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Pseudogenes are defined as "non-functional" copies of corresponding parent genes. The cognition of pseudogenes continues to be refreshed through accumulating and updating research findings. Previous studies have predominantly focused on mammals, but pseudogenes have received relatively less attention in the field of microbiology. Given the increasing recognition on the importance of pseudogenes, in this review, we focus on several aspects of microorganism pseudogenes, including their classification and characteristics, their generation and fate, their identification, their abundance and distribution, their impact on virulence, their ability to recombine with functional genes, the extent to which some pseudogenes are transcribed and translated, and the relationship between pseudogenes and viruses. By summarizing and organizing the latest research progress, this review will provide a comprehensive perspective and improved understanding on pseudogenes in microorganisms. Key points: • Concept, classification and characteristics, identification and databases, content, and distribution of microbial pseudogenes are presented. • How pseudogenization contribute to pathogen virulence is highlighted. • Pseudogenes with potential functions in microorganisms are discussed. [ABSTRACT FROM AUTHOR]

Objective: Although radiation workers are exposed to much lower doses of neutron-γ rays than those suffered in nuclear explosions and accidents, it does not mean that their health is not affected by radiation. Lower doses of radiation do not always cause morphological aberrations in chromosomes, so more sophisticated tests must be sought to specific alterations in the exposed cells. Our goal was to characterize the specific gene expression in lymphocytes from logging workers who were continuously exposed to low doses of neutron-γ radiation. We hypothesized that the combination of cell type-specific transcriptomes and open chromatin profiles would identify lymphocyte-specific gene alterations induced by long-term radiation with low-dose neutron^-rays and discover new regulatory pathways and transcriptional regulatory elements. Methods: Lymphocytes were extracted from workers who have been occupationally exposed to neutron-γ and workers unexposed to radiation in the same company. mRNA-seq and ATAC-seq (Assay for Transposase-Accessible Chromatin with high-throughput sequencing) were performed, followed integrative analysis to identify specific gene regulatory regions induced by neutron-γ radiation. A qPCR assay was then performed to verify the downregulation of RNA coding for ribosomal proteins and flow cytometry was used to detect ribosomal protein expression and cell cycle alterations. Results: We identified transcripts that were specifically induced by neutron-γ radiation and discovered differential open chromatin regions that correlated with these gene activation patterns. Notably, we observed a downward trend in the expression of both differentially expressed genes and open chromatin peaks. Our most significant finding was that the differential peak upregulated in ATAC-seq, while the differential gene was downregulated in the ribosome pathway. We confirmed that neutron-γ radiation leads to transcriptional inhibition by analyzing the most enriched promoters, examining RPS18 and RPS27A expression by qPCR, and analyzing protein-protein interactions of the differential genes. Ribosomal protein expression and cell cycle were also affected by neutron-γ as detected by flow cytometry. Conclusion: We have comprehensively analyzed the genetic landscape of human lymphocytes based on chromatin accessibility and transcript levels, enabling the identification of novel neutron-γ induced signature genes not previously known. By comparing fine-mapping of open chromatin and RNA reads, we have determined that neutron-γ specifically leads to downregulation of genes in the ribosome pathway, with pseudogenes potentially playing a crucial role. [ABSTRACT FROM AUTHOR]

In the current study, we sequenced the complete plastome of Monstera deliciosa Liebm. (1849), an attractive foliage plant. The total length of the plastome of M. deliciosa is 163,499 bp and it consists of three distinct regions: a large single-copy (90,092 bp), a small single-copy (21,737 bp), and a pair of inverted repeats (IRs, 25,835 bp). The overall GC content is 36.2%, and the genome contains 110 functional genes, excluding pseudogenes. These functional genes encompass 77 protein-coding genes, 29 transfer RNA genes, and four ribosomal RNA genes. Notably, both the infA and rpl23 genes have been identified as pseudogenes. Phylogenetic analysis based on 14 representative plastomes from seven subfamilies indicates that Monsteroideae is monophyletic and sister to Pothoideae. Furthermore, M. deliciosa and M. adansonii were shown to share a recent common ancestor, the finding for which is supported by a strong bootstrap value. The sequenced plastome of M. deliciosa can serve as a valuable resource for establishing phylogenetic relationships and enhancing species identification within the genus Monstera. In addition, it can facilitate investigations into the genetic characteristics of this plant. [ABSTRACT FROM AUTHOR]

Background: Voltage-dependent anion selective channels (VDACs) are the most abundant mitochondrial outer membrane proteins, encoded in mammals by three genes, VDAC1, 2 and 3, mostly ubiquitously expressed. As 'mitochondrial gatekeepers', VDACs control organelle and cell metabolism and are involved in many diseases. Despite the presence of numerous VDAC pseudogenes in the human genome, their significance and possible role in VDAC protein expression has not yet been considered. Results: We investigated the relevance of processed pseudogenes of human VDAC genes, both in physiological and in pathological contexts. Using high-throughput tools and querying many genomic and transcriptomic databases, we show that some VDAC pseudogenes are transcribed in specific tissues and pathological contexts. The obtained experimental data confirm an association of the VDAC1P8 pseudogene with acute myeloid leukemia (AML). Conclusions: Our in-silico comparative analysis between the VDAC1 gene and its VDAC1P8 pseudogene, together with experimental data produced in AML cellular models, indicate a specific over-expression of the VDAC1P8 pseudogene in AML, correlated with a downregulation of the parental VDAC1 gene. [ABSTRACT FROM AUTHOR]

Most pseudogenes are generated when an RNA transcript is reverse-transcribed and integrated into the genome at a new location. Pseudogenes are often considered as an imperfect and silent copy of a functional gene because of the accumulation of numerous mutations in their sequence. Here we report the presence of Pfh8-ps, a Phf8 retrotransposed pseudogene in the mouse genome, which has no disruptions in its coding sequence. We show that this pseudogene is mainly transcribed in testis and can produce a PHF8-PS protein in vivo. As the PHF8-PS protein has a well-conserved JmjC domain, we characterized its enzymatic activity and show that PHF8-PS does not have the intrinsic capability to demethylate H3K9me2 in vitro compared to the parental PHF8 protein. Surprisingly, PHF8-PS does not localize in the nucleus like PHF8, but rather is mostly located at the cytoplasm. Finally, our proteomic analysis of PHF8-PS-associated proteins revealed that PHF8-PS interacts not only with mitochondrial proteins, but also with prefoldin subunits (PFDN proteins) that deliver unfolded proteins to the cytosolic chaperonin complex implicated in the folding of cytosolic proteins. Together, our findings highlighted PHF8-PS as a new pseudogene-derived protein with distinct molecular functions from PHF8. [ABSTRACT FROM AUTHOR]

The circadian clock allows the body to coordinate physiology with the light and dark cycles of the day. One of the critical components of the molecular clock machinery is the PER family of circadian clock genes, including PER1, PER2, and PER3. The PER3P1 (or PER4) is the only molecular clock-related pseudogene that has been derived from the PER3 gene. Pseudogenes have an essential role in the regulation of gene expression. In this study, we investigated the PER3P1 fluctuations and their effects on the PER3 expression level. We hypothesized that the PER3P1 pseudogene suppresses clock-related miRNAs and positively affects clock genes levels. The results showed that PER3P1 and its parent gene had a corresponding oscillatory expression pattern. The overexpression of PER3P1 increased the mRNA level of PER3. In contrast, the downregulation of PER3 decreased PER3P1 expression. Furthermore, PER3P1 modulates the expression level of other clock genes, including PER1, PER2, CRY1, and CRY2. Our results suggest that the PER3P1 pseudogene may positively affect the PER3 and other clock-related gene expressions. [ABSTRACT FROM AUTHOR]

