|Testis derived transcript (3 LIM domains)|
The 3rd LIM domain of TES (green) bound to the EVH1 domain of Mena (blue). The EVH1 FPPPP binding site (magenta) is blocked by some residues from TES (yellow).
|Symbols||TES (; TESS; TESS-2)|
Testin also known as TESS is a protein that in humans is encoded by the TES gene located on chromosome 7. TES is a 47 kDa protein composed of 421 amino acids found at focal adhesions and is thought to have a role in regulation of cell motility. In addition to this, TES functions as a tumour suppressor. The TES gene is located within a fragile region of chromosome 7, and the promoter elements of the TES gene have been shown to be susceptible to methylation - this prevents the expression of the TES protein. TES came to greater prominence towards the end of 2007 as a potential mechanism for its tumour suppressor function was published.
Tes is composed of the following domains:
|Domain Name||Boundaries||Domain type|
|Cysteine rich domain||1 - 90||No Homology|
|PET domain||90 - 200||PET domain - no structure|
|Linker .||201 - 233||no domain|
|LIM1||234 - 300||LIM domain|
|LIM2||300 - 365||LIM domain|
|LIM3||366 - 421||LIM domain|
The structures of the Cysteine rich domain and the PET domain are not known. LIM domains, however, are known as modulators of protein interactions. LIM domain consist of 2 zinc fingers separated by 2 hydrophobic amino acids (generally a phenylalanine and then a leucine).
TES does not appear to be an enzyme; rather it is a protein that mediates/regulates cellular functions via Protein:protein interactions. Pull down experiments reveal that TES has putative interactions mediated by the indicated domain:
|mENA/VASP||LIM3||||Yeast two Hybrid, Pull-down assay, Structure, ITC|
|Arp7a||???||||Yeast two Hybrid|
|Zyxin||LIM1||||Yeast two Hybrid, Pull-down assay|
Some of the potential binding partners (Zyxin, mENA) can be found in focal adhesion complexes; the range of binding partners indicates a potential role for TES in-between 'privileged' Actin polymerisation and focal adhesion contacts to the extracellular matrix. This tallies with the observation that GFP-tagged TES can be seen at focal adhesions.
TES as a tumour suppressor
In December 2007, Boeda, Briggs et al. showed that the third LIM domain of TES displaces Mena from its usual subcellular positions (focal adhesions or the cell leading edge). The ENA/VASP protein family (of which Mena is a member) are anchored to specific proteins within the cell by a peptide motif consisting of a phenylalanine residue, followed by four proline residues - known as a FPPPP motif. It is the EVH1 domains of VASP/EVL proteins that directly contact the FPPPP motif. The precise architecture of TES:MENA binding was revealed by X-ray crystallography, and showed that the 3rd LIM domain of TES covered up the FPPPP binding site within Menas EVH1 domain. Isothermal titration calorimetry showed that TES has a greater affinity for Mena than its canonical FPPPP ligand, as presented in the focal adhesion protein zyxin. Using microscopy it was shown that either over-expression of GFP-tagged TES, or just the tagged third LIM domain displaced Mena from focal adhesions and reduced mean cell velocity.
These finding were significant given that Mena is often over-expressed in cancer cells, and is thought to be partly responsible for cancer cell motility, and therefore a factor in cancer metastasis. TES is conversely often not produced in cancer cells. It is possible that a drug designed to mimic TES's interaction with Mena could be used to prevent metastasis and thus development of secondary tumours in cancer patients. The work was widely reported in the British press (the work was carried out by Cancer Research UK), and also in the international press.
Based on the observations that:
- Mammalian cell derived TES binding Zyxin
- E. coli produced recombinant TES (rTES) does not bind Zyxin
- An rTES construct composed of residues 201-421 (i.e., the linker and all 3 LIM domains) does bind Zyxin
- The above rTES construct binds an N-terminal rTES construct, consisting of the cysteine rich and PET domains - IE, the two halves of TES interact with each other.
Garvalov et al. propose that TES exists in two conformational states: A 'closed' state where the N & C halves of TES interact, obscuring the Zyxin binding site in LIM1, and an 'open' state where the Zyxin binding site is accessible and the two halves no-longer interact in the same fashion, if at all. The regulatory mechanism switching between the two states is not presently fully understood.
In gene knockout experiments, transgenic mice lacking both copies of the TES gene displayed an increased susceptibility to tumour formation when challenged with a carcinogen. Mice retaining the TES gene were less susceptible: thus, TES is a tumour suppressor gene.
- PDB 2iyb; Boëda B, Briggs DC, Higgins T, Garvalov BK, Fadden AJ, McDonald NQ, Way M (December 2007). "Tes, a specific Mena interacting partner, breaks the rules for EVH1 binding". Mol. Cell 28 (6): 1071–82. doi:10.1016/j.molcel.2007.10.033. PMID 18158903.
- Tatarelli C, Linnenbach A, Mimori K, Croce CM (August 2000). "Characterization of the human TESTIN gene localized in the FRA7G region at 7q31.2". Genomics 68 (1): 1–12. doi:10.1006/geno.2000.6272. PMID 10950921.
- Coutts AS, MacKenzie E, Griffith E, Black DM (March 2003). "TES is a novel focal adhesion protein with a role in cell spreading". J. Cell. Sci. 116 (Pt 5): 897–906. doi:10.1242/jcs.00278. PMID 12571287.
- Drusco A, Zanesi N, Roldo C, Trapasso F, Farber JL, Fong LY, Croce CM (August 2005). "Knockout mice reveal a tumor suppressor function for Testin". Proc. Natl. Acad. Sci. U.S.A. 102 (31): 10947–51. doi:10.1073/pnas.0504934102. PMC 1182460. PMID 16033868.
- Dawid IB, Breen JJ, Toyama R (April 1998). "LIM domains: multiple roles as adapters and functional modifiers in protein interactions". Trends Genet. 14 (4): 156–62. doi:10.1016/S0168-9525(98)01424-3. PMID 9594664.
- Garvalov BK, Higgins TE, Sutherland JD, Zettl M, Scaplehorn N, Köcher T, Piddini E, Griffiths G, Way M (April 2003). "The conformational state of Tes regulates its zyxin-dependent recruitment to focal adhesions". J. Cell Biol. 161 (1): 33–9. doi:10.1083/jcb.200211015. PMC 2172870. PMID 12695497.
- "Drug target to stop cancer spread". BBC News. 2010-10-06. Retrieved 2010-10-06.
- Jha A (2007-12-28). "New light shed on how cancers spread". Science. The Guardian. Retrieved 2010-10-06.
- Fletcher V (2007-12-28). "The Cancer 'Life Saver'". UK News. Express.co.uk. Retrieved 2010-10-06.
- "Cancer's spreading mechanism found". Science. The Sydney Morning Herald. 2007-12-29. Retrieved 2010-10-06.
- "Clue found to checking cancer spread". The Times of India. 2007-12-29. Retrieved 2010-10-06.