DISCUSSION The Pogona vitticeps selenoproteome, cysteine homologues and associated machinery were predicted by comparing it with the Anolis carolinensis and the human genome. The methionine problem First of all, we chose the Anolis carolinensis genome from SelenoDB 2.0 because it was phylogenetically close to Pogona vitticeps. However, we observed that most of the proteins annotated in the data base did not start with a methionine and, what is more, some proteins were very poorly annotated. Therefore, we decided to compare the Pogona vitticeps with the human genome from the SelenoDB 1.0 as it is better annotated. If the results obtained from the comparison between Pogona vitticeps and Homo sapiens were similar, the Anolis carolinensis annotation would be chosen as the reference genome for the phylogenetically reason. Once we started the analysis, we noticed that a big number of alignments did not predict a methionine at the beginning of the Pogona vitticeps protein. In order to solve this problem we took into account the results obtained from the Seblastian, Genewise prediction and we also searched for an ATG codon in the file resulted from the fastasubseq analysis. However, in the case of the cysteine homologues and the machinery proteins, we could only use the results from the Genewise and the ATG codon search. In some cases, no methionine was found and we could not conclude that this protein was present in the Pogona vitticeps genome. After all the analysis, we observed that most of the proteins found in the Pogona vitticeps genome did not start with a methionine and therefore we are not sure if these proteins could be functional even if high homology was observed. In the cases where we found high homology, selenocysteine and a SECIS element but no methionine, we think that this could be due to poor annotation of the Pogona vitticeps genome. An article from Touriol C, et al (2003)[14], mentions that a small number of mammals mRNAs initiate translation from a non-AUG codon (Leu or Val codon in most of cases). The use of a non-AUG codon depends on the sequence context and the secondary structure of the next codon. Therefore, some of the proteins that did not start with a methionine could have this alternative translation or maybe they would not be correctly translated and therefore they would not be functional proteins. We have found that no methionine was present in some selenoproteins that have been described as ancestral selenoproteins [6]. In this case, although no methionine was found and we classified them as “grey” in the previous results section, we have considered that these selenoproteins are more likely to be present in the Pogona vitticeps genome because they are ancestral and no loss or conversion has been previously described. These proteins are: GPX1, TR1, TR3, SelM, SelN, MSRB1 and SelW2 . In the following section we present the selenoproteins, cysteine homologues of selenoproteins and machinery involved in the biosynthesis of selenoproteins that we found. They will be distinguished using the following criteria: - Green → Selenoproteins - Blue → Cysteine homologues of selenoproteins - Red → Machinery for the biosynthesis of selenoproteins Here you can access each protein directly: GPx1 GPx2 GPx3 GPx4 GPx5 GPx6 GPx7 GPx8 DI1 DI2 DI3 TR1 TR2 TR3 SecS Sel15 SelH SelI SelK SelM SelN SelO SelP SelR1 SelR2 SelR3 SelS SelT SelU1 SelU2 SelU3 SelV SelW1 SelW2 MsrA MSRB1 MSRB2 MSRB3 SBP2 Secp43 PSTK SEPHS1 SPS2 eEFsec GPx Glutathione peroxidases are the largest selenoprotein family in vertebrates. In mammals we find 8 GPx homologues, from which five (GPx1-4 and GPx6) contain a Sec residue in their active site and the other three (GPx5, GPx7 and GPx8) the active site Sec is replaced by Cys. GPxs are involved in hydrogen peroxidase (H2O2) signaling, detoxification of hydroperoxides and maintaining cellular redox homeostasis. We wanted to find if these selenoproteins also exist in Pogona vitticeps [6][13]. - GPx1 GPx1 is a cytosolic enzyme responsible for catalyzing glutathione GSH-dependent reduction of H2O2 to water, that is why many physiological roles of H2O2 such as cell proliferation, apoptosis and stress response are modulated by this enzyme. GPx1 is an ubiquitous enzyme expressed in all cell types, especially liver and kidney. It is also the first mammalian protein whose gene was found to contain Sec-encoding UGA [13]. Both lizard and human GPx1 proteins predict an homologous protein in the same genomic region of Pogona vitticeps. Even though none of the predictions aligns with methionine, the human protein shows a better annotation as it starts with a methionine. The selenocysteine present in the human query also aligns with a selenocysteine in the Pogona vitticeps genome. We also could not find a ATG upstream the protein in the file from the fastasubseq. Seblastian predicted a selenoprotein that corresponds with the one obtained from Exonerate. Moreover, a SECIS can be found in the 3’UTR.       