Inherited copy number variations (CNVs) can provide valuable information for cancer susceptibility and prognosis. However, their association with oropharynx squamous cell carcinoma (OPSCC) is still poorly studied. Using microarrays analysis, we identified three inherited CNVs associated with OPSCC risk, of which one was validated in 152 OPSCC patients and 155 controls and related to pseudogene-microRNA-mRNA interaction. Individuals with three or more copies of ADAM3A and ADAM5 pseudogenes (8p11.22 chromosome region) were under 6.49-fold increased risk of OPSCC. ADAM5 shared a highly homologous sequence with the ADAM9 3′-UTR, predicted to be a binding site for miR-122b-5p. Individuals carrying more than three copies of ADAM3A and ADAM5 presented higher ADAM9 expression levels. Moreover, patients with total deletion or one copy of pseudogenes and with higher expression of miR-122b-5p presented worse prognoses. Our data suggest, for the first time, that ADAM3A and ADAM5 pseudogene-inherited CNV could modulate OPSCC occurrence and prognosis, possibly through the interaction of ADAM5 pseudogene transcript, miR-122b-5p, and ADAM9. [ABSTRACT FROM AUTHOR]

Prokaryotic genomes are usually densely packed with intact and functional genes. However, in certain contexts, such as after recent ecological shifts or extreme population bottlenecks, broken and nonfunctional gene fragments can quickly accumulate and form a substantial fraction of the genome. Identification of these broken genes, called pseudogenes, is a critical step for understanding the evolutionary forces acting upon, and the functional potential encoded within, prokaryotic genomes. Here, we present Pseudofinder, an open-source software dedicated to pseudogene identification and analysis in bacterial and archaeal genomes. We demonstrate that Pseudofinder's multi-pronged, reference-based approach can detect a wide variety of pseudogenes, including those that are highly degraded and typically missed by gene-calling pipelines, as well newly formed pseudogenes containing only one or a few inactivating mutations. Additionally, Pseudofinder can detect genes that lack inactivating substitutions but experiencing relaxed selection. Implementation of Pseudofinder in annotation pipelines will allow more precise estimations of the functional potential of sequenced microbes, while also generating new hypotheses related to the evolutionary dynamics of bacterial and archaeal genomes. [ABSTRACT FROM AUTHOR]

HMGN2 have functions in inflammatory response. However, the role of HMGN2 in severe acute pancreatitis (SAP) remains unclear. Here, our study was to discuss the role and regulatory mechanism ofHMGN2 in SAP. In this study, the SAP cell model of AR42J was used to study the function and mechanism of HMGN2 in SAP. The protein expression in cells and serums were examined by western blot and ELISA assay. qPCR was used to test the transcriptional RNA level. Cell viability were examined by MTT assay. Luciferase assay was used to evaluate the interaction between gene and gene. Our results showed that HMGN2 was significantly upregulated in SAP patients. The database predicted and luciferase assay data indicated the HMGN2 was directly binding with miR-590-3p. ELISA, MTT and western blot experiments showed that the HMGN2 were promoted the cell proliferation, reduced the inflammation, and repressed the cell autophagy. Mechanism studies showed that the pseudogene HMGN2P46 level was positively correlated with HMGN2 and upregulated HMGN2 expression by competing for miR-590-3p in SAP. Taken together, all over these results showed upregulation of HMGN2 alleviates SAP, this process was regulated by HMGN2P46 competitively binding with miR-590-3p, which may provide a new insight for the treatment and intervention in SAP. Pseudogene HMGN2P46 was a miRNA sponge to regulate HMGN2 level by competing for miR-590-3p to alleviate the process of SAP. It provided a novel strategy for the diagnosis and treatment of severe acute pancreatitis. [ABSTRACT FROM AUTHOR]

The outcomes of ovarian cancer are complicated and usually unfavorable due to their diagnoses at a late stage. Identifying the efficient prognostic biomarkers to improve the survival of ovarian cancer is urgently warranted. The survival-related pseudogenes retrieved from the Cancer Genome Atlas database were screened by univariate Cox regression analysis and further assessed by least absolute shrinkage and selection operator (LASSO) method. A risk score model based on the prognostic pseudogenes was also constructed. The pseudogene-mRNA regulatory networks were established using correlation analysis, and their potent roles in the ovarian cancer progression were uncovered by functional enrichment analysis. Lastly, ssGSEA and ESTIMATE algorithms was used to evaluate the levels of immune cell infiltrations in cancer tissues and explore their relationship with risk signature. A prediction model of 10-pseudogenes including RPL10P6, AC026688.1, FAR2P4, AL391840.2, AC068647.2, FAM35BP, GBP1P1, ARL4AP5, RPS3AP2, and AMD1P1 was established. The 10-pseudogenes signature was demonstrated to be an independent prognostic factor in patient with ovarian cancer in the random set (hazard ratio [HR] = 2.512, 95% confidence interval [CI] = 2.03-3.11, P < 0.001) and total set (HR = 1.71, 95% CI = 1.472-1.988, P < 0.001). When models integrating with age, grade, stage, and risk signature, the Area Under Curve (AUC) of the 1-year, 3-year, 5-year and 10-year Receiver Operating Characteristic curve in the random set and total set were 0.854, 0.824, 0.855, 0.805 and 0.679, 0.697, 0.739, 0.790, respectively. The results of functional enrichment analysis indicated that the underlying mechanisms by which these pseudogenes influence cancer prognosis may involve the immune-related biological processes and signaling pathways. Correlation analysis showed that risk signature was significantly correlated with immune cell infiltration and immune score. We identified a novel 10-pseudogenes signature to predict the survival of patients with ovarian cancer, and that may serve as novel possible prognostic biomarkers and therapeutic targets for ovarian cancer. [ABSTRACT FROM AUTHOR]

Pseudogenes were originally regarded as non-functional components scattered in the genome during evolution. Recent studies have shown that pseudogenes can be transcribed into long non-coding RNA and play a key role at multiple functional levels in different physiological and pathological processes. microRNAs (miRNAs) are a type of non-coding RNA, which plays important regulatory roles in cells. Numerous studies have shown that pseudogenes and miRNAs have interactions and form a ceRNA network with mRNA to regulate biological processes and involve diseases. Exploring the associations of pseudogenes and miRNAs will facilitate the clinical diagnosis of some diseases. Here, we propose a prediction model PMGAE (Pseudogene–MiRNA association prediction based on the Graph Auto-Encoder), which incorporates feature fusion, graph auto-encoder (GAE), and eXtreme Gradient Boosting (XGBoost). First, we calculated three types of similarities including Jaccard similarity, cosine similarity, and Pearson similarity between nodes based on the biological characteristics of pseudogenes and miRNAs. Subsequently, we fused the above similarities to construct a similarity profile as the initial representation features for nodes. Then, we aggregated the similarity profiles and associations of nodes to obtain the low-dimensional representation vector of nodes through a GAE. In the last step, we fed these representation vectors into an XGBoost classifier to predict new pseudogene–miRNA associations (PMAs). The results of five-fold cross validation show that PMGAE achieves a mean AUC of 0.8634 and mean AUPR of 0.8966. Case studies further substantiated the reliability of PMGAE for mining PMAs and the study of endogenous RNA networks in relation to diseases. [ABSTRACT FROM AUTHOR]

