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Search indexed bioRxiv preprints in genomics, neuroscience, cell biology and bioinformatics. Read source abstracts and check manuscript versions; preprints are not peer reviewed.

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bioRxiv2018

The core genome m5C methyltransferase JHP1050 (M.Hpy99III) plays an important role in orchestrating gene expression in Helicobacter pylori

Helicobacter pylori encodes a large number of Restriction-Modification (R-M) systems despite its small genome.R-M systems have been described as \"primitive immune systems\" in bacteria, but the role of methylation in bacterial gene regulation and other processes is increasingly accepted. Every H.pylori strain harbours a unique set of R-M systems resulting in a highly diverse methylome. We identified a highly conserved GCGC-specific m5C MTase (JHP1050) that was predicted to be active in all of 459 H.pylori genome sequences analyzed. Transcriptome analysis of two H.pylori strains and their respective MTase mutants showed that inactivation of the MTase led to changes in the expression of 225 genes in strain J99, and 29 genes in strain BCM-300.10 genes were differentially expressed in both mutated strains. Combining bioinformatic analysis and site-directed mutagenesis, we demonstrated that motifs overlapping the promoter influence the expression of genes directly, while methylation of other motifs might cause secondary effects.Thus, m5C methylation modifies the transcription of multiple genes, affecting important phenotypic traits that include adherence to host cells, natural competence for DNA uptake, bacterial cell shape, and susceptibility to copper.

microbiology↗

bioRxiv2018

Association of Prevotella enterotype with polysomnographic data in obstructive sleep apnea/hypopnea syndrome patients

Intermittent hypoxia and sleep fragmentation are critical pathophysiological processes involved in obstructive sleep apnea/hypopnea syndrome (OSAHS). These manifestation independently affect similar brain regions and contribute to OSAHS-related comorbidities that are known to be related to the host gut alteration microbiota. We hypothesized that microbiota disruption influences the pathophysiological processes of OSAHS through a microbiota-gut-brain axis. Thus, we aim to survey enterotypes and polysomnographic data of OSAHS patients. Subjects were diagnosed by polysomnography, from whom fecal samples were obtained and analyzed for the microbiome composition by variable regions 3-4 of 16S rRNA pyrosequencing and bioinformatic analyses. We examined blood cytokines level of all subjects. Three enterotypes Bacteroides (n=73), Ruminococcus (n=14), and Prevotella (n=26) were identified. Central apnea indices, mixed apnea indices, N1 sleep stage, mean apnea-hypopnea duration, and arousal indices were increased in apnea-hypopnea indices (AHI) [≥]15 patients with the Prevotella enterotype. However, for AHI<15 subjects, obstructive apnea indices and systolic blood pressure were significantly observed in Ruminococcus and Prevotella enterotypes, respectively. The present study indicates the possibility of pathophysiological interplay between enterotypes and sleep structure disruption in sleep apnea through a microbiota-gut-brain axis and offers some new insight toward the pathogenesis of OSAHS.\n\nImportanceIntermittent hypoxia (IH) and sleep fragmentation (SF) are hallmarks of are the predominant mechanism underlying obstructive sleep apnea/hypopnea syndrome (OSAHS). Moreover, IH and SF of pathophysiological roles in the gut microbiota dysbiosis in OSAHS have been demonstrated. We hypothesized that gut microbiota disruption may cross-talk the brain function via microbiota-gut-brain axis. Indeed, we observed central apnea indices and other parameters of disturbances during sleep were significantly elevated in AHI[≥]15 patients with the Prevotella enterotype. This enterotype prone to endotoxin production, driving systemic inflammation, ultimately contributes to OSAHS-linked comorbidities. Vice versa, increasing the arousal index leads to systemic inflammatory changes and accompanies metabolic dysfunction. We highlight that the possibility that the microbiota-gut-brain axis operates a bidirectional effect on the development of OSAHS pathology.

microbiology↗

bioRxiv2018

Circular RNAs regulate cancer stem cells by FMRP against CCAR1 complex in hepatocellular carcinoma

