
pages=226–30
pmid=3918546
doi=10.1002/art.1780280221 ] More recently, recombinant exosome proteins have become available and these have been used to develop line immunoassays (LIAs) and enzyme linked immunosorbent assays (ELISAs) for detecting these antibodies.

In these diseases, antibodies are mainly directed against two of the proteins of the complex, called "PM/Scl-100" (the RNase D like protein) and "PM/Scl-75" (one of the RNase PH like proteins from the ring) and antibodies recognizing these proteins are found in approximately 30% of patients with the PM/Scl overlap syndrome. [cite journal
author=Raijmakers "et al."
title=PM-Scl-75 is the main autoantigen in patients with the polymyositis/scleroderma overlap syndrome
journal=Arthritis & Rheumatism
year=2004
volume=50
pages=565–9
pmid=14872500
doi=10.1002/art.20056 ] Although these two proteins are the main target of the autoantibodies, other exosome subunits and associated proteins (like C1D) can be targeted in these patients. [cite journal
author= Brouwer "et al."
title=Autoantibodies directed to novel components of the PM/Scl complex, the human exosome.
journal=Arthritis Research
year=2002
volume=4
pages=134–8
pmid=11879549
doi=10.1186/ar389 ] [cite journal
author= Schilders "et al."
title=C1D is a major autoantibody target in patients with the polymyositis-scleroderma overlap syndrome
journal=Arthritis & Rheumatism
year=2007
volume=56
pages=2449–54
pmid=17599775
doi=10.1002/art.22710 ] Currently, the most sensitive way to detect these antibodies is by using a peptide, derived from the PM/Scl-100 protein, as the antigen in an ELISA, instead of complete proteins. By this method, autoantibodies are found in up to 55% of patients with the PM/Scl overlap syndrome, but they can also be detected in patients suffering from either scleroderma, polymyositis or dermatomyositis alone. [cite journal
author=Mahler "et al."
title=Clinical evaluation of autoantibodies to a novel PM/Scl peptide antigen
journal=Arthritis Research & Therapy
year=2005
volume=7
pages=R704–13
pmid=15899056
doi=10.1186/ar1729]

As the autobodies are mainly found in patients that have characteristics of several different autoimmune diseases, the clinical symptoms of these patients can vary widely. The symptoms that are seen most often are the typical symptoms of the individual autoimmune diseases and include Raynaud's phenomenon, arthritis, myositis and scleroderma. [cite journal
author=Mahler "et al."
title=Novel aspects of autoantibodies to the PM/Scl complex: Clinical, genetic and diagnostic insights
journal=Autoimmunity Reviews
year=2007
volume=6
pages=432–7
pmid=17643929
doi=10.1016/j.autrev.2007.01.013 ] Treatment of these patients is symptomatic and is similar to treatment for the individual autoimmune disease, often involving either immunosuppressive or immunomodulating drugs. [cite journal
author= Jablonska "et al."
title=Scleromyositis: a scleroderma/polymyositis overlap syndrome
journal=Clinical Rheumatology
year=1998
volume=17
pages=465–7
pmid=9890673
doi=10.1007/BF01451281 ]

Cancer

The exosome has been shown to be inhibited by the antimetabolite drug fluorouracil, which is a drug used in chemotherapy treatment of cancer. It is one of the most successful drugs for treating solid tumors. In yeast cells treated with fluorouracil, defects were seen in the processing of ribosomal RNA, identical to those seen when the activity of the exosome was blocked by molecular biological strategies. Lack of correct ribosomal RNA processing is lethal to cells, explaining the antimetabolic effect of the drug. [cite journal
author=Lum "et al."
title=Discovering modes of action for therapeutic compounds using a genome-wide screen of yeast heterozygotes
journal=Cell
year=2004
volume=116
pages=121–37
pmid=14718172
doi=10.1016/S0092-8674(03)01035-3 ]

List of subunits

*note label|Note_B|A|AIn archaea several exosome proteins are present in multiple copies, to form the full core of the exosome complex.
*note label|Note_B|B|BAlthough the yeast protein Rrp44 is part of the yeast complex, its human homologue hDis3 has never been found to be associated with the human complex.

See also

* The proteasome, the main protein degrading machinery of cells
* The spliceosome, a complex involved in RNA splicing, that also contains an RNA binding ring structure

References

Further reading

* cite journal
author=Vanacova "et al."
title=The exosome and RNA quality control in the nucleus
journal=EMBO reports
year=2007
volume=8
pages=651–657
url=http://npg.nature.com/embor/journal/v8/n7/full/7401005.html
doi=10.1038/sj.embor.7401005
*cite journal
author=Houseley "et al."
title=RNA-quality control by the exosome
journal=Nature Reviews Molecular Cell Biology
year=2006
volume=7
pages=529–539
url=http://www.nature.com/nrm/journal/v7/n7/abs/nrm1964.html
doi=10.1038/nrm1964 –-- subscription required
*cite journal
author=Büttner "et al."
title=The exosome: a macromolecular cage for controlled RNA degradation
journal=Molecular Microbiology
year=2006
volume=61
pages=1372–1379
url=http://www.blackwell-synergy.com/doi/abs/10.1111/j.1365-2958.2006.05331.x
doi=10.1111/j.1365-2958.2006.05331.x
*cite journal
author=Lorentzen "et al."
title=The Exosome and the Proteasome: Nano-Compartments for Degradation
journal=Cell
year=2006
volume=125
pages=651–654
doi=10.1016/j.cell.2006.05.002
*cite journal
author=G.J.M. Pruijn
title=Doughnuts dealing with RNA
journal=Nature Structural & Molecular Biology
year=2005
volume=12
pages=562–564
url=http://www.nature.com/nsmb/journal/v12/n7/full/nsmb0705-562.html
doi=10.1038/nsmb0705-562

External links

* [http://www.rcsb.org/pdb/explore/explore.do?structureId=2NN6 Structure of the human exosome at the RCSB Protein Data Bank]
* [http://www.rcsb.org/pdb/explore.do?structureId=2JE6 Structure of an archael exosome at the RCSB Protein Data Bank]
* [http://www.rcsb.org/pdb/explore.do?structureId=2JEA Structure of an archael exosome bound to RNA at the RCSB Protein Data Bank]
* [http://www.rcsb.org/pdb/explore.do?structureId=2HBJ Structure of the yeast exosome protein Rrp6 at the RCSB Protein Data Bank]