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Pseudoalteromonas viridis strain BBR56 was isolated from seawater at Dutungan Island, South Sulawesi, Indonesia. Bacterial DNA was isolated using Promega Genomic DNA TM050. DNA purity and quantity were assessed using NanoDrop spectrophotometers and Qubit fluorometers. The DNA library and sequencing were prepared using Oxford Nanopore Technology GridION MinKNOW 20.06.9 with long read, direct, and comprehensive analysis. High accuracy base calling was assessed with Guppy version 4.0.11. Filtlong and NanoPlot were used for filtering and visualizing the FASTQ data. Flye (2.8.1) was used for de novo assembly analysis. Variant calls and consensus sequences were created using Medaka. The annotation of the genome was elaborated by DFAST. The assembled genome and annotation were tested using Busco and CheckM. Herein, we found that the highest similarity of the BBR56 isolate was 98.37% with the 16 S rRNA gene sequence of P. viridis G-1387. The genome size was 5.5 Mb and included chromosome 1 (4.2 Mbp) and chromosome 2 (1.3 Mbp), which encoded 61 pseudogenes, 4 noncoding RNAs, 113 tRNAs, 31 rRNAs, 4,505 coding DNA sequences, 4 clustered regularly interspaced short palindromic repeats, 4,444 coding genes, and a GC content of 49.5%. The sequence of the whole genome of P. viridis BBR56 was uploaded to GenBank under the accession numbers CP072425-CP072426, biosample number SAMN18435505, and bioproject number PRJNA716373. The sequence read archive (SRR14179986) was successfully obtained from NCBI for BBR56 raw sequencing reads. Digital DNA-DNA hybridization results showed that the genome of BBR56 had the potential to be a new species because no other bacterial genomes were similar to the sample. Biosynthetic gene clusters (BGCs) were assessed using BAGEL4 and the antiSMASH bacterial version. The genome harbored diverse BGCs, including genes that encoded polyketide synthase, nonribosomal peptide synthase, RiPP-like, NRP-metallophore, hydrogen cyanide, betalactone, thioamide-NRP, Lant class I, sactipeptide, and prodigiosin. Thus, BBR56 has considerable potential for further exploration regarding the use of its secondary metabolite products in the human and fisheries sectors., (© 2024. The Author(s).)

Glioblastoma (GBM) is the most aggressive and frequent type of primary brain cancer in adult patients. One of the key molecular features associated with GBM pathogenesis is the dysfunction of PTEN oncosuppressor. In addition to PTEN gene, humans and several primates possess processed PTEN pseudogene (PTENP1) that gives rise to long non-coding RNA lncPTENP1-S. Regulation and functions of PTEN and PTENP1 are highly interconnected, however, the exact molecular mechanism of how these two genes affect each other remains unclear. Here, we analyzed the methylation level of the CpG islands (CpGIs) in the promoter regions of PTEN and PTENP1 in patient-derived GBM neurospheres. We found that increased PTEN methylation corelates with decreased PTEN mRNA level. Unexpectedly, we showed the opposite trend for PTENP1. Using targeted methylation and demethylation of PTENP1 CpGI, we demonstrated that DNA methylation increases lncPTENP1-S expression in the presence of wild type PTEN protein but decreases lncPTENP1-S expression if PTEN protein is absent. Further experiments revealed that PTEN protein binds to PTENP1 promoter region and inhibits lncPTENP1-S expression if its CpGI is demethylated. Interestingly, we did not detect any effect of lncPTENP1-S on the level of PTEN mRNA, indicating that in GBM cells PTENP1 is a downstream target of PTEN rather than its upstream regulator. Finally, we studied the functions of lncPTENP1-S and demonstrated that it plays a pro-oncogenic role in GBM cells by upregulating the expression of cancer stem cell markers and decreasing cell adhesion., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier B.V. and Société Française de Biochimie et Biologie Moléculaire (SFBBM). All rights reserved.)

Background: 21-Hydroxylase deficiency (21-OHD) is caused by pathogenic CYP21A2 variations. CYP21A2 is arranged in tandem with its highly homologous pseudogene CYP21A1P; therefore, it is prone to mismatch and rearrangement, producing different types of complex variations. There were few reports on using only one method to detect different CYP21A2 variants simultaneously., Aims: Targeted long-read sequencing method was used to detect all types of CYP21A2 variants in a series of patients with 21-OHD., Methods: A total of 59 patients with 21-OHD were enrolled from Peking Union Medical College Hospital. Long-range locus-specific PCR and long-read sequencing (LRS) were performed to detect the pathogenic variants in CYP21A2., Results: Copy-number variants of CYP21A2 were found in 25.4% of patients, including 5.1% with 3 copies of CYP21A2, 16.9% with 1 copy of CYP21A2, and 3.4% with 0 copy of CYP21A2. The remaining 74.6% of patients had 2 copies of CYP21A2. Pathogenic variants were identified in all 121 alleles of 59 patients. Specifically, single-nucleotide variants and small insertions/deletions (< 50 bp) were detected in 79 alleles, of which conversed from CYP21A1P were detected in 63 alleles, and rare variants were found in the other 16 alleles. Large gene conversions (> 50 bp) from pseudogene were detected in 10 alleles, and different chimeric genes (CYP21A1P/CYP21A2 or TNXA/TNXB) formed by large deletions were detected in 32 alleles. Of all variants, p.I173N was the most common variant (19.0%)., Conclusions: Our study demonstrated that targeted long-read sequencing is a comprehensive method for detecting CYP21A2 variations, which is helpful for genetic diagnosis in 21-OHD patients., (© 2023. The Author(s), under exclusive licence to Italian Society of Endocrinology (SIE).)