Circular RNA (circRNA) possesses great pre-clinical diagnostic and therapeutic potentials in multiple cancers. However, the underlying correlation between circRNAs and cancer stem cells (CSCs) has not been reported. The absence of circZKSCAN1 endowed several malignant properties including cancer stemness and tightly correlated with worse overall and recurrence-free survival rate in HCC cells in vitro and in vivo. Bioinformatics analysis and RNA immunoprecipitation-sequencing (RIP-seq) results revealed that circZKSCAN1 exerted its inhibitive role by competitively binding FMRP, therefore, block the binding between FMRP and {beta}-catenin-binding protein-cell cycle and apoptosis regulator 1 (CCAR1) mRNA, and subsequently restraining the transcriptional activity of Wnt signaling. In addition, RNA-splicing protein Quaking 5 was found downregulated in HCC tissues and responsible for the reduction of circZKSCAN1. Collectively, this study revealed the mechanisms underlying the regulatory role of circZKSCAN1 in HCC CSCs and identified the newly discovered Qki5- circZKSCAN1-FMRP-CCAR1-Wnt signaling axis as a potentially important therapeutic target for HCC treatment.\n\nStatement of significanceO_LICircZKSCAN1, a novel negative regulator for cancer stem cells, was firstly identified with reverse correlation with HCC prognosis.\nC_LIO_LICircZKSCAN1 directly targets FMRP, and competitive binding with {beta}-catenin-binding protein cell cycle and apoptosis regulator 1 (CCAR1) for its activity.\nC_LIO_LIThe decreased expression of Quaking 5 is responsible for the absence of circZKSCAN1 in HCC.\nC_LI

cancer biology↗

bioRxiv2018

Topokaryotyping demonstrates single cell variability and stress dependent variations in nuclear envelope associated domains

Analysis of large-scale interphase genome positioning with reference to a nuclear landmark has recently been studied using sequencing-based single cell approaches. However, these approaches are dependent upon technically challenging, time consuming and costly high throughput sequencing technologies, requiring specialized bioinformatics tools and expertise. Here, we propose a novel, affordable and robust microscopy-based single cell approach, termed Topokaryotyping, to analyze and reconstruct the interphase positioning of genomic loci relative to a given nuclear landmark, detectable as banding pattern on mitotic chromosomes. This is accomplished by proximity-dependent histone labeling, where biotin ligase BirA fused to nuclear envelope marker Emerin was coexpressed together with Biotin Acceptor Peptide (BAP)-histone fusion followed by (i) biotin labeling, (ii) generation of mitotic spreads, (iii) detection of the biotin label on mitotic chromosomes and (iv) their identification by karyotyping. Using Topokaryotyping, we identified both cooperativity and stochasticity in the positioning of emerin-associated chromatin domains in individual cells. Furthermore, the chromosome-banding pattern showed dynamic changes in emerin-associated domains upon physical and radiological stress. In summary, Topokaryotyping is a sensitive and reliable technique to quantitatively analyze spatial positioning of genomic regions interacting with a given nuclear landmark at the single cell level in various experimental conditions.

cell biology↗

bioRxiv2018

Pervasive contaminations in sequencing experiments are a major source of false genetic variability: a Mycobacterium tuberculosis meta-analysis

Contaminant DNA is a well-known confounding factor in molecular biology and in genomic repositories. Strikingly, analysis workflows for whole-genome sequencing (WGS) data usually neglect the errors introduced by potential contaminations. We performed a comprehensive evaluation of the extent and impact of contaminant DNA in WGS by analyzing more than 4,000 bacterial samples from 20 different studies. We found that contaminations are pervasive and can introduce large biases in variant analysis. We showed that these biases can translate in hundreds of false positive and negative SNPs, even for samples with slight contaminations. Studies investigating complex biological traits from sequencing data can be completely biased if contaminations are neglected during the bioinformatic analysis. We used both real and simulated data to evaluate and implement reliable, contamination-aware analysis pipelines. Our results urge for the implementation of such pipelines as sequencing technologies consolidate as a precision tool in the research and clinical context.

