ACRATA: a novel electron transfer domain associated to apoptosis and cancer
© Sanchez-Pulido et al; licensee BioMed Central Ltd. 2004
Received: 05 October 2004
Accepted: 29 December 2004
Published: 29 December 2004
Recently, several members of a vertebrate protein family containing a six trans-membrane (6TM) domain and involved in apoptosis and cancer (e.g. STEAP, STAMP1, TSAP6), have been identified in Golgi and cytoplasmic membranes. The exact function of these proteins remains unknown.
We related this 6TM domain to distant protein families using intermediate sequences and methods of iterative profile sequence similarity search.
Here we show for the first time that this 6TM domain is homolog to the 6TM heme binding domain of both the NADPH oxidase (Nox) family and the YedZ family of bacterial oxidoreductases.
This finding gives novel insights about the existence of a previously undetected electron transfer system involved in apoptosis and cancer, and suggests further steps in the experimental characterization of these evolutionarily related families.
A family of vertebrate proteins containing a six transmembrane domain (6TM) has been recently implicated in apoptosis and cancer. Up to date, this family contains four vertebrate members: STEAP, STAMP1/STEAP2, TIARP and TSAP6/pHyde. STEAP (six-transmembrane epithelial antigen of the prostate) was the first described member of this family and identified as a prostate-specific cell-surface antigen overexpressed in cancer, located at the cell-cell junction of the secretory epithelium of prostate, and found as well in both colon and bladder cancer cell lines [1, 2]. STAMP1 (six transmembrane protein of prostate 1), also known as STEAP2, also overexpressed in prostate cancer, has been located in the trans-Golgi network and shuttles to plasma membranes, which suggest a role in the secretory/endocytic pathways [3, 4]. TIARP (Tumor necrosis factor-alpha-induced adipose-related protein) is a cell surface protein induced by TNF-α and IL-6, probably implicated in resistance to insulin [5, 6]. TSAP6 (tumor suppressor activated pathway-6), also known as pHyde [7–10], is a p53 inducible protein which regulates apoptosis and the cell cycle via direct interaction with Nix (a pro-apoptotic Bcl-2 related protein) and Myt1 kinase (a negative regulator of the G2/M transition) . TSAP6 has been shown to be interacting with TCTP (Translationally controlled tumor protein) and could be implicated in its secretion .
To do the sequence analysis of the new domain we took advantage of the possibility of connecting distant protein families via intermediate sequences  and methods of iterative profile sequence similarity search: HMMer [20, 21] and PSI-BLAST  over the Uniprot 90% non redundant sequence database . We used NAIL to view and analyse the HMMer results . The alignment of transmembrane regions using standard substitution matrices might be inaccurate because of the different roles played by amino acids in globular proteins and in transmembrane media . In the case of the ACRATA domain, the regions to be aligned mostly consist of amino acids located in transmembrane regions (Fig. 1). For this reason, we used a method based on a hidden Markov model (HMMer hmmalign), which does not rely on a general substitution matrix [20, 21], using as a guide both the transmembrane predictions from TMHMM [26, 27] and the results of multiple sequence alignment using T-Coffee [28, 29]. The genomic neighborhood of the bacterial sequences (YedZ family) was analyzed to find potentially related genes in operons using STRING [17, 18].
To investigate the consistency of our results we performed iterative database searches using the PSI-BLAST program . We used as query the most conserved region of the ACRATA domain in E. coli YedZ protein (residues 71–166). These searches detected all of the ACRATA domain-containing families after 15 iterations (using a cut-off of E = 0.005 for the inclusion of retrieved sequences in the profile). None of these profile searches retrieved new unrelated sequences, and reciprocal searches produced convergent results. Therefore, we have concluded that ACRATA is a previously undetected, conserved domain that is commonly found in members of the STEAP, YedZ and Nox protein families. The similarity between these proteins was suspected before . However, no alignment, domain definition or substantial statistical evidence was provided to demonstrate the evolutionary relationships of these proteins.
The complete conservation of two histidines in all ACRATA domain containing proteins (Fig. 1) indicates that the STEAP protein family could bind at least an heme group, as was previously experimentally characterized for Nox and YedZ families [13, 16], or for other analogous proteins such as cytochromes .
Although the mechanism of action of the ACRATA domain could be the same in all the proteins containing it, its variable cellular role is made conspicuous by the different effects produced by the modifications in the expression of the corresponding genes. For example, the knock-out of the whole YedYZ operon seems not to affect E. coli in a number of conditions tested (Brokx, S.J. and Weiner, J.H. personal communication). Very differently, changes in the expression patterns of human genes containing the ACRATA domain could be related with apoptosis or cancer [1–14]. The well known functional flexibility of oxidoreductases is exemplified in bacterial proteins such as cupredoxins and cytochromes, normally involved in electron transfer during respiration but that can enter in eukaryotic cells to induce apoptosis or inhibition of cell growth , or in the Nox family, with functions as different as host defence in phagocytes or extracellular matrix modification [11, 12].
We have described for the first time a 6TM domain present in three protein families (STEAP, YedZ and Nox): the ACRATA domain. The common functions of the proteins of those families suggest that this domain is involved in electron transfer, mediated by its heme binding capability. We hypothesize that STEAP, STAMP1, TSAP6, and TIARP have this function, and that they form part of electron transfer systems involved in cellular regulation, apoptosis, and cancer. Additional experimental approaches using different members of the ACRATA domain-containing families are required to confirm these hypotheses.
six-transmembrane epithelial antigen of the prostate
six transmembrane protein of prostate 1
tumor suppressor activated pathway-6
Tumor necrosis factor-alpha-induced adipose-related protein
Translationally controlled tumor protein
reactive oxygen species
flavin adenine dinucleotide
Hidden Markov Models
We are grateful to S.J. Brokx and J.H. Weiner (University of Alberta, Edmonton, Canada) for useful comments on early versions of the manuscript. Thanks also to G.R. Reina and J. Menéndez (Hospital Puerta de Hierro, Madrid, Spain) for their encouragement and moral support.
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