We conclude that this protein may exist in the Pogona vitticeps genome in scaffold CEMB01545308 between positions 4557477-4560307 in the direct strand. As mentioned at the beginning of the discussion, it is suggested that GPX1 is one of the ancestral vertebrate selenoproteins and therefore we think that this protein is more likely to be present in the Pogona vitticeps genome. - GPx2 It is mainly found in the epithelium of the gastrointestinal tract and has specificity for hydrogen peroxide and other hydroperoxides. It has been shown to play a role in the development of cancer, but it is still unknown if it protects against cancer or promotes it [13]. Both lizard and human GPx2 proteins predict an homologous protein in the same genomic region of Pogona vitticeps. Both predictions start with methionine and therefore we chose the lizard protein as a reference because it is phylogenetically closer compared with the human. The selenocysteine present in the lizard query also aligns with a selenocysteine in the Pogona vitticeps genome. Seblastian predicted a selenoprotein that corresponds with the one obtained from Exonerate. Moreover, a SECIS can be found in the 3’UTR.       We conclude that this protein is in the Pogona vitticeps genome in scaffold CEMB01012720.1 between positions 1089793-1093950 in the direct strand. - GPx3 GPx3 is secreted primarily from kidney and is the major GPx form in plasma. It protects cells against oxidative stress and downexpression of this gene has been related to various cancers [13]. Both lizard and human GPx3 proteins predict an homologous protein in the same genomic region of Pogona vitticeps. However, the human protein shows a better alignment as the predicted protein starts with a methionine, which aligns with the pogona genome. The selenocysteine present in the human query also aligns with a selenocysteine in Pogona vitticeps genome. Seblastian predicted a selenoprotein that corresponds with the one obtained from Exonerate. Moreover, a SECIS can be found in the 3’UTR.      We conclude that this protein is in the Pogona vitticeps genome in scaffold CEMB01545298.1 between positions 148504 - 230484 in the direct strand. - GPx4 It is expressed in a wide range of cell types and tissues. GPx4 inhibits lipid peroxidation and also prevents oxidative stress-induced apoptosis [13]. Both lizard and human GPx4 proteins predict an homologous protein in the same genomic region of Pogona vitticeps. Even though none of the predictions align with methionine, the human protein shows a better annotation as the human selenoprotein begins with a methionine. The selenocysteine present in the human query also aligns with a selenocysteine in Pogona vitticeps genome. We examined the results obtained with the Genewise and no methionine alignment was present. However, Seblastian predicted a selenoprotein that corresponds with the one obtained from Exonerate and in this case it begins with a methionine. Moreover, a SECIS can be found in the 3’UTR.      We conclude that this protein exists in the Pogona vitticeps genome in scaffold CEMB01020348.1 between positions 449454 - 458335 in the reverse strand. - GPx5 This protein is specifically expressed in the epididymis in the mammalian male reproductive tract, and is androgen-regulated. It has been proposed to play a role in protecting the membranes of spermatozoa from lipid peroxidation [13]. This protein is only annotated for the human genome. The predicted protein starts with methionine and aligns the cysteine. However, we find a X residue followed by a bad alignment so we think that this could correspond to a stop codon and, so that, this protein could be shorter in Pogona vitticeps.       We conclude that this protein exists in the Pogona vitticeps genome in scaffold CEMB01006746.1 between positions 201027 - 201269 in the reverse strand. - GPx6 It is found in olfactory epithelium and during embryonic development. The exact function of this gene is not known [13]. This protein is only annotated for the human genome. Seblastian predicted a selenoprotein that corresponds with the GPx3 and not the GPx6. Moreover, the scaffold for both proteins is the same.      We conclude that this protein is not present in the Pogona vitticeps genome because in the Exonerate prediction a X residue that could correspond to a stop codon is present before the selenocysteine and no methionine was present. - GPx7 and GPx8 GPx7 suppresses ROS and protexts against oxidative DNA damage and double-strand breaks.GPx8 is found in the cellular membrane [13]. GPx7 is only annotated for the human genome whereas GPx8 is annotated for both human and lizard. In this case we decided to take the human protein for further analysis as it is better annotated. For GPx7 we have predicted five exons, three of which match with the three exons predicted for GPx8 in the same scaffold. We think that these two proteins could be two forms of alternative splicing. Although none of the predictions start with methionine, the GPx7 start with a valine which can be a start codon [14].      