genomics↗

bioRxiv2018

Chiral DNA sequences as commutable reference standards for clinical genomics

Chirality is a geometric property describing any object that is inequivalent to a mirror image of itself. Due to its 5-3 directionality, a DNA sequence is distinct from a mirrored sequence arranged in reverse nucleotide order, and is therefore chiral. A given sequence and its opposing chiral partner sequence share many properties, such as nucleotide composition and sequence entropy. Here we demonstrate that chiral DNA sequence pairs also perform equivalently during molecular and bioinformatic techniques that underpin modern genetic analysis, including PCR amplification, hybridization, whole-genome, target-enriched and nanopore sequencing, sequence alignment and variant detection. Given these shared properties, synthetic DNA sequences that directly mirror clinically relevant and/or analytically challenging regions of the human genome are ideal reference standards for clinical genomics. We show how the addition of chiral DNA standards to patient tumor samples can prevent false-positive and false-negative mutation detection and, thereby, improve diagnosis. Accordingly, we propose that chiral DNA standards can fulfill the unmet need for commutable internal reference standards in precision medicine.

genomics↗

bioRxiv2018

Eps8 is a convergence point integrating EGFR and integrin trafficking and crosstalk

Crosstalk between adhesion and growth factor receptors plays a critical role in tissue morphogenesis and repair, and aberrations contribute substantially to neoplastic disease. However, the mechanisms by which adhesion and growth factor receptor signalling are integrated, spatially and temporally, are unclear.\n\nWe used adhesion complex enrichment coupled with quantitative proteomic analysis to identify rapid changes to adhesion complex composition and signalling following growth factor stimulation. Bioinformatic network and ontological analyses revealed a substantial decrease in the abundance of adhesion regulatory proteins and co-ordinators of endocytosis within 5 minutes of EGF stimulation. Together these data suggested a mechanism of EGF-induced receptor endocytosis and adhesion complex turnover.\n\nCombinatorial interrogation of the networks allowed a global and dynamic view of adhesion and growth factor receptor crosstalk to be assembled. By interrogating network topology we identified Eps8 as a putative node integrating 5{beta}1 integrin and EGFR functions. Importantly, EGF stimulation promoted internalisation of both 5{beta}1 and EGFR. However, perturbation of Eps8 increased constitutive internalisation of 5{beta}1 and EGFR; suggesting that Eps8 constrains 5{beta}1 and EGFR endocytosis in the absence of EGF stimulation. Consistent with this, Eps8 regulated Rab5 activity and was required for maintenance of adhesion complex organisation and for EGF-dependent adhesion complex disassembly. Thus, by co-ordinating 5{beta}1 and EGFR trafficking mechanisms, Eps8 is able to control adhesion receptor and growth factor receptor bioavailability and cellular contractility.\n\nWe propose that during tissue morphogenesis and repair, Eps8 functions to spatially and temporally constrain endocytosis, and engagement, of 5{beta}1 and EGFR in order to precisely co-ordinate adhesion disassembly, cytoskeletal dynamics and cell migration.

cell biology↗

bioRxiv2018

Analysis of a structured intronic region of the LMP2 pre-mRNA from EBV reveals associations with human regulatory proteins.

Objective: The pre-mRNA of the Epstein-Barr virus LMP2 (latent membrane protein 2) has a region of unusual RNA structure that partially spans two consecutive exons and the entire intervening intron; suggesting RNA folding might affect splicing--particularly via interactions with human regulatory proteins. To better understand the roles of protein associations with this structured intronic region, we undertook a combined bioinformatics (motif searching) and experimental analysis (biotin pulldowns and RNA immunoprecipitations) of protein binding. Result: Characterization of the ribonucleoprotein composition of this region revealed several human proteins as interactors: regulatory proteins hnRNP A1 (heterogeneous nuclear ribonucleoprotein A1), hnRNP U, HuR (human antigen R), and PSF (polypyrimidine tract-binding protein-associated splicing factor), as well as, unexpectedly, the cytoskeletal protein actin. Treatment of EBV-infected cells with drugs that alter actin polymerization specifically showed marked effects on splicing in this region. This suggests a potentially novel role for nuclear actin in regulation of viral RNA splicing.

molecular biology↗

bioRxiv2018

Relative Reduction of the Biological and phylogenetic diversity of oral microbiome in diabetic and pre-diabetic subjects

