Mediator complexes are variable at the evolutionary, compositional and conformational levels.[3] Figure 1 shows only one "snapshot" of what a particular complex might comprise,[b] but it is an inaccurate depiction of the conformationin vivo. During evolution, Mediator has complexified. The yeastSaccharomyces cerevisiae (a simpleeukaryote) is thought to have up to 21 subunits in the core Mediator (exclusive of the CDK module), while mammals have up to 26.
Individual subunits can be absent or replaced by other subunits under different conditions. Also, there are manyintrinsically disordered regions in Mediator proteins, which may contribute to the conformational flexibility seen both with and without other bound proteins or protein complexes. A more realistic model of Mediator without the CDK module is shown inFigure 2.[4]
Mediator is required for successfultranscription of genes by RNA polymerase II, and contacts the polymerase in thetranscription preinitiation complex.[3] A recent model showing the polymerase associating with Mediator without DNA is shown inFigure 3.[4] In addition to RNA polymerase II, Mediator must also associate with transcription factors and DNA; a model of such interactions is shown inFigure 4.[5] Note that the different morphologies of Mediator do not necessarily mean that a particular model is correct; rather those differences may reflect the flexibility of Mediator as it interacts with other molecules.[c] For example, after binding theenhancer and core promoter, the Mediator complex compositionally changes, dissociating thekinase module and associating withRNA polymerase II for transcriptional activation.[6]
Figure 5: Mediator complex architecture with focus on the disordered "spline" of MED14[9]
The yeast Mediator complex is approximately as massive as asmall subunit of a eukaryotic ribosome. The yeast Mediator has 25 subunits, while the mammalian Mediator is slightly larger.[3] Mediator comprises 4 main parts: the head, middle, tail, and the transiently associated CDK8 kinase module.[10]
Mediator subunits have manyintrinsically disordered regions called "splines", which may be important to allow the structural changes of Mediator that change the function of the complex.[3][d] Figure 5 shows the splines of theMED14 subunit connecting a large portion of the complex together while still allowing flexibility.[4][e]
Mediator complexes lacking a subunit have been found or produced. These smaller complexes can still function normally in some activity, but lack other capabilities.[3] This indicates a somewhat independent function of some of the subunits while composing the larger complex.
Another example of structural variability is seen in vertebrates, in which 3paralogues of subunits of thecyclin-dependent kinase (CDK) module have evolved by 3 independentgene duplication events followed by sequence divergence.[3]
There is a report that Mediator stably associates with a particular type ofnon-coding RNA, ncRNA-a.[11][f] These stable associations regulate gene expressionin vivo, and are prevented by mutations in MED12 that produce the human diseaseFG syndrome.[11] Thus, the structure of a Mediator complex can be augmented by RNA as well as proteinaceous transcription factors.[3]
Figure 3: Structural model of Mediator's tail and middle bound to RNA polymerase II[9]
Mediator was originally discovered because it was important for RNA polymerase II function, but it has many more functions than just interactions at the transcription start site.[3]
Figure 4: Model of Mediator with some transcription factors, Pol II and DNA
Mediator is a crucial component for transcription initiation. Mediator interacts with the pre-initiation complex, composed of RNA Polymerase II and general transcription factors TFIIB, TFIID, TFIIE, TFIIF, and TFIIH to stabilize and initiate transcription.[12] Studies of Mediator–RNA Pol II contacts in budding yeast showed the importance of TFIIB-Mediator contacts in the formation of the complex. Interactions of Mediator with TFIID in the initiation complex has been shown.[10]
The structure of a core Mediator (cMed) while associated with a core pre-initiation complex was elucidated.[12]
The preinitiation complex, which contains Mediator, transcription factors, a nucleosome[13][14][g] and RNA polymerase II, is important for positioning the polymerase for the start of transcription. Before RNA synthesis starts, the polymerase dissociates from Mediator. This is seemingly via phosphorylation of the polymerase by a kinase. Importantly, Mediator and transcription factors do not dissociate from the DNA when the polymerase begins transcription. Rather, the complex remains at the promoter to recruit another RNA polymerase to begin another round of transcription.[3][h]
There is some evidence to suggest that Mediator inSchizosaccharomyces pombe helps regulateRNA polymerase III (Pol III) transcripts oftRNAs.[15] An independent report confirmed Mediator specifically associating with Pol III inSaccharomyces cerevisiae.[16] Those authors also reported specific associations withRNA polymerase I and proteins involved in transcription elongation and RNA processing, supporting other evidence of Mediator's involvement in elongation and processing.[16]