Mallotus paniculatus (Lam.) Müll. Arg. 1865 (Euphorbiaceae) is a shrub or small tree with medicinal properties that is distributed across Southeast Asia. In this study, we sequenced the complete chloroplast genome of M. paniculatus to study phylogenetic relationships within the family Euphorbiaceae Juss. The complete chloroplast genome of M. paniculatus was 164,455 bp in length, with an overall GC content of 35.3%. It was found to consist of a long single copy region of 89,021 bp, a small single copy region of 18,524 bp, and a pair of inverted repeats of 28,455 bp. Results indicated that the chloroplast genome contains a total of 131 genes, including 78 protein-coding genes, 37 tRNA genes, eight rRNA genes, and eight pseudogenes. The phylogenetic tree showed that M. paniculatus is closely related to Mallotus japonicus and Mallotus peltatus. [ABSTRACT FROM AUTHOR]

Gynura japonica (Asteraceae) is a folk herbal medicine with multi-pharmacological functions involving analgesic, hemostatic and antiangiogenic activities. The study was conducted to assemble the complete chloroplast (cp) genome of G. japonica through a genome-skimming approach. The assembled cp genome was 151,023 bp in size, with 62.8% AT content, consisting of a large single copy (LSC) of 83,185 bp, two copies of inverted repeat (IRa and IRb) of 24,847 bp, and a small single copy (SSC) of 18,144 bp. The cp genome of G. japonica contained 133 genes, including eight ribosomal RNA genes (rRNAs), 37 transfer RNA genes (tRNAs), 86 protein-coding genes (PCGs), and two pseudogenes (ψycf1 and ψrps19). Our phylogenomic analysis based on whole plastid genomes strongly supports G. japonica is a sister to the clade including Crassocephalum crepidioides and Jacobaea vulgaris. [ABSTRACT FROM AUTHOR]

We report the complete chloroplast genome (plastome) sequences of Pedicularis cephalantha (147,087 bp) and P. nigra (145,726 bp), endemic to southwestern China. Both plastomes have typical quadripartite structures with one large-single copy region, one small-single copy region, and two inverted repeat regions. Both plastome sequences contained 37 tRNA genes and eight rRNA genes, but they differed in the numbers of protein-coding genes: P. cephalantha had 76 functional genes and 12 pseudogenes while P. nigra had 74 functional genes and 13 pseudogenes. Phylogenetic analysis shows that P. cephalantha and P. nigra are closely related, then sister to P. oederi in the family Orobanchaceae. [ABSTRACT FROM AUTHOR]

Genetic degeneration is an extraordinary feature of sex chromosomes, with the loss of functions of Y-linked genes in species with XY systems, and W-linked genes in ZW systems, eventually affecting almost all genes. Although degeneration is familiar to most biologists, important aspects are not yet well understood, including how quickly a Y or W chromosome can become completely degenerated. I review the current understanding of the time-course of degeneration. Degeneration starts after crossing over between the sex chromosome pair stops, and theoretical models predict an initially fast degeneration rate and a later much slower one. It has become possible to estimate the two quantities that the models suggest are the most important in determining degeneration rates—the size of the sex-linked region, and the time when recombination became suppressed (which can be estimated using Y–X or W–Z sequence divergence). However, quantifying degeneration is still difficult. I review evidence on gene losses (based on coverage analysis) or loss of function (by classifying coding sequences into functional alleles and pseudogenes). I also review evidence about whether small genome regions degenerate, or only large ones, whether selective constraints on the genes in a sex-linked region also strongly affect degeneration rates, and about how long it takes before all (or almost all) genes are lost. This article is part of the theme issue 'Challenging the paradigm in sex chromosome evolution: empirical and theoretical insights with a focus on vertebrates (Part I)'. [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]

Immune reactions in the tumor microenvironment are an important hallmark of cancer, and emerging immune therapies have been proven effective against several types of cancers. To investigate cancer genome-immune interactions and the role of immunoediting or immune escape mechanisms in cancer development, we analyzed 2834 whole genome and RNA sequencing datasets across 31 distinct tumor types with respect to key immunogenomic aspects and provided comprehensive immunogenomic profiles of pan-cancers. We found that selective copy number changes in immune-related genes may contribute to immune escape. Furthermore, we developed an index of the immunoediting history of each tumor sample based on the information of mutations in exonic regions and pseudogenes and evaluated the immunoediting history of each tumor. Our immuno-genomic analyses of pan-cancers have the potential to identify a subset of tumors with immunogenicity and diverse backgrounds or intrinsic pathways associated with their immune status and immunoediting history. [ABSTRACT FROM AUTHOR]

OCT4 plays critical roles in self-renewal and pluripotency maintenance of embryonic stem cells, and is considered as one of the main stemness markers. It also has pivotal roles in early stages of embryonic development. Most studies on OCT4 have focused on the expression and function of OCT4A, which is the biggest isoform of OCT4 known so far. Recently, many studies have shown that OCT4 has various transcript variants, protein isoforms, as well as pseudogenes. Distinguishing the expression and function of these variants and isoforms is a big challenge in expression profiling studies of OCT4. Understanding how OCT4 is functioning in different contexts, depends on knowing of where and when each of OCT4 transcripts, isoforms and pseudogenes are expressed. Here, we review OCT4 known transcripts, isoforms and pseudogenes, as well as its interactions with other proteins, and emphasize the importance of discriminating each of them in order to understand the exact function of OCT4 in stem cells, normal development and development of diseases. [ABSTRACT FROM AUTHOR]