The epithelial sodium channel (ENaC) plays a key role in salt and water homeostasis in tetrapod vertebrates. There are four ENaC subunits (α, β, γ, δ), forming heterotrimeric αβγ- or δβγ-ENaCs. Although the physiology of αβγ-ENaC is well understood, for decades the field has stalled with respect to δβγ-ENaC due to the lack of mammalian model organisms. The SCNN1D gene coding for δ-ENaC was previously believed to be absent in rodents, hindering studies using standard laboratory animals. We analyzed all currently available rodent genomes and discovered that SCNN1D is present in rodents but was independently lost in five rodent lineages, including the Muridae (mice and rats). The independent loss of SCNN1D in rodent lineages may be constrained by phylogeny and taxon-specific adaptation to dry habitats, however habitat aridity does not provide a selection pressure for maintenance of SCNN1D across Rodentia. A fusion of two exons coding for a structurally flexible region in the extracellular domain of δ-ENaC appeared in the Hystricognathi (a group that includes guinea pigs). This conserved pattern evolved at least 41 Ma and represents a new autapomorphic feature for this clade. Exon fusion does not impair functionality of guinea pig (Cavia porcellus) δβγ-ENaC expressed in Xenopus oocytes. Electrophysiological characterization at the whole-cell and single-channel level revealed conserved biophysical features and mechanisms controlling guinea pig αβγ- and δβγ-ENaC function as compared with human orthologs. Guinea pigs therefore represent commercially available mammalian model animals that will help shed light on the physiological function of δ-ENaC. [ABSTRACT FROM AUTHOR]

The interferon-induced transmembrane (IFITM) gene family performs multiple functions in immunity, including inhibition of virus entry into cells. The IFITM repertoire varies widely between species and consists of protein-coding genes and pseudogenes. The selective forces driving pseudogenization within gene families are rarely understood. In this issue, the human pseudogene IFITM4P is characterized as a virus-induced, long noncoding RNA that contributes to restriction of influenza A virus by regulating mRNA levels of IFITM1, IFITM2, and IFITM3. [ABSTRACT FROM AUTHOR]

Processed pseudogenes may serve as a genetic reservoir for evolutionary innovation. Here, we argue that through the activity of long interspersed element‐1 retrotransposons, processed pseudogenes disperse coding and noncoding sequences rich with regulatory potential throughout the human genome. While these sequences may appear to be non‐functional, a lack of contemporary function does not prohibit future development of biological activity. Here, we discuss the dynamic evolution of certain processed pseudogenes into coding and noncoding genes and regulatory elements, and their implication in wide‐ranging biological and pathological processes. Also see the video abstract here: https://youtu.be/iUY_mteVoPI [ABSTRACT FROM AUTHOR]

Background: Pseudogenes are defined as key regulators in cancer initiation and progression. But their biological function and clinical significance in hepatocellular carcinoma (HCC) remain to be elucidated. In the current study, we identified a novel pseudogene, Annexin A2 pseudogene 1 (ANXA2P1), in HCC and explored its underlining molecular mechanism. Methods and Results: We analyzed the expression pattern of ANXA2P1 in a TCGA dataset and an HCC sample cohort and evaluated its clinical significance. The biological effects on HCC cells proliferation, migration, invasion and epithelial-mesenchymal transition (EMT) process were assessed by Cell Counting Kit-8 assay, Transwell assay and Western blot, respectively. The ANXA2P1/miR-376a-3p/ANXA2 axis was determined by bioinformatics analysis and dual-luciferase reporter assays. ANXA2P1 exerted as an oncogene that was significantly overexpressed in HCC tissues and was associated with disease progression and unfavorable prognosis of HCC patients. ANXA2P1 knockdown suppressed cell growth, cell migration and invasion and reversed EMT phenotype in HCC. Mechanistically, ANXA2P1 acts as a competing endogenous RNA for miR-376a-3p, thereby leading to the upregulation of its cognate gene ANXA2. Conclusions: ANXA2P1/miR-376a-3p/ANXA2 axis plays an important role in the progression of HCC. Our findings may provide valuable therapeutic target for treating HCC. [ABSTRACT FROM AUTHOR]

Pseudogenes have been classified as functionless and their annotation is an ongoing problem. The Adh6-ps1—a mouse pseudogene belonging to the alcohol dehydrogenase gene complex (Adh) was analyzed to review the conservation, homology, expression, and interactions and identify any role it plays in disease phenotypes using bioinformatics databases. Results showed that Adh6-ps1 have 2 transcripts (processed and unprocessed) which may have emerged from a transposition and duplication event, respectively, and that induced inversions (Uox gene, In(3)11Rk) involving gene complexes associated with Adh6-ps1 have been implicated in a diverse range of diseases. Adh6-ps1 is highly conserved in vertebrates particularly rodents and expressed in the liver. The top 5 MirRNA targets were Mir455, Mir511, Mir1903, Mir361, and Mir669o markers. While much is unknown about Mir1903 and Mir669o, the silencing of Mir455 and Mir511 is linked with hepatocellular carcinoma (HCC), and Mir361 is implicated in endometrial cancers. Given the identified MirRNA interactions with Adh6-ps1 and its expression in HCC and reproductive systems, it may well have a role in tumorigenesis and disease phenotypes. Nonetheless, further studies are required to establish these facts to add to the growing efforts to understand pseudogenes and their potential involvement in disease conditions. [ABSTRACT FROM AUTHOR]

Metabarcoding of Metazoa using mitochondrial genes may be confounded by both the accumulation of PCR and sequencing artefacts and the co‐amplification of nuclear mitochondrial pseudogenes (NUMTs). The application of read abundance thresholds and denoising methods is efficient in reducing noise accompanying authentic mitochondrial amplicon sequence variants (ASVs). However, these procedures do not fully account for the complex nature of concomitant sequences and the highly variable DNA contribution of specimens in a metabarcoding sample. We propose, as a complement to denoising, the metabarcoding Multidimensional Abundance Threshold Evaluation (metaMATE) framework, a novel approach that allows comprehensive examination of multiple dimensions of abundance filtering and the evaluation of the prevalence of unwanted concomitant sequences in denoised metabarcoding datasets. metaMATE requires a denoised set of ASVs as input, and designates a subset of ASVs as being either authentic (mitochondrial DNA haplotypes) or nonauthentic ASVs (NUMTs and erroneous sequences) by comparison to external reference data and by analysing nucleotide substitution patterns. metaMATE (i) facilitates the application of read abundance filtering strategies, which are structured with regard to sequence library and phylogeny and applied for a range of increasing abundance threshold values, and (ii) evaluates their performance by quantifying the prevalence of nonauthentic ASVs and the collateral effects on the removal of authentic ASVs. The output from metaMATE facilitates decision‐making about required filtering stringency and can be used to improve the reliability of intraspecific genetic information derived from metabarcode data. The framework is implemented in the metaMATE software (available at https://github.com/tjcreedy/metamate). see also the Perspective by Natalie R. Graham, Rosemary G. Gillespie and Henrik Krehenwinkel [ABSTRACT FROM AUTHOR]