We conclude that this protein may be in the Pogona vitticeps genome in scaffold CEMB01025174.1 between positions 1189829 - 1249111. However, the GPx8 prediction does not start with methionine so we can not conclude that this protein would be functional in Pogona vitticeps. DI1, DI2 and DI3 The iodothyronine deodinase family of selenoproteins are involved in regulation of thyroid hormone activity by reductive deodination, that is, they are involved in the activation or deactivation of thyroid hormones. In mammals, we find three proteins: DI1, DI2 and DI3. DI1 and DI3 are found on the plasma membrane, but DI2 is localized to the endoplasmatic reticulum. Moreover, DI1 and DI2 catalyze the deionidation of T4 to T3 (active), while DI3 converts T4 to rT3 (inactive). We wanted to know if these proteins were also found in Pogona vitticeps [13]. - DI1 Both lizard and human DI1 proteins predict an homologous protein in the same genomic region of Pogona vitticeps. Even though none of the predictions align with methionine, the human protein shows a better annotation as it begins with a methionine. The selenocysteine present in the human query also aligns with a selenocysteine in the Pogona vitticeps genome. However Seblastian predicted a selenoprotein that corresponds with the one obtained from Exonerate and it aligns a Methionine at the beginning. Moreover, a SECIS can be found in the 3’UTR.      We conclude that this protein is in the Pogona vitticeps genome in scaffold CEMB01015858.1 between positions 845129- 851790 in the direct strand. - DI2 Both lizard and human DI2 proteins predict an homologous protein in the same genomic region of Pogona vitticeps. Both predictions start with a methionine and therefore we chose the lizard protein as a reference because it is phylogenetically closer compared with the human. The selenocysteine present in the lizard query also aligns with a selenocysteine in Pogona vitticeps genome. However, Seblastian did not predict a selenoprotein but a SECIS can was found in the 3’UTR.      We conclude that this protein is in the Pogona vitticeps genome in scaffold CEMB01024191.1 between positions 69236 - 86155 in the direct strand. - DI3 Only human DI3 protein predicts an homologous protein in Pogona vitticeps. The human DI3 selenoprotein was chosen because the DI3 annotation was not found in the lizard genome from SelenoDB 2.0. Even the prediction does not start with a Methionine. The selenocysteine present in the human query also aligns with a selenocysteine in the Pogona vitticeps genome. In this case we examined the prediction obtained from Genewise and it was very similar to the Exonerate prediction. No methionine alignment was found at the beginning of the protein. Moreover we could not find an ATG codon in the file resulted from fastasubseq. Seblastian predicted a selenoprotein that corresponds with the one obtained from Exonerate. Moreover, a SECIS can be found in the 3’UTR. As the predicted protein starts with a Leucine, it is possible that the translation could start with this amino acid [14].      We conclude that this protein is in the Pogona vitticeps genome in scaffold CEMB01002674.1 between positions 1655498 - 1656337 in the reverse strand. TR1, TR2 and TR3 They are part of the pyridine nucleotide-disulfide oxidoreductase family, which participates in many aspects of cell function based on protection against oxidant injury, cell growth and transformation, and the recycling of ascorbate from its oxidative form. Apart from reducing Trx, it also reduces other substrates. We find TR1, TR2 and TR3, located in cytosol or nucleus, mitochondria and testis, respectively. TR1 and TR2 are important during development [37]. - TR1 The Exonerate prediction does not start with methionine but we found high homology and the selenocysteines aligned. Seblastian could predict a selenoprotein where the selenocysteine aligns. Moreover, a SECIS could be found in the 3’UTR.      We conclude that this protein may exist in the Pogona vitticeps genome in scaffold CEMB01031819.1 between positions 1005603 - 1033533 in the direct strand. As mentioned at the beginning of the discussion, it is suggested that TR1 is one of the ancestral vertebrate selenoproteins and therefore we think that this protein is more likely to be present in the Pogona vitticeps genome. - TR2 and TR3 We decided to take the human proteins for further analysis as they were better annotated. For TR3 the Exonerate predicted sixteen exons, twelve of which match with the twelve exons predicted for TR2 in the same scaffold. The Seblastian predicted the same selenoprotein for both and we could find a SECIS in the 3’UTR.      We think that these two proteins could be two forms of alternative splicing. None of the predictions start with methionine so we can not conclude that these proteins would be functional in Pogona vitticeps. However, As mentioned at the beginning of the discussion, it is suggested that TR3 is one of the ancestral vertebrate selenoproteins and therefore we think that this protein is more likely to be present in the Pogona vitticeps genome. SecS Sec synthase (SecS) uses monoselenophosphate to convert Ser-tRNASec into Sec-tRNASec to allow Sec to be incorporated into proteins [7]. Both lizard and human SecS proteins predict an homologous protein in the same genomic region of Pogona vitticeps. As the prediction against the Lizard starts with methionine, the Lizard protein was chosen as a reference. The cysteine residues are also aligned.      