Mealybugs are insects that maintain intracellular bacterial symbionts to supplement their nutrient-poor plant sap diets. Some mealybugs have a single betaproteobacterial endosymbiont, a Candidatus Tremblaya species (hereafter Tremblaya) that alone provides the insect with its required nutrients. Other mealybugs have two nutritional endosymbionts that together provision these same nutrients, where Tremblaya has gained a gammaproteobacterial partner that resides in its cytoplasm. Previous work had established that Pseudococcus longispinus mealybugs maintain not one but two species of gammaproteobacterial endosymbionts along with Tremblaya. Preliminary genomic analyses suggested that these two gammaproteobacterial endosymbionts have large genomes with features consistent with a relatively recent origin as insect endosymbionts, but the patterns of genomic complementarity between members of the symbiosis and their relative cellular locations were unknown. Here, using long-read sequencing and various types of microscopy, we show that the two gammaproteobacterial symbionts of P. longispinus are mixed together within Tremblaya cells, and that their genomes are somewhat reduced in size compared with their closest nonendosymbiotic relatives. Both gammaproteobacterial genomes contain thousands of pseudogenes, consistent with a relatively recent shift from a free-living to an endosymbiotic lifestyle. Biosynthetic pathways of key metabolites are partitioned in complex interdependent patterns among the two gammaproteobacterial genomes, the Tremblaya genome, and horizontally acquired bacterial genes that are encoded on the mealybug nuclear genome. Although these two gammaproteobacterial endosymbionts have been acquired recently in evolutionary time, they have already evolved codependencies with each other, Tremblaya , and their insect host. [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]

Intelectins are a family of multimeric secreted proteins that bind microbe-specific glycans. Both genetic and functional studies have suggested that intelectins have an important role in innate immunity and are involved in the etiology of various human diseases, including inflammatory bowel disease. Experiments investigating the role of intelectins in human disease using mouse models are limited by the fact that there is not a clear one-to-one relationship between intelectin genes in humans and mice, and that the number of intelectin genes varies between different mouse strains. In this study we show by gene sequence and gene expression analysis that human intelectin-1 (ITLN1) has multiple orthologues in mice, including a functional homologue Itln1; however, human intelectin-2 has no such orthologue or homologue. We confirm that all sub-strains of the C57 mouse strain have a large deletion resulting in retention of only one intelectin gene, Itln1. The majority of laboratory strains have a full complement of six intelectin genes, except CAST, SPRET, SKIVE, MOLF and PANCEVO strains, which are derived from different mouse species/subspecies and encode different complements of intelectin genes. In wild mice, intelectin deletions are polymorphic in Mus musculus castaneus and Mus musculus domesticus. Further sequence analysis shows that Itln3 and Itln5 are polymorphic pseudogenes due to premature truncating mutations, and that mouse Itln1 has undergone recent adaptive evolution. Taken together, our study shows extensive diversity in intelectin genes in both laboratory and wild-mice, suggesting a pattern of birth-and-death evolution. In addition, our data provide a foundation for further experimental investigation of the role of intelectins in disease. [ABSTRACT FROM AUTHOR]

Copy Number Variations (CNVs) account for a large proportion of human genome and are a primary contributor to human phenotypic variation, in addition to being the molecular basis of a wide spectrum of disease. Multiallelic CNVs represent a considerable fraction of large CNVs and are strictly related to segmental duplications according to their prevalent duplicate alleles. RCCX CNV is a complex, multiallelic and tandem CNV located in the major histocompatibility complex (MHC) class III region. RCCX structure is typically defined by the copy number of a DNA segment containing a series of genes – the serine/threonine kinase 19 (STK19), the complement 4 (C4), the steroid 21-hydroxylase (CYP21), and the tenascin-X (TNX) – lie close to each other. In the Caucasian population, the most common RCCX haplotype (69%) consists of two segments containing the genes STK19-C4A-CYP21A1P-TNXA-STK19B-C4B-CYP21A2-TNXB , with a telomere-to-centromere orientation. Nonallelic homologous recombination (NAHR) plays a key role into the RCCX genetic diversity: unequal crossover facilitates large structural rearrangements and copy number changes, whereas gene conversion mediates relatively short sequence transfers. The results of these events increased the RCCX genetic diversity and are responsible of specific human diseases. This review provides an overview on RCCX complexity pointing out the molecular bases of Congenital Adrenal Hyperplasia (CAH) due to CYP21A2 deficiency, CAH-X Syndrome and disorders related to CNV of complement component C4. [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]

Congenital adrenal hyperplasia (CAH) due to 21-hydroxylase deficiency is a complicated condition genetically, clinically, and treatment wise. Genetically, there are numerus mutations with different effect on enzyme activity that make genetic diagnosis a challenge. Clinically, there are a wide range of presentations from asymptomatic patients to the severe life-threatening classic CAH. Both an asymptomatic heterozygote and a mildly affected non-classical patient can carry a 'severe' mutation and endow it to their offspring. We present a case of non-classic CAH and discuss the problematic relations between biochemical and genetic diagnosis. By integrating the seemingly contradicting literature, we provide a new simple tool to assess the risk of such patients to give birth to a child with classic CAH. Congenital adrenal hyperplasia (CAH) due to 21-hydroxylase deficiency varies clinically from asymptomatic or mild disease (non-classic), through 'simple virilizing' type, to a severe life-threatening condition (classic) due to adrenal insufficiency. 'Classic' CAH patient must harbor two 'severe' mutations, one on each of the alleles of the 21OH gene. Prevention of the birth of a child with classic disease is a challenge as the heterozygote carrier of 'severe' mutation is usually asymptomatic with normal biochemical tests, and a 'non-classic' patient can still carry a 'severe' mutation. Genetic diagnosis is a challenge, as there are numerus mutations including deletions, intronic mutations, double and triple mutations, and pseudogenes. In this article, we provide a new simple tool to assess the risk of a couple to give birth to a child with 'classic' disease. [ABSTRACT FROM AUTHOR]