Long read sequencing technologies have the potential to accurately detect and phase variation in genomic regions that are difficult to fully characterize with conventional short read methods. These difficult to sequence regions include several clinically relevant genes with highly homologous pseudogenes, many of which are prone to gene conversions or other types of complex structural rearrangements. We present PB-Motif, a new method for identifying rearrangements between two highly homologous genomic regions using PacBio long reads. PB-Motif leverages clustering and filtering techniques to efficiently report rearrangements in the presence of sequencing errors and other systematic artifacts. Supporting reads for each high-confidence rearrangement can then be used for copy number estimation and phased variant calling. First, we demonstrate PB-Motif's accuracy with simulated sequence rearrangements of PMS2 and its pseudogene PMS2CL using simulated reads sweeping over a range of sequencing error rates. We then apply PB-Motif to 26 clinical samples, characterizing CYP21A2 and its pseudogene CYP21A1P as part of a diagnostic assay for congenital adrenal hyperplasia. We successfully identify damaging variation and patient carrier status concordant with clinical diagnosis obtained from multiplex ligation-dependent amplification (MLPA) and Sanger sequencing. The source code is available at: github.com/zstephens/pb-motif. [ABSTRACT FROM AUTHOR]

Background: Pseudogenes show multiple functions in various cancer types, and immunotherapy is a promising cancer treatment. Therefore, this study aims to identify immune-related pseudogene signature in endometrial cancer (EC).Methods: Gene transcriptome data of EC tissues and corresponding clinical information were downloaded from The Cancer Genome Atlas (TCGA) through UCSC Xena browser. Spearman correlation analysis was performed to identify immune-related pseudogenes (IRPs) between the immune genes and pseudogenes. Univariate Cox regression, LASSO, and multivariate were performed to develop a risk score signature to investigate the different overall survival (OS) between high- and low-risk groups. The prognostic significance of the signature was assessed by the Kaplan-Meier curve, time-dependent receiver operating characteristic (ROC) curve. The abundance of 22 immune cell subtypes of EC patients was evaluated using CIBERSORT.Results: Nine IRPs were used to build a prognostic signature. Survival analysis revealed that patients in the low-risk group presented longer OS than those in the high-risk group as well as in multiple subgroups. The signature risk score was independent of other clinical covariates and was associated with several clinicopathological variables. The prognostic signature reflected infiltration by multiple types of immune cells and revealed the immunotherapy response of patients with anti-programmed death-1 (PD-1) and anti-programmed cell death 1 ligand 1 (PD-L1) therapy. Function enrichment analysis revealed that the nine IRPs were mainly involved in multiple cancer-related pathways.Conclusion: We identified an immune-related pseudogene signature that was strongly correlated with the prognosis and immune response to EC. The signature might have important implications for improving the clinical survival of EC patients and provide new strategies for cancer treatment. [ABSTRACT FROM AUTHOR]

Background: Four ohnologous genes (sox1, sox2, sox3, and sox15) were generated by two rounds of whole-genome duplication in a vertebrate ancestor. In eutherian mammals, Sox1, Sox2, and Sox3 participate in central nervous system (CNS) development. Sox15 has a function in skeletal muscle regeneration and has little functional overlap with the other three ohnologs. In contrast, the frog Xenopus laevis and zebrafish orthologs of sox15 as well as sox1-3 function in CNS development. We previously reported that Sox15 is involved in mouse placental development as neofunctionalization, but is pseudogenized in the marsupial opossum. These findings suggest that sox15 might have evolved with divergent gene fates during vertebrate evolution. However, knowledge concerning sox15 in other vertebrate lineages than therian mammals, anuran amphibians, and teleost fish is scarce. Our purpose in this study was to clarify the fate and molecular evolution of sox15 during vertebrate evolution. Results: We searched for sox15 orthologs in all vertebrate classes from agnathans to mammals by significant sequence similarity and synteny analyses using vertebrate genome databases. Interestingly, sox15 was independently pseudogenized at least twice during diversification of the marsupial mammals. Moreover, we observed independent gene loss of sox15 at least twice during reptile evolution in squamates and crocodile-bird diversification. Codon-based phylogenetic tree and selective analyses revealed an increased dN/dS ratio for sox15 compared to the other three ohnologs during jawed vertebrate evolution. Conclusions: The findings revealed an asymmetric evolution of sox15 among the four ohnologs during vertebrate evolution, which was supported by the increased dN/dS values in cartilaginous fishes, anuran amphibians, and amniotes. The increased dN/dS value of sox15 may have been caused mainly by relaxed selection. Notably, independent pseudogenizations and losses of sox15 were observed during marsupial and reptile evolution, respectively. Both might have been caused by strong relaxed selection. The drastic gene fates of sox15, including neofunctionalization and pseudogenizations/losses during amniote diversification, might be caused by a release from evolutionary constraints. [ABSTRACT FROM AUTHOR]

Background: Pseudogenes are non-functional copies of protein coding genes that typically follow a different molecular evolutionary path as compared to functional genes. The inclusion of pseudogene sequences in DNA barcoding and metabarcoding analysis can lead to misleading results. None of the most widely used bioinformatic pipelines used to process marker gene (metabarcode) high throughput sequencing data specifically accounts for the presence of pseudogenes in protein-coding marker genes. The purpose of this study is to develop a method to screen for nuclear mitochondrial DNA segments (nuMTs) in large COI datasets. We do this by: (1) describing gene and nuMT characteristics from an artificial COI barcode dataset, (2) show the impact of two different pseudogene removal methods on perturbed community datasets with simulated nuMTs, and (3) incorporate a pseudogene filtering step in a bioinformatic pipeline that can be used to process Illumina paired-end COI metabarcode sequences. Open reading frame length and sequence bit scores from hidden Markov model (HMM) profile analysis were used to detect pseudogenes. Results: Our simulations showed that it was more difficult to identify nuMTs from shorter amplicon sequences such as those typically used in metabarcoding compared with full length DNA barcodes that are used in the construction of barcode libraries. It was also more difficult to identify nuMTs in datasets where there is a high percentage of nuMTs. Existing bioinformatic pipelines used to process metabarcode sequences already remove some nuMTs, especially in the rare sequence removal step, but the addition of a pseudogene filtering step can remove up to 5% of sequences even when other filtering steps are in place. Conclusions: Open reading frame length filtering alone or combined with hidden Markov model profile analysis can be used to effectively screen out apparent pseudogenes from large datasets. There is more to learn from COI nuMTs such as their frequency in DNA barcoding and metabarcoding studies, their taxonomic distribution, and evolution. Thus, we encourage the submission of verified COI nuMTs to public databases to facilitate future studies. [ABSTRACT FROM AUTHOR]

Many early studies of ribosomal RNA gene (rDNA) suggested that rDNA tandem repeats within species are homogeneous. However, increasing number of reports have found intra-individual rDNA polymorphism across a range of taxa. Here, we reported a high level of intra-individual polymorphism of 18S-ITS1-5.8S rDNA in the genome of Cynoglossus melampetalus (Pleuronectiformes: Cynoglossidae), indicating a non-concerted evolution manner. Sequence alignments found two distinct types of 18S and 5.8S (Type A and B) and five types of ITS1 sequence (Type A - E) coexisted in the genome differing in length, GC content, secondary structure stability and minimum free energy. Based on the unique features of pseudogene and comparison of the conserved 18S rDNA sequence and 5.8S secondary structure of 22 flatfishes revealed that Type B sequences of 18S, 5.8S and their linked ITS1 were putative pseudogenes. So far, detection of rRNA pseudogenes from the multiple rDNA copies has been an intricate puzzle. Our results, as a result, provide a new ideal for rRNA pseudogene identification. • Five types of ITS1-5.8S-ITS2 sequence were found, indicating a non-concerted evolution pattern. • Type B sequences of 18S, 5.8S and their linked ITS1 were putative pseudogenes. • Variations in the conserved 18S and 5.8S revealed the putative pseudogenes. • ►These findings provide a new ideal for rRNA pseudogene identification. [ABSTRACT FROM AUTHOR]