We conclude that this protein is in the Pogona vitticeps genome in scaffold CEMB01019488.1 between positions 3072240-3123698 in the direct strand. Sel15 Little is known about Sel15, but it is believed to play a role in protection against prostate cancer, as it is located on a chromosome often affected by cancer and selenium supplementation appears to decrease incidence of this cancer [15]. Both lizard and human Sel15 proteins predict an homologous protein in the same genomic region of Pogona vitticeps. Even though none of the predictions align with methionine, the human protein shows a better annotation because it begins with a methionine. However, a gap of amino acids was found at the beginning of the Pogona vitticeps selenoprotein. The selenocysteine present in the human query also aligns with a selenocysteine in Pogona vitticeps genome. However Seblastian predicted a selenoprotein that started with a methionine. Moreover, a SECIS can be found in the 3’UTR.      We conclude that this protein is in the Pogona vitticeps genome in scaffold CEMB01029493.1 between positions 105905 - 155503 in the reverse strand. SelH SelH may be related in redox-related processes [16]. Both lizard and human SelH proteins predict an homologous protein in the same genomic region of Pogona vitticeps. We chose the human protein because it shows a better annotation because it begins with a methionine. The selenocysteine present in the human query also aligns with a selenocysteine in Pogona vitticeps genome. However Seblastian did not predict a selenoprotein. Moreover, any SECIS can not be found in the 3’UTR.      We cannot conclude that this protein is in the Pogona vitticeps genome because the results from the Exonerate do not match the results obtained from the Seblastian and SECIS prediction. A SECIS element is necessary in order to have a selenoprotein. So, we think it is more likely that the SelH is not present in the Pogona vitticeps genome. This result was surprising to us because SelH it has been suggested to be one of the ancestral vertebrate selenoproteins. No loss or conversion to cysteine has been described and therefore we hypothesise that, the result from the SECIS and Seblastian prediction could be due a to a bad annotation in the P. vitticeps genome or a bad prediction by Seblastian and it is more likely that SelH is present in the Pogona vitticeps genome. SelI SelI helps to form and maintain vesicular membranes by catalyzing phosphatidylethanolamine biosynthesis from CDP-ethanolamine [17]. Both lizard and human SelI proteins predict an homologous protein in the same genomic region of Pogona vitticeps. None of the predictions with Exonerate start with a methionine and none of them align with a selenocysteine. However, the Genewise against the human protein starts with a methionine. Seblastian could predict a selenoprotein where the selenocysteine aligns. Moreover, a SECIS could be found in the 3’UTR.      We conclude that this protein is in the Pogona vitticeps genome in scaffold CMB01020366.1 between positions 114743 - 208479 in the reverse strand and that the issue with the methionine could be due to a bad annotation. SelK SelK is important in immune cells, where it is required for Ca2+ flux, T-cell proliferation and neutrophil migration. It also protects cells from ER stress-induced apoptosis, among other functions [18]. Both lizard and human SelK proteins predict an homologous protein in the same genomic region of Pogona vitticeps. Both predictions start with methionine and therefore we chose the lizard protein as a reference because it is phylogenetically closer compared with the human. The selenocysteine present in the lizard query also aligns with a selenocysteine in Pogona vitticeps genome. we also examined the prediction obtained with the Genewise and it is very similar to the prediction from Exonerate. Seblastian predicted a selenoprotein that corresponds with the one obtained from Exonerate. Moreover, a SECIS can be found in the 3’UTR.      We conclude that this protein is in the Pogona vitticeps genome in scaffold CEMB01006654.1 between positions 1858186 - 1863288 in the reverse strand. SelM Function still not clear, it may function as a thiol-disulfide oxidoreductase that participates in disulfide bond formation [19]. Both lizard and human SelM proteins predict an homologous protein in the same genomic region of Pogona vitticeps. Even though none of the predictions align with methionine, the human protein shows a better annotation because the human selenoprotein begins with a methionine. The selenocysteine present in the human query also aligns with a selenocysteine in Pogona vitticeps genome. However, a gap of amino acids was found at the beginning of the Pogona vitticeps selenoprotein. Moreover we could not find an ATG codon in the fastasubseq file. Seblastian predicted a selenoprotein that corresponds with the one obtained from Exonerate but it did not start with a methionine. Moreover, a SECIS was found in the 3’UTR.      We conclude that this protein may exist in the Pogona vitticeps genome in scaffold CEMB01008420.1 between positions 924112 - 162538 in the positive strand. As mentioned at the beginning of the discussion, it is suggested that SelM is one of the ancestral vertebrate selenoproteins and therefore we think that this protein is more likely to be present in the Pogona vitticeps genome. SelN SelN protects the cell against oxidative stress and participates in the regulation of redox-related calcium homeostasis through ryanodine receptor. It is essential for muscle regeneration and satellite cell maintenance in skeletal muscle [20]. Both lizard and human SelN proteins predict an homologous protein in the same genomic region of Pogona vitticeps. In this case, the human protein has two selenocysteines whereas the lizard protein has only the second one. None of the predictions start with a methionine and no methionine could be found in the fastasubseq file. The predicted gene only aligns with the second selenocysteine and this alignment shows more homology with the lizard protein. We think that the first selenocysteine has been lost along evolution. Seblastian could not predict a selenoprotein. However, a SECIS could be found in the 3’UTR.      