LINE-1-mediated retrotransposition of protein-coding mRNAs is an active process in modern humans for both germline and somatic genomes. Prior works that surveyed human data mostly relied on detecting discordant mappings of paired-end short reads, or exon junctions contained in short reads. Moreover, there have been few genome-wide comparisons between gene retrocopies in great apes and humans. In this study, we introduced a more sensitive and accurate method to identify processed pseudogenes. Our method utilizes long-read assemblies, and more importantly, is able to provide full-length retrocopy sequences as well as flanking regions which are missed by short-read based methods. From 22 human individuals, we pinpointed 40 processed pseudogenes that are not present in the human reference genome GRCh38 and identified 17 pseudogenes that are in GRCh38 but absent from some input individuals. This represents a significantly higher discovery rate than previous reports (39 pseudogenes not in the reference genome out of 939 individuals). We also provided an overview of lineage-specific retrocopies in chimpanzee, gorilla, and orangutan genomes. [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]

Transposable elements (TEs) help shape the structure and function of the human genome. When inserted into some locations, TEs may disrupt gene regulation and cause diseases. Here, we present xTea (x-Transposable element analyzer), a tool for identifying TE insertions in whole-genome sequencing data. Whereas existing methods are mostly designed for short-read data, xTea can be applied to both short-read and long-read data. Our analysis shows that xTea outperforms other short read-based methods for both germline and somatic TE insertion discovery. With long-read data, we created a catalogue of polymorphic insertions with full assembly and annotation of insertional sequences for various types of retroelements, including pseudogenes and endogenous retroviruses. Notably, we find that individual genomes have an average of nine groups of full-length L1s in centromeres, suggesting that centromeres and other highly repetitive regions such as telomeres are a significant yet unexplored source of active L1s. xTea is available at https://github.com/parklab/xTea. Identification of transposable element (TE) insertions from whole genome sequencing data remains challenging. Here the authors developed a comprehensive TE insertion detection algorithm xTea that can be applied to both short-read and long-read sequencing data. [ABSTRACT FROM AUTHOR]

Hepatica is a small genus of Ranunculaceae with medicinal and horticultural value. We characterized nine complete chloroplast (cp) genomes of Hepatica , which ranged from 159,549 to 161,081 bp in length and had a typical quadripartite structure with a large single-copy region (LSC; 80,270–81,249 bp), a small single-copy region (SSC; 17,029–17,838 bp), and two copies of inverted repeat (IR; 31,008–31,100 bp). The cp genomes of Hepatica possess 76 protein-coding genes (PCGs), 29 tRNAs, and four rRNA genes. Comparative analyses revealed a conserved ca. 5-kb IR expansion in Hepatica and other Anemoneae; moreover, multiple inversion events occurred in Hepatica and its relatives. Analyses of selection pressure (dN / dS) showed that most of the PCGs are highly conserved except for rpl20 and rpl22 in Hepatica falconeri , Hepatica americana , and Hepatica acutiloba. Two genes (rps16 and infA) were identified as pseudogenes in Hepatica. In contrast, rpl32 gene was completely lost. The phylogenetic analyses based on 76 PCGs resolved the phylogeny of Hepatica and its related genera. Non-monophyly of Anemone s.l. indicates that Hepatica should be reclassified as an independent genus. In addition, Hepatica nobilis var. japonica is not closely related to H. nobilis. [ABSTRACT FROM AUTHOR]

Cervical epithelial cells play a central role in cervical remodeling (CR) during pregnancy and cervical events during menstrual cycle, including mounting physical and immunological barriers, proliferation and differentiation, maintenance of fluid balance, and likely in withstanding the mechanical force exerted by the growing fetus prior to term. In the present study, we attempt to decipher the specific roles of VEGF in fetal human cervical epithelial cells by delineating VEGF signature genes using RNA sequencing in order to characterize the specific biological effects of VEGF in these cells. Out of a total of 25,000 genes screened, 162 genes were found to be differentially expressed in human cervical epithelial cells, of which 12 genes were found to be statistically significantly differentially expressed. The differentially expressed genes (162) were categorized by biological function, which included (1) proliferation, (2) immune response, (3) structure/matrix, (4) mitochondrial function, and (5) cell adhesion/communication and others (pseudogenes, non-coding RNA, miscellaneous genes, and uncharacterized genes). We conclude that VEGF plays a key role in CR by altering the expression of genes that regulate proliferation, immune response, energy metabolism and cell structure, and biological processes that are essential to development and likely CR. [ABSTRACT FROM AUTHOR]

While the samples and data from the Pima Indians of the Gila River Indian Community have been included in many international HLA workshops and conferences and have been the focus of numerous population reports and the source of novel alleles at the classical HLA loci, they have not been studied for the non-classical loci. In order to expand our HLA -disease association studies, we typed over 300 whole genome sequences from full Pima heritage members, controlled for first degree relationship, and employed recently developed computer algorithms to resolve HLA alleles. Both classical— HLA-A, -B , and - C — and non-classical— HLA-E, -F, -G, -J, -L, -W, -Y, -DPA2, -DPB2, -DMA, -DMB, -DOA, -DRB2, -DRB9 , TAP1 — loci were typed at the 4-field level of resolution. We present allele and selected haplotype frequencies, test the genotype distributions for population structure, discuss the issues that are created for tests of Hardy-Weinberg equilibrium over the four sample spaces of high resolution HLA typing, and address the implications for the evolution of non-classical pseudogenes that are no longer expressed in a phenotype subject to natural selection. [ABSTRACT FROM AUTHOR]

Rod monochromacy is a rare condition in vertebrates characterized by the absence of cone photoreceptor cells. The resulting phenotype is colourblindness and low acuity vision in dim-light and blindness in bright-light conditions. Early reports of xenarthrans (armadillos, sloths and anteaters) suggest that they are rod monochromats, but this has not been tested with genomic data. We searched the genomes of Dasypus novemcinctus (nine-banded armadillo), Choloepus hoffmanni (Hoffmann's two-toed sloth) and Mylodon darwinii (extinct ground sloth) for retinal photoreceptor genes and examined them for inactivating mutations. We performed PCR and Sanger sequencing on cone phototransduction genes of 10 additional xenarthrans to test for shared inactivating mutations and estimated the timing of inactivation for photoreceptor pseudogenes. We concluded that a stem xenarthran became an long-wavelength sensitive-cone monochromat following a missense mutation at a critical residue in SWS1, and a stem cingulate (armadillos, glyptodonts and pampatheres) and stem pilosan (sloths and anteaters) independently acquired rod monochromacy early in their evolutionary history following the inactivation of LWS and PDE6C, respectively. We hypothesize that rod monochromacy in armadillos and pilosans evolved as an adaptation to a subterranean habitat in the early history of Xenarthra. The presence of rod monochromacy has major implications for understanding xenarthran behavioural ecology and evolution.