Mediator is involved inchromatin looping, which brings distant regions of a chromosome into closer physical proximity.[3] The ncRNA-a mentioned above[11] is involved in such looping.[i]Enhancer RNAs (eRNAs) can function similarly.[3]
Involvement of Mediator in various human diseases has been reviewed.[24][25][26][27][28][29][30][31][32][33][34][excessive citations] Since inhibiting one interaction of a disease-causing signaling pathway with a subunit of Mediator may not inhibit general transcription needed for normal function, Mediator subunits are attractive candidates for therapeutic drugs.[3]
Mediator interactome in Saccharomyces cerevisiae[16]
Very gentle cell lysis in yeast followed by co-immunoprecipitation with an antibody to aMED17 has confirmed almost all previously reported or predicted interactions and revealed many previously unsuspected specific interactions of various proteins with Mediator.[16]
MicroRNAs help regulate the expression of many proteins. MED1 is targeted by miR-1, which is important in gene regulation in cancers.[35] Thetumor suppressor miR-137 also regulates MED1.[36]
Null mutants die early (embryonic day 11.5).[37][38] Investigatinghypomorphic mutants (which survive 2 days longer) found that placental defects were primarily lethal and that there were also defects in cardiac and hepatic development, but many other organs were normal.[38]
In mice,conditional mutations can be produced to affect only specific cells or tissues at specific times, so that the mouse can develop to adulthood to have its adultphenotype studied. In one case, MED1 was found to participate in controlling the timing of events ofmeiosis in male mice.[39] Conditional mutants inkeratinocytes differ in skin wound healing.[40] A conditional mutation in mice changed dentalepithelium intoepidermal epithelium, which caused hair to grow beside the incisors.[41]
Thesubunits form at least threestructurally distinct submodules. The head and the middle modulesinteract directly with RNA polymerase II, whereas the elongated tail moduleinteracts with gene-specific regulatoryproteins. Mediator containing the CDK8 module is less active than Mediator lacking this module in supportingtranscriptional activation.
The head module contains: MED6, MED8, MED11, SRB4/MED17, SRB5/MED18, ROX3/MED19, SRB2/MED20 and SRB6/MED22.
The middle module contains: MED1, MED4, NUT1/MED5, MED7, CSE2/MED9, NUT2/MED10, SRB7/MED21 and SOH1/MED31. CSE2/MED9 interacts directly with MED4.
The tail module contains: MED2, PGD1/MED3, RGR1/MED14, GAL11/MED15 and SIN4/MED16.
The CDK8 module contains: MED12, MED13, CCNC and CDK8. Individual preparations of the Mediator complex lacking one or more distinctsubunits have been variously termed ARC, CRSP, DRIP, PC2, SMCC and TRAP.
^Mediator is also referred to in scientific literature as thevitamin Dreceptor interacting protein (DRIP) coactivator complex and the thyroid hormone receptor-associated proteins (TRAP).
^However note that more recently it has been found that the CDK module and MED26 cannot be present concurrently in a complex.[3]
^The sharp bend in the DNA associated with thetranscription bubble is shown in the graphical abstract and first figure of thisresearch paper
^Some of those changes are diagrammed infigure 1 of the review article, which can be viewed in slightly larger form by clicking it at that site.
^Note that Med 17 (shown in blue) also has that sort of spline
^These non-codingactivating RNAs have not been mentioned yet in the ncRNA article as of 16 February 2017
^This is the +1 nucleosome, which "covers" the transcription start site during the preinitiation phase.
^This is diagrammed infigure 2 of the review article, which can be viewed in slightly larger form by clicking it at that site.
^This is diagrammed infigure 3 of the review article, which can be viewed in slightly larger form by clicking it at that site. That figure also shows Pol II disengaged from mediator,etc, which remains on the DNA
^Björklund S, Gustafsson CM (May 2005). "The yeast Mediator complex and its regulation".Trends in Biochemical Sciences.30 (5):240–4.doi:10.1016/j.tibs.2005.03.008.PMID15896741.
^Croce S, Chibon F (2015). "MED12 and uterine smooth muscle oncogenesis: State of the art and perspectives".Eur J Cancer.51 (12):1603–10.doi:10.1016/j.ejca.2015.04.023.PMID26037152.
^Schiano C, Casamassimi A, Rienzo M, de Nigris F, Sommese L, Napoli C (2014). "Involvement of Mediator complex in malignancy".Biochim Biophys Acta.1845 (1):66–83.doi:10.1016/j.bbcan.2013.12.001.PMID24342527.
^Schiano C, Casamassimi A, Vietri MT, Rienzo M, Napoli C (2014). "The roles of mediator complex in cardiovascular diseases".Biochim Biophys Acta.1839 (6):444–51.doi:10.1016/j.bbagrm.2014.04.012.PMID24751643.
^Napoli C, Sessa M, Infante T, Casamassimi A (2012). "Unraveling framework of the ancestral Mediator complex in human diseases".Biochimie.94 (3):579–87.doi:10.1016/j.biochi.2011.09.016.PMID21983542.