Pseudogenes have been classified as functionless and their annotation is an ongoing problem. The Adh6-ps1-a mouse pseudogene belonging to the alcohol dehydrogenase gene complex (Adh) was analyzed to review the conservation, homology, expression and interactions, and identify any role it plays in disease phenotypes using bioinformatics databases. Results showed that Adh6-ps1 have 2 transcripts (processed and unprocessed) which may have emerged from a transposition and duplication event, respectively, and that induced inversions (Uox gene, In(3)11Rk) involving gene complexes associated with Adh6-ps1 have been implicated in a diverse range of diseases. Adh6-ps1 is highly conserved in vertebrates particularly rodents, and expressed in the liver. The top 5 MirRNA targets identified are Mir455, Mir511, Mir1903, Mir361, and Mir669o markers. While much is unknown about Mir1903 and Mir669o, the silencing of Mir455 and Mir511 is linked with hepatocellular carcinoma (HCC), and Mir361 is implicated in endometrial cancers. Given the identified MirRNA interactions with Adh6-ps1 and their expression in cancer phenotypes, and Adh6-ps1 associations with induced inversions, it may well have a role in tumorigenesis and disease phenotypes. Nonetheless, further studies are required to establish these facts to add to the growing efforts to understand pseudogenes and their potential involvement in disease conditions. [ABSTRACT FROM AUTHOR]

Numerous cancer drivers have been identified, but they are specific to a given cancer type and condition; universal cancer drivers and universal cancer mechanisms still remain largely unclear. Here, we identified the deadliest universal drivers for all cancers via developing algorithms to analyze massive RNAseqs and clinical data from The Cancer Genome Atlas (TCGA). In general, noncoding RNAs primarily serve as the most important inducers and suppressors for all types of cancers. In particular, pseudogenes are primary inducers, and specifically the antisense RNA RP11-335K5.2 serves as the most universal cancerous driver, independently of the cancer type and condition. Therefore, noncoding RNAs, instead of proteins as conventionally thought, primarily drive cancer, which establishes a novel field for future cancer research and therapy. [ABSTRACT FROM AUTHOR]

We take advantage of synteny blocks, the analytical construct enabled at the evolutionary moment of speciation or polyploidization, to follow the independent loss of duplicate genes in two sister species or the loss through fractionation of syntenic paralogs in a doubled genome. By examining how much sequence remains after a contiguous series of genes is deleted, we find that this residue remains at a constant low level independent of how many genes are lost—there are few if any relics of the missing sequence. Pseudogenes are rare or extremely transient in this context. The potential exceptions lie exclusively with a few examples of speciation, where the synteny blocks in some larger genomes tolerate degenerate sequence during genomic divergence of two species, but not after whole genome doubling in the same species where fractionation pressure eliminates virtually all non-coding sequence. [ABSTRACT FROM AUTHOR]

Pseudogenes, which are long noncoding RNAs that originate from protein-coding genes, have been suggested to play important roles in disease. Although studies have revealed high expression of legumain (LGMN) in many types of tumors, the regulation of LGMN remains largely unknown. Here, we found that a novel LGMN pseudogene (LGMNP1) was upregulated in glioblastoma (GBM) tissues and high LGMNP1 expression in GBM cells enhanced proliferation and invasion. Biochemical analysis showed that cytoplasmic LGMNP1 functionally targeted miR-495-3p in a manner involving an RNA-induced silencing complex. Dual-luciferase reporter assays demonstrated that LGMN was a target of miR-495-3p, and LGMN was upregulated and positively correlated with LGMNP1 in GBM. Moreover, miR-495-3p was downregulated and negatively correlated with LGMNP1 in GBM tissues. Notably, the tumor-promoting effects of LGMNP1 upregulation could be alleviated by miR-495-3p mimics. Furthermore, GBM cells overexpressing LGMNP1 exhibited more aggressive tumor progression and elevated LGMN expression in vivo. Thus, our data illustrate that LGMNP1 exerts its oncogenic activity, at least in part, as a competitive endogenous RNA (ceRNA) that elevates LGMN expression by sponging miR-495-3p. CeRNA-mediated miRNA sequestration might be a novel therapeutic strategy in GBM. [ABSTRACT FROM AUTHOR]

The incidence and mortality rate of hepatocellular carcinoma (HCC) nowadays is still at high levels. The regulatory roles of pseudogene in cancers have been gradually recognized in recent years. However, comprehensive investigation of abnormally expressed pseudogene and related mechanisms in HCC remains lacking. GSE124535 dataset was used to identify differentially expressed pseudogenes in HCC tissues compared with normal tissues. Prognostic value of these differentially expressed pseudogenes was analyzed at GEPIA. StarBase used to analyze microRNAs (miRNAs) can bind with pseudogene, while the targets for these miRNAs were analyzed at miRTarBase. Protein–protein interaction (PPI) network was then established for miRNA targets, after that hub genes were selected. Expression correlation of pseudogene and hub genes was analyzed at StarBase. In total, 16 upregulated and 17 downregulated pseudogenes were identified. Pseudogene HSPB1P1 was identified abnormally expressed in 20 types of human cancers and could be used as an indicator for poorer overall survival of patients with HCC. Functional analyses showed that HSPB1P1 was strongly correlated with signaling pathways related to cancer progression. Further studied revealed that HSPB1P1 could direct regulate the EZH2 expression in HCC. In summary, our study indicated that HSPB1P1 was a predictor for poorer overall survival of patients with HCC and may be potential therapeutic target against HCC. [ABSTRACT FROM AUTHOR]

Pseudogenes are indicating more and more functional potentials recently, though historically were regarded as relics of evolution. Computational methods for predicting pseudogene functions on Gene Ontology is important for directing experimental discovery. However, no pseudogene-specific computational methods have been proposed to directly predict their Gene Ontology (GO) terms. The biggest challenge for pseudogene function prediction is the lack of enough features and functional annotations, making training a predictive model difficult. Considering the close functional similarity between pseudogenes and their parent coding genes that share great amount of DNA sequence, as well as that coding genes have rich annotations, we aim to predict pseudogene functions by borrowing information from coding genes in a graph-based way. Here we propose Pseudo2GO, a graph-based deep learning semi-supervised model for pseudogene function prediction. A sequence similarity graph is first constructed to connect pseudogenes and coding genes. Multiple features are incorporated into the model as the node attributes to enable the graph an attributed graph, including expression profiles, interactions with microRNAs, protein-protein interactions (PPIs), and genetic interactions. Graph convolutional networks are used to propagate node attributes across the graph to make classifications on pseudogenes. Comparing Pseudo2GO with other frameworks adapted from popular protein function prediction methods, we demonstrated that our method has achieved state-of-the-art performance, significantly outperforming other methods in terms of the M-AUPR metric. [ABSTRACT FROM AUTHOR]