Here we present the results of a large-scale bioinformatics annotation of non-coding RNA loci in 48 avian genomes. Our approach uses probabilistic models of hand-curated families from the Rfam database to infer conserved RNA families within each avian genome. We supplement these annotations with predictions from the tRNA annotation tool, tRNAscan-SE and microRNAs from miRBase. We identify 34 lncRNA-associated loci that are conserved between birds and mammals and validate 12 of these in chicken. We report several intriguing cases where a reported mammalian lncRNA, but not its function, is conserved. We also demonstrate extensive conservation of classical ncRNAs (e.g., tRNAs) and more recently discovered ncRNAs (e.g., snoRNAs and miRNAs) in birds. Furthermore, we describe numerous "losses" of several RNA families, and attribute these to either genuine loss, divergence or missing data. In particular, we show that many of these losses are due to the challenges associated with assembling avian microchromosomes. These combined results illustrate the utility of applying homology-based methods for annotating novel vertebrate genomes.

Interferon lambda 4 gene (IFNL4) encodes IFN-λ4, a new member of the IFN-λ family with antiviral activity. In humans IFNL4 open reading frame is truncated by a polymorphic frame-shift insertion that eliminates IFN-λ4 and turns IFNL4 into a polymorphic pseudogene. Functional IFN-λ4 has antiviral activity but the elimination of IFN-λ4 through pseudogenization is strongly associated with improved clearance of hepatitis C virus (HCV) infection. We show that functional IFN-λ4 is conserved and evolutionarily constrained in mammals and thus functionally relevant. However, the pseudogene has reached moderately high frequency in Africa, America, and Europe, and near fixation in East Asia. In fact, the pseudogenizing variant is among the 0.8% most differentiated SNPs between Africa and East Asia genome-wide. Its raise in frequency is associated with additional evidence of positive selection, which is strongest in East Asia, where this variant falls in the 0.5% tail of SNPs with strongest signatures of recent positive selection genome-wide. Using a new Approximate Bayesian Computation (ABC) approach we infer that the pseudogenizing allele appeared just before the out-of-Africa migration and was immediately targeted by moderate positive selection; selection subsequently strengthened in European and Asian populations resulting in the high frequency observed today. This provides evidence for a changing adaptive process that, by favoring IFN-λ4 inactivation, has shaped present-day phenotypic diversity and susceptibility to disease.

BRAF inhibitors (BRAFi) selectively target oncogenic BRAFV600E/K and are effective in 80% of advanced cutaneous malignant melanoma cases carrying the V600 mutation. However, the development of drug resistance limits their clinical efficacy. Better characterization of the underlying molecular processes is needed to further improve treatments. We previously demonstrated that transcription of PTEN is negatively regulated by the PTEN pseudogene antisense RNA, PTENP1-AS, and here we investigated the impact of this transcript on clinical outcome and BRAFi resistance in melanoma. We observed that increased expression levels of PTENP1-AS in BRAFi resistant cells associated with enrichment of EZH2 and H3K27me3 at the PTEN promoter, consequently reducing the expression levels of PTEN. Further, we showed that targeting of the PTENP1-AS transcript sensitized resistant cells to BRAFi treatment and that high expression of PTENP1-AS in stage III melanoma correlated with poor survival. Collectively, the data presented here show that PTENP1-AS is a promising target for re-sensitizing cells to BRAFi and also a possible prognostic marker for clinical outcome in stage III melanoma. [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]

B Cover Legend b Water lily ( I Nymphaea colorata i ), an early diverging angiosperm, whose genome contains pseudogenes as "footprints" of the seed maturation regulators. Image courtesy of Mariko Nonogaki (Nishiyama I et al i ., pp. 889-901). [Extracted from the article]

Recently, biallelic variants in the SORD gene were identified as causal for axonal hereditary neuropathy (HN). We ascertained the spectrum and frequency of SORD variants among a large cohort of Czech patients with unknown cause of HN. Exome sequencing data were analysed for SORD (58 patients). The prevalent c.757del variant was tested with fragment analysis (931 patients). Sanger sequencing in additional 70 patients was done. PCR primers were designed to amplify the SORD gene with the exclusion of the pseudogene SORD2P. Sequence differences between gene and pseudogene were identified and frequencies of SNPs were calculated. Eighteen patients from 16 unrelated families with biallelic variants in the SORD gene were found and the c.757del was present in all patients on at least one allele. Three novel, probably pathogenic, variants were detected, always in a heterozygous state in combination with the c.757del on the second allele. Patients presented with a slowly progressive axonal HN. Almost all patients had moderate pes cavus deformity. SORD neuropathy is frequent in Czech patients and the third most common cause of autosomal recessive HN. The c.757del is highly prevalent. Specific amplification of the SORD gene with the exclusion of the pseudogene is essential for a precise molecular diagnostics. [ABSTRACT FROM AUTHOR]

