TES (protein)

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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)
TES
Identifiers
Symbol TESS
Entrez 26136
HUGO 14620
OMIM 606085
PDB 2iyb
UniProt Q9UGI8
Other data
Locus Chr. 7 q31.2

'TES' (aka 'Testin') is the protein product of the TESS gene, located on chromosome 7[1] in Homo sapiens. 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. [2] In addition to this, TES functions as a tumour suppressor [3]; the TESS gene is located within a fragile region of chromosome 7, and the promoter elements of the TESS 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.

Contents

[edit] Domain Organisation

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 [4]. LIM domain consist of 2 zinc fingers separated by 2 hydrophobic amino acids (generally a Phenylalanine and then a Leucine).

[edit] Binding Partners

TES does not appear to be an enzyme; rather it is a protein that mediates/regulates cullular functions via Protein:protein interactions. Pull down experiments [5] reveal that TES has putative interactions mediated by the indicated domain:

Partner Domain ref Method
mENA/VASP LIM3 Coutts et al, Garalov et al & Boeda, Briggs et al Yeast two Hybrid, Pull-down assay, Structure, ITC
Arp7a  ??? Coutts et al Yeast two Hybrid
Zyxin LIM1 Coutts et al & Garalov et al Yeast two Hybrid, Pull-down assay
Actin PET? Garalov et al Pull-down assay
α-Actinin PET? Garalov et al Pull-down assay
Paxillin PET? Garalov et al Pull-down assay

Garalov et al showed that the interaction between TES & zyxin were direct, using recombinant proteins expressed in E.coli.

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.

[edit] TES as a tumour suppressor

In December 2007, Boeda, Briggs et al[6] showed that the 3rd 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 comprising of a Phenylalanine residue, followed by 4 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 the 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 3rd 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 is cancer Metastasis. TES is conversely, often not produced in cancer cells. It is possible that a drug designed to mimic TESs interaction with Mena could be used to prevent Metastasis and thus development of secondary tumours in cancer patients. Given this, the work was widely reported in the British press (the work was carried out by Cancer Research UK), [7][8] [9] and also in the international press [10][11].

[edit] Conformational Change

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.

Garalov 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.

[edit] Phenotype

In RNAi experiments, cells that had impaired TES expression showed an inability to correctly organise their focal adhesions and actin stress fibres.

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.

[edit] References

  1. ^ Tatarelli C, et al, (2002) Characterization of the Human TESTIN Gene Localized in the FRA7G Region at 7q31.2, Genomics, 68, P 1-12
  2. ^ Coutts, AS et al, TES is a novel focal adhesion protein with a role in cell spreading, (2003) Journal of Cell Science 116, 897-906
  3. ^ Drusco, A et al, Knockout mice reveal a tumour suppressor function for Testin, (2005) Proc Natl Acad Sci U S A 102 p10947–10951.
  4. ^ Dawida, IB et al, LIM domains: multiple roles as adapters and functional modifiers in protein interactions, (1998), Trends in Genetics, 14, pp156-162
  5. ^ Garalov, B et al, The conformational state of TES regulates its Zyxin-dependent recruitment to focal adhesions (2003), JCB
  6. ^ Boeda, B., Briggs, D.C. et al, TES, a specific Mena interacting partner, breaks the rules for EVH1 binding, (2007), Molecular Cell, 28, pp1071-1082
  7. ^ BBC NEWS | Health | Drug target to stop cancer spread
  8. ^ New light shed on how cancers spread | Science | The Guardian
  9. ^ Sunday Express: The World's Greatest Newspaper :: Posts
  10. ^ Cancer's spreading mechanism found - Science - Specials - smh.com.au
  11. ^ Clue found to checking cancer spread-Health/Sci-The Times of India