We can conclude that this protein may exist in the Pogona vitticeps genome in scaffold CEMB01037816.1 between positions 1211310-1227196 in the direct strand. However,as mentioned at the beginning of the discussion, it is suggested that SelN is one of the ancestral vertebrate selenoproteins and therefore we think that this protein is more likely to be present in the Pogona vitticeps genome. SelO SelO function is still not well known, but it may be a redox-active mitochondrial selenoprotein which interacts with a redox target protein [21]. Both lizard and human SelO proteins predict an homologous protein in the same genomic region of Pogona vitticeps. Even though none of the predictions start with methionine, for the human protein we have found an ATG codon upstream the predicted gene in the file obtained from fastasubseq. Moreover, the selenocysteine aligns with a selenocysteine in Pogona vitticeps genome whereas in the lizard this alignment is not found. Seblastian predicted a selenoprotein that corresponds with the one obtained from Exonerate. Moreover, a SECIS can be found in the 3’UTR.      We conclude that this protein is found in the Pogona vitticeps genome in scaffold CEMB01025919.1 between positions 171404-282103 in the direct strand and that the issue with the methionine could be due to a bad annotation. SelP SelP function is still not well known, but it is thought that this protein could be involved in supplying selenium to tissues such as brain and testis, as well as creating some extracellular antioxidant defense [22]. Both lizard and human SelP proteins predict an homologous protein in the same genomic region of Pogona vitticeps. However, the human protein shows a better alignment as the predicted gene starts with a methionine. Seblastian predicted a selenoprotein that also starts with methionine and that has the selenocysteines aligned. Moreover, a SECIS can be found in the 3’UTR.      We conclude that this protein is in the Pogona vitticeps genome in scaffold CEMB01031586.1 between positions 3789864 - 3884508 in the direct strand. SelR1, SelR2 and SelR3 This protein is a methionine sulfoxide reductase that works together with MsrA to reduce methionine sulfoxide and regulate biological processes and cope with oxidative stress. There are three of them: SelR1, SelR2 and SelR3[23]. After doing the analysis and searching in the bibliography [6], we have found that MSRB (extracted from SelenoDB 2.0) and SelR (extracted from SelenoDB 1.0) are the same proteins. We can observe that the scaffolds predicted are the same and in the case of MSRB1 and SelR1 the alignment is identical. - SelR1 Only human SelR1 protein predicts an homologous protein in Pogona vitticeps. The human SelR1 selenoprotein from SelenoDB 1.0 was chosen because the SelR1 annotation was not found in the lizard genome from SelenoDB 2.0. The selenocysteine present in the human query aligns with a selenocysteine in Pogona vitticeps genome. However, the Pogona vitticeps genome does not start with a Methionine. Moreover, we could not find a ATG codon at the beginning of the predicted protein in the file obtained from fastasubseq. Seblastian predicted a selenoprotein that corresponds with the one obtained from Exonerate and it did not start with a methionine. Moreover, a SECIS can be found in the 3’UTR.      We can conclude that this protein may exist in the Pogona vitticeps genome in scaffold CEMB01034240.1 between positions 245530- 325002 in the direct strand. - SelR2 Only human SelR2 protein predicts an homologous protein in Pogona vitticeps. The human SelR1 selenoprotein from SelenoDB 1.0 was chosen because the SelR2 annotation was not found in the lizard genome from SelenoDB 2.0. The algiment obtained showed a gap of amino acids at the beginning and at the end of the Pogona vitticeps protein. The cysteine present in the human query aligns with a cysteine in the P. vitticeps genome. However, before this alignment, some X residues could be found. In this case, we examined the results obtained from the Genewise prediction and they were similar to the Exonerate results: big gaps of amino acids were found, and a X residue appeared before the cysteines alignment.      Taking into account that the alignment with the human query has two big gaps of amino acids in the two predictions and some X residues (that could be stop codons)are found before the cysteine alignment, we conclude that this protein can not be found in the Pogona vitticeps genome. - SelR3 Only human SelR3 protein predicts an homologous protein in Pogona vitticeps. The human SelR3 protein from SelenoDB 1.0was chosen because the SelR3 annotation was not found in the lizard genome from SelenoDB 2.0. The cysteine present in the human query aligns with a cysteine in Pogona vitticeps genome. However, the methionine did not align at the beginning. We examined the results obtained with the Genewise prediction and they were very similar to the Exonerate results. They also predicted the cysteine alignment and did not align the methionine. Moreover we could not find an ATG codon at the beginning of the predicted protein in the file obtained from fastasubseq.      