The Bemisia cassava whitefly complex includes species that cause severe crop damage through vectoring cassava viruses in eastern Africa. Currently, this whitefly complex is divided into species and subgroups (SG) based on very limited molecular markers that do not allow clear definition of species and population structure. Based on 14,358 genome-wide SNPs from 62 Bemisia cassava whitefly individuals belonging to sub-Saharan African species (SSA1, SSA2 and SSA4), and using a well-curated mtCOI gene database, we show clear incongruities in previous taxonomic approaches underpinned by effects from pseudogenes. We show that the SSA4 species is nested within SSA2, and that populations of the SSA1 species comprise well-defined south-eastern (Madagascar, Tanzania) and north-western (Nigeria, Democratic Republic of Congo, Burundi) putative sub-species. Signatures of allopatric incipient speciation, and the presence of a 'hybrid zone' separating the two putative sub-species were also detected. These findings provide insights into the evolution and molecular ecology of a highly cryptic hemipteran insect complex in African, and allow the systematic use of genomic data to be incorporated in the development of management strategies for this cassava pest. [ABSTRACT FROM AUTHOR]

Background: Commelinaceae (Commelinales) comprise 41 genera and are widely distributed in both the Old and New Worlds, except in Europe. The relationships among genera in this family have been suggested in several morphological and molecular studies. However, it is difficult to explain their relationships due to high morphological variations and low support values. Currently, many researchers have been using complete chloroplast genome data for inferring the evolution of land plants. In this study, we completed 15 new plastid genome sequences of subfamily Commelinoideae using the Mi-seq platform. We utilized genome data to reveal the structural variations and reconstruct the problematic positions of genera for the first time. Results: All examined species of Commelinoideae have three pseudogenes (accD, rpoA, and ycf15), and the former two might be a synapomorphy within Commelinales. Only four species in tribe Commelineae presented IR expansion, which affected duplication of the rpl22 gene. We identified inversions that range from approximately 3 to 15 kb in four taxa (Amischotolype, Belosynapsis, Murdannia, and Streptolirion). The phylogenetic analysis using 77 chloroplast protein-coding genes with maximum parsimony, maximum likelihood, and Bayesian inference suggests that Palisota is most closely related to tribe Commelineae, supported by high support values. This result differs significantly from the current classification of Commelinaceae. Also, we resolved the unclear position of Streptoliriinae and the monophyly of Dichorisandrinae. Among the ten CDS (ndhH, rpoC2, ndhA, rps3, ndhG, ndhD, ccsA, ndhF, matK, and ycf1), which have high nucleotide diversity values (Pi > 0.045) and over 500 bp length, four CDS (ndhH, rpoC2, matK, and ycf1) show that they are congruent with the topology derived from 77 chloroplast protein-coding genes. Conclusions: In this study, we provide detailed information on the 15 complete plastid genomes of Commelinoideae taxa. We identified characteristic pseudogenes and nucleotide diversity, which can be used to infer the family evolutionary history. Also, further research is needed to revise the position of Palisota in the current classification of Commelinaceae. [ABSTRACT FROM AUTHOR]

The bitter taste sensation is important to warn mammals of the ingestion of potentially toxic food compounds. For mammals, whose nutrition relies on highly specific food sources, such as blood in the case of vampire bats, it is unknown if bitter sensing is involved in prey selection. By contrast to other bat species, vampire bats exhibit numerous bitter taste receptor pseudogenes, which could point to a decreased importance of bitter taste. However, electrophysiological and behavioural studies suggest the existence of functional bitter taste transmission. To determine the agonist spectra of the three bitter taste receptors that are conserved in all three vampire bat species, we investigated the in vitro activation of Desmodus rotundus T2R1, T2R4 and T2R7. Using a set of 57 natural and synthetic bitter compounds, we were able to identify agonists for all three receptors. Hence, we confirmed a persisting functionality and, consequently, a putative biological role of bitter taste receptors in vampire bats. Furthermore, the activation of the human TAS2R7 by metal ions is shown to be conserved in D. rotundus. [ABSTRACT FROM AUTHOR]

Lung cancer is one of the deadliest, most aggressive cancers. Abrupt changes in gene expression represent an important challenge to understand and fight the disease. Gene co-expression networks (GCNs) have been widely used to study the genomic regulatory landscape of human cancer. Here, based on 1,143 RNA-Seq experiments from the TCGA collaboration, we constructed GCN for the most common types of lung tumors: adenocarcinoma (TAD) and squamous cells (TSCs) as well as their respective control networks (NAD and NSC). We compared the number of intra-chromosome (cis-) and inter-chromosome (trans-) co-expression interactions in normal and cancer GCNs. We compared the number of shared interactions between TAD and TSC, as well as in NAD and NSC, to observe which phenotypes were more alike. By means of an over-representation analysis, we associated network topology features with biological functions. We found that TAD and TSC present mostly cis- small disconnected components, whereas in control GCNs, both types have a giant trans- component. In both cancer networks, we observed cis- components in which genes not only belong to the same chromosome but to the same cytoband or to neighboring cytobands. This supports the hypothesis that in lung cancer, gene co-expression is constrained to small neighboring regions. Despite this loss of distant co-expression observed in TAD and TSC, there are some remaining trans- clusters. These clusters seem to play relevant roles in the carcinogenic processes. For instance, some clusters in TAD and TSC are associated with the immune system, response to virus, or control of gene expression. Additionally, other non-enriched trans- clusters are composed of one gene and several associated pseudo-genes, as in the case of the FTH1 gene. The appearance of those common trans- clusters reflects that the gene co-expression program in lung cancer conserves some aspects for cell maintenance. Unexpectedly, 0.48% of the edges are shared between control networks; conversely, 35% is shared between lung cancer GCNs, a 73-fold larger intersection. This suggests that in lung cancer a process of de-differentiation may be occurring. To further investigate the implications of the loss of distant co-expression, it will become necessary to broaden the investigation with other omic-based approaches. However, the present approach provides a basis for future work toward an integrative perspective of abnormal transcriptional regulatory programs in lung cancer. [ABSTRACT FROM AUTHOR]