DNA barcoding has been actively used as a method for species identification, and it will become an increasingly important method in the future. However, DNA barcoding can occasionally encounter a major problem, namely "nuclear mitochondrial DNA pseudogenes (NUMTs)". In this paper, we report the discovery of a pseudogene in the mitochondrial COI region from aquatic heteropterans. PCR with Folmer's universal primer set produced two different lengths of DNA sequences: a standard 658 bp sequence and a slightly shorter 652 bp sequence. The presence of an "indel" and "stop codon" in the shorter sequence made them separable. We recommend using the "HCOoutout" primer as a downward primer for amplification of the COI region of aquatic heteropterans, especially corixid water bugs. [ABSTRACT FROM AUTHOR]

Background: Plastid gene loss and pseudogenization has been widely documented in parasitic and mycoheterotrophic plants, which have relaxed selective constraints on photosynthetic function. More enigmatic are sporadic reports of pseudogenization and loss of important photosynthesis genes in lineages thought to be fully photosynthetic. Here we report the complete plastid genome of Saniculiphyllum guangxiense, a critically endangered and phylogenetically isolated plant lineage, along with genomic evidence of reduced chloroplast function. We also report 22 additional plastid genomes representing the diversity of its containing clade Saxifragales, characterizing gene content and placing variation in a broader phylogenetic context. Results: We find that the plastid genome of Saniculiphyllum has experienced pseudogenization of five genes of the ndh complex (ndhA, ndhB, ndhD, ndhF, and ndhK), previously reported in flowering plants with an aquatic habit, as well as the surprising pseudogenization of two genes more central to photosynthesis (ccsA and cemA), contrasting with strong phylogenetic conservatism of plastid gene content in all other sampled Saxifragales. These genes participate in photooxidative protection, cytochrome synthesis, and carbon uptake. Nuclear paralogs exist for all seven plastid pseudogenes, yet these are also unlikely to be functional. Conclusions: Saniculiphyllum appears to represent the greatest degree of plastid gene loss observed to date in any fully photosynthetic lineage, perhaps related to its extreme habitat specialization, yet plastid genome length, structure, and substitution rate are within the variation previously reported for photosynthetic plants. These results highlight the increasingly appreciated dynamism of plastid genomes, otherwise highly conserved across a billion years of green plant evolution, in plants with highly specialized life history traits. [ABSTRACT FROM AUTHOR]

Background: We aimed to identify differentially expressed pseudogenes and explore their potential functions in four types of common gynecological malignancies (e.g., cervical squamous cell carcinoma, ovarian serous cystadenocarcinoma, uterine corpus endometrial carcinoma, and uterine carcinosarcoma) using bioinformatics technology. Materials and methods: We identified up-regulated and down-regulated pseudogenes and built a pseudogene-miRNA-mRNA regulatory network through public datasets to explore their potential functions in carcinogenesis and cancer prognosis. Results: Among the 63 up-regulated pseudogenes identified, LDHAP5 demonstrated the greatest potential as a candidate pseudogene due to its significant association with poor overall survival in ovarian serous cystadenocarcinoma. KEGG pathway analysis revealed that LDHAP5 showed significant enrichment in MicroRNAs in cancer, Pathway in cancer and PI3K-AKT signaling pathway. Further analysis revealed that EGFR was the potential target mRNA of LDHAP5, which may play an important role in ovarian serous cystadenocarcinoma. Conclusions: LDHAP5 was associated with the occurrence and prognosis of ovarian serous cystadenocarcinoma, and thus shows potential as a novel therapeutic target against such cancer. [ABSTRACT FROM AUTHOR]

Head and neck squamous cell carcinoma (HNSCC) is the sixth malignancy, which is characterized by poor prognosis or high mortality because of the lack of predicting markers. Aberrant cancer pseudogenes have been found predictive for prognosis. We aim to identify a pseudogene-based prognosis signature for HNSCC by machine learning. RNA-seq data were downloaded from The Cancer Genome Atlas, and 700 differentially-expressed pseudogenes were identified. The survival-related pseudogenes were screened through COX-regression analysis, which includes univariate regression, least absolute shrinkage and selection operator regression, and multivariate regression, and a five-pseudogene signature was constructed. The value of prediction for the signature was validated in multiple subgroups in terms of survival. Gene set enrichment analysis (GSEA) and coexpression analysis were used to determine the underlying biological functions. Seven hundred dysregulated pseudogenes were identified, and the five-pseudogene signature can distinguish the low-risk and high-risk patients for both training and testing sets and predicted prognosis with high sensitivity and specificity. Furthermore, the signature was applicable to patients of different genders, ages, stages, and grades. Coexpression analysis revealed that the five-pseudogene is associated with immune system. GSEA showed cancer-related biological process and pathways the five-pseudogene involved in. The five-pseudogene signature is not only a novel marker for prognosis but also a promising signature for monitoring therapeutic schedule. Therefore, our findings may have potential clinical significance. [ABSTRACT FROM AUTHOR]

Background: Glioma is the most common primary brain tumor with a dismal prognosis. It is urgent to develop novel molecular biomarkers and conform to individualized schemes. Methods: Differentially expressed pseudogenes between low grade glioma (LGG) and glioblastoma multiforme (GBM) were identified in the training cohort. Least absolute shrinkage and selection operator (LASSO) regression and multivariate Cox proportional hazards regression analyses were used to select pseudogenes associated with prognosis of glioma. A risk signature was constructed based on the selected pseudogenes for predicting the survival of glioma patients. A pseudogene-miRNA-mRNA regulatory network was established and visualized using Cytoscape 3.5.1. Gene Oncology (GO) and signaling pathway analyses were performed on the targeted genes to investigate functional roles of the risk signature. Results: Five pseudogenes (ANXA2P2, EEF1A1P9, FER1L4, HILS1, and RAET1K) correlating with glioma survival were selected and used to establish a risk signature. Time-dependent receiver operating characteristic (ROC) curves revealed that the risk signature could accurately predict the 1, 3, and 5-year survival of glioma patients. GO and signaling pathway analyses showed that the risk signature was involved in regulation of proliferation, migration, angiogenesis, and apoptosis in glioma. Conclusions: In this study, a risk signature with five pseudogenes was constructed and shown to accurately predict 1-, 3-, and 5-year survival for glioma patient. The risk signature may serve as a potential target against glioma. [ABSTRACT FROM AUTHOR]

Previous researches have provided evidence for the significant involvement of pseudogenes in immune-related functions across different types of cancer. However, the mechanisms by which pseudogenes regulate immunity in ovarian cancer (OV) and their potential impact on clinical outcomes remain unclear. To address this gap in knowledge, our study utilized a novel computational framework to analyze a total of 491 samples from three public datasets. We employed a combination of 10 machine-learning algorithms to construct a signature known as the tumor-infiltrating immune cells-related pseudogenes signature (TIICPS). The TIICPS, consisting of 12 pseudogenes, demonstrated independent prognostic value for overall survival, surpassing conventional clinical traits, 62 published signatures, and TP53 and BRCA mutation status in three cohorts. Patients with low TIICPS exhibited heightened immune-related pathways, intricate genomic alterations, substantial immune infiltration, and greater potential for immunotherapy efficacy. Consequently, TIICPS holds promise as a predictive tool for prognosis and immunotherapy response in ovarian cancer. • We constructed a tumor-infiltrating immune cells-related pseudogenes signature (TIICPS) based on integrative machine learning. • TIICPS as an independent prognostic indicator is superior to clinical traits, 62 published signatures, and TP53 and BRCA mutation. • Patients with low TIICPS are featured by substantial immune infiltration and potential sensitivity for immunotherapy. [ABSTRACT FROM AUTHOR]