We can not conclude that this protein selenocysteine would be functional in the Pogona vitticeps genome in scaffold CEMB01015116.1 between positions 223908 - 2293719 in the reverse strand. SelS SelS is involved in the degradation process of misfolded endoplasmatic reticulum (ER) luminal proteins. It transfers the misfolded proteins from the ER to cytosol, where they are destroyed [24]. Both lizard and human SelS proteins predict an homologous protein in the same genomic region of Pogona vitticeps. Even though none of the predictions start with methionine, the human protein shows a better alignment as the selenocysteine aligns with a selenocysteine in Pogona vitticeps genome whereas in the lizard this alignment is not found. Seblastian could not predict a selenoprotein. However, a SECIS could be found in the 3’UTR. As the predicted proteins starts with Leucine, thi could be a start codon for the predicted protein [14].      We conclude that this protein may be found in the Pogona vitticeps genome in scaffold CMB01545307.1 between positions 2383099-2451895 in the direct strand. SelT SelT has thioredoxin reductase-like oxidoreductase activity. It protects dopaminergic neurons against oxidative stress and cell death by taking part in processes of calcium mobilization and neuroendocrine secretion, among other functions [25]. Both lizard and human SelT proteins predict an homologous protein in the same genomic region of Pogona vitticeps. We chose the human selenoprotein because it shows a better annotation as the human selenoprotein starts with a methionine and it aligns with a methionine in the Pogona vitticeps genome. The selenocysteine present in the human query also aligns with a selenocysteine in the Pogona vitticeps genome. Seblastian predicted a selenoprotein that corresponds with the one obtained from Exonerate. Moreover, a SECIS can be found in the 3’UTR.      We conclude that this protein is in the Pogona vitticeps genome in scaffold CEMB01025449.1 between positions 2390177-2399217 in the direct strand. SelU1, SelU2 and SelU3 It catalyzes the transfer of selenium from selenophosphate for conversion conversion of 2-thiouridine to 2-selenouridine at the wobble position in tRNA. There are three different types of this protein: SelU1, SelU2 and SelU3 [26]. - SelU1 Both lizard and human SelU1 proteins predict an homologous protein in the same genomic region of Pogona vitticeps. Both queries were analyzed because we observed that the human genome has a cysteine annotated in its genome whereas a selenocysteine is found in the same position in the Anolis carolinensis genome. The alignment between the Pogona vitticeps and Anolis carolinensis genome showed an alignment between the two selenocysteines whereas the analysis with the human genome reported an alignment between a cysteine and a selenocysteine in the same position. Even though the annotation from the Anolis carolinensis did not start with methionine, we chose the lizard annotation because is phylogenetically closer than the human annotation. Seblastian predicted a selenoprotein that corresponds with the one obtained from Exonerate and it begins with a methionine. Moreover, a SECIS can be found in the 3’UTR.      We conclude that a selenoprotein can be found in the Pogona vitticeps genome in scaffold CEMB01004403.1 between positions 1495870 - 1503336 in the positive strand. As said before, we observed that a cysteine is present in the Human genome. We think that the Pogona vitticeps and the Anolis carolinensis have maintained a selenocysteine in this position whereas the human is a cysteine homolog. - SelU2 Both lizard and human SelU2 proteins predict an homologous protein in the same genomic region of Pogona vitticeps. Even though none of the predictions aligns with a methionine, the human protein shows a better annotation because it starts with a methionine. The cysteine present in the human query aligns with a cysteine in Pogona vitticeps genome. However a big gap of amino acids was observed in the first region of the Pogona vitticeps protein. Moreover, we could not find an ATG codon upstream in the file obtained from the fasta subseq.      We can not conclude that this protein would be functional in the Pogona vitticeps genome in scaffold CEMB01001592.1 between positions 719555 - 760989 in the direct strand. - SelU3 Both lizard and human SelU3 proteins predict an homologous protein in the same genomic region of Pogona vitticeps. Even though none of the predictions align with methionine, the human protein shows a better annotation because it starts with a methionine. However, we found an ATG codon in the file obtained from fastasubseq just two amino acids upstream the predicted protein. The cysteine present in the human query aligns with a cysteine in Pogona vitticeps genome.      