We identified and characterized the pseudogene complements of five plant species: four dicots (Arabidopsis thaliana, Vitis vinifera, Populus trichocarpa and Phaseolus vulgaris) and one monocot (Oryza sativa). Retroposition was considered of modest importance for pseudogene formation in all investigated species except V. vinifera, which showed an unusually high number of retro-pseudogenes in non coding genic regions. By using a pipeline for the classification of sequence duplicates in plant genomes, we compared the relative importance of whole genome, tandem, proximal, transposed and dispersed duplication modes in the pseudo and functional gene complements. Pseudogenes showed higher tendencies than functional genes to genomic dispersion. Dispersed pseudogenes were prevalently fragmented and showed high sequence divergence at flanking regions. On the contrary, those deriving from whole genome duplication were proportionally less than expected based on observations on functional loci and showed higher levels of flanking sequence conservation than dispersed pseudogenes. Pseudogenes deriving from tandem and proximal duplications were in excess compared to functional loci, probably reflecting the high evolutionary rate associated with these duplication modes in plant genomes. These data are compatible with high rates of sequence turnover at neutral sites and double strand break repairs mediated duplication mechanisms. [ABSTRACT FROM AUTHOR]

Next-generation sequencing (NGS) is an incredibly useful tool for genetic disease diagnosis. However, the most commonly used bioinformatics methods for analyzing sequence reads insufficiently discriminate genomic regions with extensive sequence identity, such as gene families and pseudogenes, complicating diagnostics. This problem has been recognized for specific genes, including many involved in human disease, and diagnostic labs must perform additional costly steps to guarantee accurate diagnosis in these cases. Here we report a new data analysis method based on the comparison of read depth between highly homologous regions to identify misalignment. Analyzing six clinically important genes—CYP21A2, GBA, HBA1/2, PMS2, and SMN1—each exhibiting misalignment issues related to homology, we show that our technique can correctly identify potential misalignment events and be used to make appropriate calls. Combined with long-range PCR and/or MLPA orthogonal testing, our clinical laboratory can improve variant calling with minimal additional cost. We propose an accurate and cost-efficient NGS testing procedure that will benefit disease diagnostics, carrier screening, and research-based population studies. [ABSTRACT FROM AUTHOR]

Pan-genome analysis is widely used to study the evolution and genetic diversity of species, particularly in bacteria. However, the impact of strain selection on the outcome of pan-genome analysis is poorly understood. Furthermore, a standard protocol to ensure high-quality pan-genome results is lacking. In this study, we carried out a series of pan-genome analyses of different strain sets of Bacillus subtilis to understand the impact of various strains on the performance and output quality of pan-genome analyses. Consequently, we found that the results obtained by pan-genome analyses of B. subtilis can be influenced by the inclusion of incorrectly classified Bacillus subspecies strains, phylogenetically distinct strains, engineered genome-reduced strains, chimeric strains, strains with a large number of unique genes or a large proportion of pseudogenes, and multiple clonal strains. Since the presence of these confounding strains can seriously affect the quality and true landscape of the pan-genome, we should remove these deviations in the process of pan-genome analyses. Our study provides new insights into the removal of biases from confounding strains in pan-genome analyses at the beginning of data processing, which enables the achievement of a closer representation of a high-quality pan-genome landscape of B. subtilis that better reflects the performance and credibility of the B. subtilis pan-genome. This procedure could be added as an important quality control step in pan-genome analyses for improving the efficiency of analyses, and ultimately contributing to a better understanding of genome function, evolution and genome-reduction strategies for B. subtilis in the future. [ABSTRACT FROM AUTHOR]

This study reports the complete chloroplast genome sequence of a continental island endemic, Potentilla gageodoensis. The total plastome size was 156,397 bp, comprising one large single-copy (LSC; 85,768 bp), one small single-copy (SSC; 18,589 bp), and two inverted repeat (IR) regions (IRa and IRb, each with 26,020 bp). The overall GC content was 36.92%, and the plastome contained 131 genes, comprising 84 protein-coding genes with two pseudogenes (infA and ycf1), 37 transfer RNA genes, and eight ribosomal RNA genes. Phylogenetic analysis performed using 27 representative Rosoideae plastomes suggests that the genus Potentilla is not monophyletic and that P. gageodoensis is sister to the clade containing four taxa of Potentilla (P. freyniana, P. freyniana var. chejuensis, P. stolonifera, and P. stolonifera var. quelpaertensis). The present study reveals the taxonomic distinction of P. gageodoensis from its congeneric species in Korea and the plastome sequence obtained from this study can be used to study phylogenetic relationships and taxonomic status. [ABSTRACT FROM AUTHOR]

Summary We have implemented the pypgatk package and the pgdb workflow to create proteogenomics databases based on ENSEMBL resources. The tools allow the generation of protein sequences from novel protein-coding transcripts by performing a three-frame translation of pseudogenes, lncRNAs and other non-canonical transcripts, such as those produced by alternative splicing events. It also includes exonic out-of-frame translation from otherwise canonical protein-coding mRNAs. Moreover, the tool enables the generation of variant protein sequences from multiple sources of genomic variants including COSMIC, cBioportal, gnomAD and mutations detected from sequencing of patient samples. pypgatk and pgdb provide multiple functionalities for database handling including optimized target/decoy generation by the algorithm DecoyPyrat. Finally, we have reanalyzed six public datasets in PRIDE by generating cell-type specific databases for 65 cell lines using the pypgatk and pgdb workflow, revealing a wealth of non-canonical or cryptic peptides amounting to >5% of the total number of peptides identified. Availability and implementation The software is freely available. pypgatk : https://github.com/bigbio/py-pgatk/ and pgdb : https://nf-co.re/pgdb. Supplementary information Supplementary data are available at Bioinformatics online. [ABSTRACT FROM AUTHOR]