Efforts to understand the genetic basis of evolutionary change have concentrated on proteins and their encoding DNA sequences. These studies have brought to light patterns and processes at the nucleotide level, yet the complex functional relationships between genetic variants and phenotypes remain poorly known. The realization that even a complete description of proteins and the effects of their activity will not suffice to understand the conditions under which they are time- and tissue-specifically expressed or repressed during development has refocused attention on cis-regulatory regions. In particular, promoter sequences are thought to hold the key for understanding the evolution of phenotypic differences between species. This is because of their complex organization into independent modules such that, unlike coding sequences in which mutations affect protein function every time the protein is expressed, mutations in cis-regulatory sequences may have minor or no pleiotropic effects. Complex information-encoding makes cis-regulatory regions poorly amenable to comparative methods designed for coding sequences. Some general conclusions are emerging as to how genetic variation is distributed across regulatory networks and the processes modulating the structure of this variation. We bring into this emerging scenario several recent findings pointing to different ways in which spliceosomal introns, pseudogenes and patterns of point mutation can be active players for the evolution of novel transcriptional profiles.

• Korean Salmonella biovar Gallinarum (SG) extended polyproline linker (PPL) of SpvB. • Korean SG isolates share specific pseudogenes and constitute an unique cluster. • Early outbreaks of fowl typhoid in Korea was caused by two highly related subgroups. • Korean SG evolved to 10 subgroups by changing PPL length and pseudogene acquisition. Salmonella enterica subsp. enterica serovar Gallinarum biovar Gallinarum (SG) causes fowl typhoid (FT) and substantial economic loss in Korea due to egg drop syndrome and mortality. Despite the extensive use of vaccines, FT still occurs in the field. Therefore, the emergence of more pathogenic SG or the recovered pathogenicity of a vaccine strain has been suspected. SpvB, an ADP-ribosyl transferase, is a major pathogenesis determinant, and the length of the polyproline linker (PPL) of SpvB affects pathogenic potency. SG strains accumulate pseudogenes in their genomes during host adaptation, and pseudogene profiling may provide evolutionary information. In this study, we found that the PPL length of Korean SG isolates varied from 11 to 21 prolines and was longer than that of a live vaccine strain, SG 9R (9 prolines). According to growth competition in chickens, the growth of an SG isolate with a PPL length of 17 prolines exceeded that of an SG isolate with a PPL length of 15 prolines. We investigated the pseudogenes of the field isolates, SG 9R and reference strains in GenBank by resequencing and comparative genomics. The pseudogene profiles of the field isolates were notably different from those of the foreign SG strains, and they were subdivided into 7 pseudogene subgroups. Collectively, the field isolates had gradually evolved by changing PPL length and acquiring additional pseudogenes. Thus, the characterization of PPL length and pseudogene profiling may be useful to understand the molecular evolution of SG and the epidemiology of FT. [ABSTRACT FROM AUTHOR]

Escherichia coli has approximately 4300 open reading frames and about 178 of them are annotated as pseudogenes. The existence of the pseudogenes in E. coli had raised the question of whether they contribute to any cell function. Recently, several pseudogenes have been studied; however, these genes are still considered rare and tend to be ignored as so-called junk. Our recent study has shown several pseudogenes in E. coli are essential in hydrogen metabolism. ydfW is one of the pseudogenes associated with hydrogen metabolism. Here, we found carbon dioxide and nitrate in the fermentation medium of mutant ydfW suggesting that this strain may induce nitrate pathways. The Fdh-N complex is comprised of FdnG, FdnH, and FdnI subunits which correspond to the nitrate-nitrite pathway. This finding is supported by transcription analysis that shows formate dehydrogenase (fdnG) and nitrate reductase (narG) are significantly induced (25- and 18-fold, respectively). FdnG and NarG play a significant role in the reduction of nitrate into nitrite; hence, their genes are induced by the build-up of nitrate. Based on in silico docking analysis, we predict that YdfW is covalently bound to the Fdh-N complex in the cytoplasm. Hence, deletion of ydfW is expected has influenced the nitrate-nitrite pathway. Subsequently, the situation will affect the hydrogen production in E. coli. • Escherichia coli pseudogene protein YdfW has been identified as essential in hydrogen metabolism. • Mutant ydfW ceased hydrogen production from glucose and formate. • ydfW mutation causes induction of respiratory nitrate reductase (NarG). • YdfW is predicted to play a role in nitrate respiration pathways. [ABSTRACT FROM AUTHOR]

Background: Salmonella enterica consists of over 2500 serovars and displays dichotomy in disease manifestations and host range. Except for the enrichment of pseudogenes in genomes for human-restricted serovars, no hallmark has been identified to distinguish those with host-generalist serovars. The serovar Sendai is rare and human-restricted. Notably, it exhibits an O, H antigen formula as the host-generalist serovar Miami. Results: We sequenced the complete genomes of the two serovars Sendai and Miami. Analysis at both nucleotide identity and gene content level demonstrates the same high degree of similarity between Sendai and Paratyphi A, but their distinct CRISPR spacers suggests a recent divergence history. A frameshift mutation occurred in rfbE for the entire lineage of Paratyphi A but not in Sendai, which may explain their distinct O antigens. The nucleotide sequence of Miami's fliC is nearly identical to Sendai's. The incongruent phylogeny of this gene with that of the adjacent genes suggests a recombination event responsible for Sendai and Miami possessing the same H antigen. Sendai's even greater number of pseudogenes than that of Paratyphi A and Typhi indicates its undergoing continued genomic degradation. The phylogenetically distinct human-restricted serovars/strains share pseudogenes with the same inactivation mutations, therefore suggesting that recombination may have occurred and have been facilitated by their overlap in niches. Conclusions: Analysis of Sendai's genome and comparison with others reflect the finer evolutionary signatures of Salmonella in the process of niches changing from facultative to obligate parasite. [ABSTRACT FROM AUTHOR]

Abstract Highly conserved 18S rDNA sequences encode ribosomal RNA and evolve in a concerted manner. In this study, 178 sequences of 18S rDNA from the ridge-eyed flounder, Pleuronichthys cornutus , were analyzed. The total sequences yielded five distinguishable types of 18S rDNA—A, B, R, S, and L—that were defined based on sequence alignments, clone clustering, and recombination detection. The length of 176 clones ranged from 1838 bp to 1846 bp, with one particularly short clone reaching only 1466 bp and one long clone reaching up to1869 bp. As per current criteria for pseudogene inference, Type S was inferred to be a pseudogene due to its truncated length (380 bp) and low minimum free energy (−536.9 kcal·mol−1). Type L had a unique 20-bp insertion and was also predicted to be as a pseudogene. Types A and B showed 31 differential sites, which in Type A was consistent with 18S rDNA sequences found in six other flounders, while Type B and recombinant Type R were not. Maximum K2P genetic distances were calculated within Type B (0.0232) and R (0.0313); these were much higher than that of Type A (0.0093) or between the six flounders (0.011). Only Type A was detected by reverse transcription experiments and linked to functional ITS1 and 5.8S fragments; Types B and R were linked to both functional and pseudo genes. Accordingly, Type A is likely the functional gene, and Types B and R are probable pseudogenes. The heterogeneity of the five types revealed that 18S rDNA sequences evolved in a non-concerted evolution manner in P. cornutus. Furthermore, our results suggest that more features should be used to identify functional or pseudo genes. Highlights • The 18S rDNA in Pleuronichthys cornutus evolved in non-concerted evolution. • Five types of 18S rDNA were defined including length variation and recombination events. • Four more characteristics were suggested for rDNA pseudogene inference except the previous several criteria. [ABSTRACT FROM AUTHOR]