We conclude that this protein is in the Pogona vitticeps genome in scaffold CEMB01011587.1 between positions 566369 - 574331 in the direct strand. SelV SelV may be involved in redox-related processes, but its function is still not well known [39]. It is the least conserved mammalian selenoprotein that likely arose from a duplication of SelW in the placental stem.[6] No hit was found for SelV using human reference annotation.      Therefore, we concluded that SelV was not present in the Pogona vitticeps genome. This matches the information from Mariotti M (2012) [6] where they mention that SelV is one of the less conserved proteins. SelW1 and SelW2 SelW is a glutathione (GSH)-dependent antioxidant involved in redox processes. It is highly expressed in skeletal muscle, heart and brain, where it places a role as a muscle growth and differentiation factor and neurons protector from oxidative stress during neuronal development. We find two types: SelW1 and SelW2 [27]. - SelW1 Both lizard and human SelW1 proteins predict an homologous protein in the same genomic region of Pogona vitticeps. The human protein was selected because it shows a better annotation as it starts with a methionine that aligns with a methionine in the Pogona vitticeps protein. The selenocysteine present in the human query aligns with a selenocysteine in Pogona vitticeps genome. We also examined the results of the Genewise prediction and they were similar to the Exonerate ones. Seblastian did not predict a selenoprotein but a SECIS was found in the 3’UTR.      We conclude that this protein is in the Pogona vitticeps genome in scaffold CEMB01027437.1 between positions 18032 - 23866 in the reverse strand. - SelW2 Only human SelW2 protein predicts an homologous protein in Pogona vitticeps. The human SelW2 selenoprotein was chosen because the SelW2 annotation was not found in the lizard genome from SelenoDB 2.0. The cysteine present in the human query also aligns with a cysteine in Pogona vitticeps genome, although we found a gap of amino acids at the beginning of the Pogona vitticeps sequence so the methionine did not align and we could not find an ATG codon in the file obtained from fastasubseq.      We cannot conclude that this protein would be functional in the Pogona vitticeps genome in scaffold CEMB01003800.1 between positions 1676926- 1754339 in the reverse strand. As mentioned at the beginning of the discussion, it is suggested that SelW2 is one of the ancestral vertebrate selenoproteins and therefore we think that this protein is more likely to be present in the Pogona vitticeps genome. MsrA If a protein has been inactivated by oxidation, this protein acts a a repair enzyme and catalyzes the reversible oxidation-reduction of methionine sulfoxide in proteins to methionine [28]. Both lizard and human MsrA proteins predict an homologous protein in the same genomic region of Pogona vitticeps. Even though none of the predictions aligns with methionine, the human protein shows a better annotation because it starts with a methionine. A big gap of amino acids was observed in the first region of the Pogona vitticeps protein. However an alignment of two methionines could be found after the amino acids gap. Therefore we consider that this alignment of methionines could be the start of the protein in Pogona vitticeps. Moreover, the cysteine present in the human query also aligns with a cysteine in Pogona vitticeps genome.      Although a big gap of amino acids in the first region was found, we think that this protein is present in the Pogona vitticeps genome in scaffold CEMB01004415.1 between positions 1787-119750 in the direct strand. MSRB MSRB is a family of proteins which reduce methionine (R)-sulfoxide back to methionine. We distinguish three types: - MSRB1: It helps to regulate actin assembly and thereby promoting filament repolymerization in cells such as macrophages [29]. - MSRB2: Upon oxidative stress, may play a role in the preservation of mitochondrial integrity by decreasing the intracellular reactive oxygen species [30]. - MSRB3: essential for hearing [31]. After doing the analysis and searching in the bibliography [6], we have found that MSRB (extracted from SelenoDB 2.0) and SelR (extracted from SelenoDB 1.0) are the same proteins. We can observe that the scaffolds predicted are the same and in the case of MSRB1 and SelR1 the alignment is identical. - MSRB1 Both lizard and human MSRB1 proteins predict an homologous protein in the same genomic region of Pogona vitticeps. Even though none of the predictions align with a methionine, the human protein shows a better annotation as the human selenoprotein as it starts with a methionine. In this case the human reference was obtained from SelenoDB 2.0database. The selenocysteine present in the human query also aligns with a selenocysteine in Pogona vitticeps genome. Seblastian predicted a selenoprotein that corresponds with the one obtained from Exonerate which also did not start with a Methionine. A SECIS can be found in the 3’UTR.      