Mapping reads to a reference sequence is a common step when analyzing allele effects in high-throughput sequencing data. The choice of reference is critical because its effect on quantitative sequence analysis is non-negligible. Recent studies suggest aligning to a single standard reference sequence, as is common practice, can lead to an underlying bias depending on the genetic distances of the target sequences from the reference. To avoid this bias, researchers have resorted to using modified reference sequences. Even with this improvement, various limitations and problems remain unsolved, which include reduced mapping ratios, shifts in read mappings and the selection of which variants to include to remove biases. To address these issues, we propose a novel and generic multi-alignment pipeline. Our pipeline integrates the genomic variations from known or suspected founders into separate reference sequences and performs alignments to each one. By mapping reads to multiple reference sequences and merging them afterward, we are able to rescue more reads and diminish the bias caused by using a single common reference. Moreover, the genomic origin of each read is determined and annotated during the merging process, providing a better source of information to assess differential expression than simple allele queries at known variant positions. Using RNA-seq of a diallel cross, we compare our pipeline with the single-reference pipeline and demonstrate our advantages of more aligned reads and a higher percentage of reads with assigned origins. Database URL: http://csbio.unc.edu/CCstatus/index.py?run=Pseudo.

Spectral entropy and GC content analyses reveal comprehensive structural features of DNA sequences. To illustrate the significance of these features, we analyze the β-esterase gene cluster, including the Est-6 gene and the ψEst-6 putative pseudogene, in seven species of the Drosophila melanogaster subgroup. The spectral entropies show distinctly lower structural ordering for ψEst-6 than for Est-6 in all species studied. However, entropy accumulation is not a completely random process for either gene and it shows to be nucleotide dependent. Furthermore, GC content in synonymous positions is uniformly higher in Est-6 than in ψEst-6, in agreement with the reduced GC content generally observed in pseudogenes and nonfunctional sequences. The observed differences in entropy and GC content reflect an evolutionary shift associated with the process of pseudogenization and subsequent functional divergence of ψEst-6 and Est-6 after the duplication event. The data obtained show the relevance and significance of entropy and GC content analyses for pseudogene identification and for the comparative study of gene-pseudogene evolution.

Abstract Background Increasing evidence suggests a critical role for long noncoding RNAs (LncRNAs) and pseudogenes in cancer. Renal cell carcinoma (RCC), the most common primary renal neoplasm, is highly aggressive and difficult to treat because of its resistance to chemotherapy and radiotherapy. Despite many identified LncRNAs and pseudogenes, few have been clearly elucidated. Methods This study provides new insights into LncRNAs and pseudogenes in the prognosis of RCC. We searched an online database to interrogate alterations and clinical data on cBioPortal. We analysed LncRNA and pseudogene signatures to predict the prognosis of RCC based on a Cox model. We also found potential serum biomarkers of RCC and validated them in 32 RCC patients, as well as healthy controls. Results Alterations were found in 2553 LncRNAs and 8901 pseudogenes and occurred in up to 23% of all cases. Among these, 27 LncRNAs and 45 pseudogenes were closely related to prognosis. We also identified signatures of LncRNAs and pseudogenes that can predict overall survival and recurrence of RCC. We then validated the relative levels of these LncRNAs and pseudogenes in the serum of 32 patients. Six of these, including LINC00520, PIK3CD-AS1, LINC01559, CEACAM22P, MSL3P1 and TREML3P, could be non-invasive biomarkers of RCC. Finally, we selected PIK3CD-AS1 to determine its role in RCC and found that upregulation of PIK3CD-AS1 was closely associated with higher tumour stage and metastasis. Conclusions These signatures of LncRNAs and pseudogenes can predict overall survival and recurrence of RCC. LINC00520, PIK3CD-AS1, LINC01559, CEACAM22P, MSL3P1 and TREML3P could be non-invasive biomarkers of RCC. These data suggest the important roles of LncRNAs and pseudogenes in RCC, and therefore provides us new insights into the prognosis of RCC.

Cardiovascular disease is the main disease that affects human life span. In recent years, the disease has been increasingly addressed at the molecular levels, for example, pseudogenes are now known to be involved in the pathogenesis and development of cardiovascular diseases. Pseudogenes are non-coding homologs of protein-coding genes and were once called “junk gene.” Since they are highly homologous to their functional parental genes, it is somewhat difficult to distinguish them. With the development of sequencing technology and bioinformatics, pseudogenes have become readily identifiable. Recent studies indicate that pseudogenes are closely related to cardiovascular diseases. This review provides an overview of pseudogenes and their roles in the pathogenesis of cardiovascular diseases. This new knowledge adds to our understanding of cardiovascular disease at the molecular level and will help develop new biomarkers and therapeutic approaches designed to prevent and treat the disease.

Strains belonging to the Pseudomonas syringae complex often possess quorum-sensing systems that comprise N -acyl- l -homoserine lactone (AHL) synthases (PsyI) and AHL receptors (PsyR). Here, we investigated the diversity of PsyI/PsyR quorum-sensing systems in 630 strains of the P. syringae complex. AHL production was observed in most strains of Pseudomonas amygdali and Pseudomonas meliae , and a few strains of Pseudomonas coronafaciens and P. syringae. The DNA sequences of psyIR and their upstream and downstream regions were categorized into eight types. P. amygdali pv. myricae , Pseudomonas savastanoi , and P. syringae pv. solidagae , maculicola , broussonetiae , and tomato encoded psyI , but did not produce detectable amounts of AHL. In P. savastanoi , an amino acid substitution (R27S) in PsyI caused defective AHL production. The psyI gene of P. syringae pv. tomato was converted to pseudogenes by frameshift mutations. Escherichia coli harboring psyI genes from P. amygdali pv. myricae , P. syringae pv. solidagae and broussonetiae showed high levels of AHL production. Forced expression of functional psyR restored AHL production in P. amygdali pv. myricae and P. syringae pv. solidagae. In conclusion, our study indicates that the PsyI/PsyR quorum-sensing systems in P. syringae strains are genetically and functionally diverse, with diversity being linked to phylogenetic and pathovar classifications. [ABSTRACT FROM AUTHOR]