Pseudogene is a gene copy that has lost its original function. For a long time, pseudogenes have been considered as “junk DNA” that inevitably arises as a result of ongoing evolutionary process. However, experimental data obtained during recent years indicate this understanding of the nature of pseudogenes is not entirely correct, and many pseudogenes perform important genetic functions. In the review, we have addressed classification of pseudogenes, methods of their detection in the genome, and the problem of their evolutionary conservatism and prevalence among species belonging to different taxonomic groups in the light of modern data. The mechanisms of gene expression regulation by pseudogenes and the role of pseudogenes in pathogenesis of various human diseases are discussed. [ABSTRACT FROM AUTHOR]

The rapid development of CRISPR technology is revolutionizing molecular approaches to the dissection of complex biological phenomena. Here we describe an alternative generally applicable implementation of the CRISPR-Cas9 system that allows for selective knockdown of extremely homologous genes. This strategy employs the lentiviral delivery of paired sgRNAs and nickase Cas9 (Cas9D10A) to achieve targeted deletion of splice junctions. This general strategy offers several advantages over standard single-guide exon-targeting CRISPR-Cas9 such as greatly reduced off-target effects, more restricted genomic editing, routine disruption of target gene mRNA expression and the ability to differentiate between closely related genes. Here we demonstrate the utility of this strategy by achieving selective knockdown of the highly homologous human genes FOXO3A and suspected pseudogene FOXO3B . We find the spJCRISPR strategy to efficiently and selectively disrupt FOXO3A and FOXO3B mRNA and protein expression; thus revealing that the human FOXO3B locus encodes a bona fide human gene. Unlike FOXO3A, we find the FOXO3B protein to be cytosolically localized in both the presence and absence of active Akt. The ability to selectively target and efficiently disrupt the expression of the closely-related FOXO3A and FOXO3B genes demonstrates the efficacy of the spJCRISPR approach. [ABSTRACT FROM AUTHOR]

Detection of evolutionary shifts in sensory systems is challenging. By adopting a molecular approach, our earlier study proposed a sensory trade-off hypothesis between a loss of colour vision and an origin of high-dutycycle (HDC) echolocation in Old World bats. Here, we test the hypothesis in New World bats, which include HDC echolocators that are distantly related to Old World HDC echolocators, as well as vampire bats, which have an infrared sensory system apparently unique among bats. Through sequencing the short-wavelength opsin gene (SWS1) in 16 species (29 individuals) of New World bats, we identified a novel SWS1 polymorphism in an HDC echolocator: one allele is pseudogenized but the other is intact, while both alleles are either intact or pseudogenized in other individuals. Strikingly, both alleles were found to be pseudogenized in all three vampire bats. Since pseudogenization, transcriptional or translational changes could separately result in functional loss of a gene, a pseudogenized SWS1 indicates a loss of dichromatic colour vision in bats. Thus, the same sensory trade-off appears to have repeatedly occurred in the two divergent lineages of HDC echolocators, and colour vision may have also been traded off against the infrared sense in vampire bats. [ABSTRACT FROM AUTHOR]

DNA repair pathways are essential for cellular survival as our DNA is constantly under assault from both exogenous and endogenous DNA damaging agents. Five major mammalian DNA repair pathways exist within a cell to maintain genomic integrity. Of these, the DNA mismatch repair (MMR) pathway is highly conserved among species and is well documented in bacteria. In humans, the importance of MMR is underscored by the discovery that a single mutation in any 1 of 4 genes within the MMR pathway (MLH1, MSH2, MSH6 and PMS2) results in Lynch syndrome (LS). LS is a autosomal dominant condition that predisposes individuals to a higher incidence of many malignancies including colorectal, endometrial, ovarian, and gastric cancers. In this review, we discuss the role of PMS2 in the MMR pathway, the evolving testing criteria used to identify variants in the PMS2 gene, the LS phenotype as well as the autosomal recessive condition called constitutional mismatch repair deficiency syndrome, and current methods used to elucidate the clinical impact of PMS2 mutations. [ABSTRACT FROM AUTHOR]

Heterotrophic plants provide evolutionarily independent, natural experiments in the genomic consequences of radically altered nutritional regimes. Here, we have sequenced and annotated the plastid genome of the endangered mycoheterotrophic orchid Hexalectris warnockii. This orchid bears a plastid genome that is ∼80% the total length of the leafy, photosynthetic Phalaenopsis , and contains just over half the number of putatively functional genes of the latter. The plastid genome of H. warnockii bears pseudogenes and has experienced losses of genes encoding proteins directly (e.g., psa/psb, rbcL) and indirectly involved in photosynthesis (atp genes), suggesting it has progressed beyond the initial stages of plastome degradation, based on previous models of plastid genome evolution. Several dispersed and tandem repeats were detected, that are potentially useful as conservation genetic markers. In addition, a 29-kb inversion and a significant contraction of the inverted repeat boundaries are observed in this plastome. The Hexalectris warnockii plastid genome adds to a growing body of data useful in refining evolutionary models in parasites, and provides a resource for conservation studies in these endangered orchids. [ABSTRACT FROM AUTHOR]

The enzyme CMP-N-acetylneuraminic acid hydroxylase (CMAH) is responsible for the synthesis of N-glycolylneuraminic acid (Neu5Gc), a sialic acid present on the cell surface proteins of most deuterostomes. The CMAH gene is thought to be present in most deuterostomes, but it has been inactivated in a number of lineages, including humans. The inability of humans to synthesize Neu5Gc has had several evolutionary and biomedical implications. Remarkably, Neu5Gc is a xenoantigen for humans, and consumption of Neu5Gc-containing foods, such as red meats, may promote inflammation, arthritis, and cancer. Likewise, xenotransplantation of organs producing Neu5Gc can result in inflammation and organ rejection. Therefore, knowing what animal species contain a functional CMAHgene, andare thus capable ofendogenousNeu5Gcsynthesis,has potentially far-reachingimplications. In addition to humans, other lineages are known, or suspected, to have lost CMAH; however, to date reports of absent and pseudogenicCMAHgenes are restricted to ahandful of species. Here, we analyze all available genomicdata for nondeuterostomes, and 322 deuterostome genomes, to ascertain the phylogenetic distribution of CMAH. Among nondeuterostomes,we found CMAH homologs in two green algae and a few prokaryotes. Within deuterostomes, putatively functional CMAH homologs are present in 184 of the studied genomes, and a total of 31 independent gene losses/pseudogenization events were inferred.Our work produces a list of animals inferred to be free from endogenous Neu5Gc based on the absence of CMAH homologs and are thus potential candidates for human consumption, xenotransplantation research, and model organisms for investigation of human diseases. [ABSTRACT FROM AUTHOR]