We conclude that this protein may exist in the Pogona vitticeps genome in scaffold CEMB01034240.1 between positions 245530 - 325002 in the direct strand and that the issue with the methionine could be due to a bad annotation. As mentioned at the beginning of the discussion, it is suggested that MSRB1 is one of the ancestral vertebrate selenoproteins and therefore we think that this protein is more likely to be present in the Pogona vitticeps genome. - MSRB2 Both lizard and human MSRB2 proteins predict an homologous protein in the same genomic region of Pogona vitticeps. Even though none of the proteins aligns with methionine, the human protein shows a better annotation as the human selenoprotein annotation starts with a methionine. In this case the human sequence was obtained from SelenoDB 2.0 data base. We do not know if the cysteine present in the human query aligns with a cysteine in Pogona vitticeps genome because many cysteine residues are found in the human sequence and the reference database does not provide to us which cysteine residue is the one that has changed. We also observed that the alignment between the query and the Pogona vitticeps was not very good and we could find some amino acids gaps. The Genewise prediction was similar to the Exonerate one and it showed a poor alignment, also with no methionines aligned.      We think that the MSRB2 protein is not present in the Pogona vitticeps genome because we can not confirm if there is an alignment between the cysteine residues and also big gaps of amino acids are found in the alignment. - MSRB3 Both lizard and human MSRB3 proteins predict an homologous protein in the same genomic region of Pogona vitticeps. Both predictions start with methionine and therefore we chose the lizard protein as a reference because it is phylogenetically closer compared with the human. The cysteine present in the lizard query also aligns with a cysteine in the Pogona vitticeps genome and a methionine alignment was found. The Genewise prediction was very similar to the Exonerate prediction and it confirmed our results.      We conclude that this protein is in the Pogona vitticeps genome in scaffold CEMB01015116.1 between positions 2239081 - 2312173 in the reverse strand. Secp43 Secp43 is a tRNA selenocysteine associated protein, so it takes part in the biosynthesis of selenoproteins [32]. Both lizard and human Secp43 proteins predict an homologous protein in the same genomic region of Pogona vitticeps. The human protein was chosen because it shows a better annotation as it starts with a methionine aligned with the Pogona vitticeps genome. In this case, the human sequence was obtained from SelenoDB 2.0 data base.      We can conclude that the Secp43 protein is present in the Pogona vitticeps genome in scaffold CEMB01011943.1 between positions 1979887 - 1991795 in the positive strand. PSTK This protein phosphorylates seryl-tRNA(Sec) to O-phosphoseryl-tRNA(Sec), an activated intermediate for selenocysteine biosynthesis [34], so it is part of the machinery for selenoproteins biosynthesis. Both lizard and human PSTK proteins predict an homologous protein in the same genomic region of Pogona vitticeps. Even though none of the predictions aligns with methionine, the human protein shows a better annotation as it starts with a methionine. However, we found an ATG codon in the file obtained from the fastasubseq. In this case the human sequence was obtained from SelenoDB 2.0 data base.      We can conclude that the PSTK protein is present in the Pogona vitticeps genome in scaffold CEMB01002406.1 between positions 243615 - 348310 in the positive strand. SEPHS1 and SPS2 Selenophosphate synthetase (SPS) has two paralogues, SPS1 and SPS2. SPS1 has been shown to regulate redox homeostasis and control cell growth, while SPS2 catalyses the synthesis of selenophosphate from selenide and ATP [34] [35]. SEPHS1 and the SPS1 human protein from the SelenoDB 1.0 are homologues [35]. Both T-coffees from Exonerate and Genewise show a high homology and the sequence predicted starts with a methionine.      We conclude that this protein exists in the Pogona vitticeps genome in scaffold CEMB01026985.1 between positions 17484 - 66734 in the reverse strand. SPS2 was only annotated in the SelenoDB 1.0 for the human genome, too. We have found that it is an homologue of the SEPHS2 protein annotated for both human and lizard in the SelenoDB 2.0[35]. We decided to took the SPS2 human protein for further alignment as it is better annotated. The predicted gene does not start with methionine. In addition, the selenocysteine is not aligned. Seblastian could not predict a selenoprotein and no SECIS was found in the 3’UTR.      For all these reasons we conclude that this protein does not exist in the Pogona vitticeps genome. This result was surprising to us because SPS2 it has been suggested to be one of the ancestral vertebrate selenoprotein. No loss or conversion to cysteine has been described before and therefore we hypothesise that, the results could be due a to a bad annotation in the P. vitticeps genome. eEFsec eEFsec is a translation factor necessary for the incorporation of selenocysteine into proteins. [36] Both lizard and human eEFsec proteins predict an homologous protein in the same genomic region of Pogona vitticeps. We decided to took the human protein for further analysis as it is better annotated. The prediction obtained does not start with methionine and we did not find an ATG codon in the file resulted from fastasubseq.      Even though the results were not clear, high homology was present in the alignment and moreover, eEFsec is an essential machinery protein so we think that it may exist in the Pogona vitticeps genome in scaffold CEMB01008168.1 between positions 332147 - 553028 in the direct strand.