BackgroundThere is a suggested reciprocal relationship between oral health and systemic disease such as type 2 diabetes. In this relationship, a systemic disease predisposing to oral infection, and when that infection is present, the oral infection aggravates the progression of the systemic disease. Several studies suggested that some oral microbiome constituents are linked to both diabetes, metabolic syndrome and obesity. This study aims to compare the microbial diversity and population structure of oral microbiome among normoglycemic, impaired glucose tolerance (IGT) and diabetic subjects.\n\nMethodologyThis study followed a case-control design (15 T2D patients, 10 IGTs and, 19 controls). Patient records were screened as per the inclusion and exclusion criteria. Assessment of periodontitis and oral health was performed to all subjects. DNA Isolation purification and quantification from collected Saliva samples were performed. 16SrRNA hypervariable regions were amplified and sequenced. Generated sequences were subjected to bioinformatics analysis. Statistical analysis and diversity indices were computed with the statistical software R, the vegan R-package, and Past318 software.\n\nResultsA total observed number of 551 OTUs. A clear reduction of the number of species (OTUs) was observed in both IGT (412) and diabetic group (372) compared with the normoglycemic group (502). This was associated with a similar pattern of biological diversity among the three groups. Phylogenetic diversity (PD-SBL) value in the normoglycemic group was higher than the diabetic group. The diabetic group had the highest evenness value and the highest microbiome bacterial pathogenic content.\n\nConclusionWe observed a clear reduction in the biological and phylogenetic diversity in the diabetic and pre-diabetic oral microbiome in comparison with the normoglycemic oral microbiome. However, this reduction was associated with an increase in the pathogenic content of the hyperglycemic microbiomes.

microbiology↗

bioRxiv2018

Transcripts from multicopy gene families localizing to mouse Y long arm encode piRNAs and proteins

Heterochromatic long arm of mouse Y chromosome harbors the multicopy species-specific sequences Ssty, Sly, Asty, and Orly that are transcribed in testis. Of these Ssty and Sly genes encode proteins - yet all the copies of these RNAs are not translated. Using bioinformatic approaches, small RNA northern blots and electrophoretic mobility shift assays, we demonstrate here that these multicopy gene families from mouse Y-long arm generate piRNAs predominantly in testis. Thus, we identified a piRNA cluster on mouse Y chromosome and also unraveled the dual role of Y-chromosome-encoded transcripts to act as primary transcripts of piRNAs in addition to their role as protein-coding RNAs.\n\nHIGHLIGHTSO_LIFirst report of a cluster of piRNAs on a mammalian Y chromosome\nC_LIO_LIReport of primary transcripts of piRNAs\nC_LIO_LIThese piRNAs putatively regulate autosomal genes expressed in mouse testis Ssty and Sly genes code for proteins as well as generate piRNAs\nC_LI

molecular biology↗

bioRxiv2018

Metagenomic profiling of ticks: identification of novel rickettsial genomes and detection of tick-borne canine parvovirus

BackgroundAcross the world, ticks act as vectors of human and animal pathogens. Ticks rely on bacterial endosymbionts, which often share close and complex evolutionary links with tick-borne pathogens. As the prevalence, diversity and virulence potential of tick-borne agents remain poorly understood, there is a pressing need for microbial surveillance of ticks as potential disease vectors.\n\nMethodology/Principal FindingsWe developed a two-stage protocol that includes 16S-amplicon screening of pooled samples of hard ticks collected from dogs, sheep and camels in Palestine, followed by shotgun metagenomics on individual ticks to detect and characterise tick-borne pathogens and endosymbionts. Two ticks isolated from sheep yielded an abundance of reads from the genus Rickettsia, which were assembled into draft genomes. One of the resulting genomes was highly similar to Rickettsia massiliae strain MTU5. Analysis of signature genes showed that the other represents the first genome sequence of the potential pathogen Candidatus Rickettsia barbariae. Ticks from a dog and a sheep yielded draft genome sequences of strains of the Coxiella-like endosymbiont Candidatus Coxeilla mudrowiae. A sheep tick yielded sequences from the sheep pathogen Anaplasma ovis, while Hyalomma ticks from camels yielded sequences belonging to Francisella-like endosymbionts. From the metagenome of a dog tick from Jericho, we generated a genome sequence of a canine parvovirus.\n\nSignificanceHere, we have shown how a cost-effective two-stage protocol can be used to detect and characterise tick-borne pathogens and endosymbionts. In recovering genome sequences from an unexpected pathogen (canine parvovirus) and a previously unsequenced pathogen (Candidatus Rickettsia barbariae), we demonstrate the open-ended nature of metagenomics. We also provide evidence that ticks can carry canine parvovirus, raising the possibility that ticks might contribute to the spread of this troublesome virus.\n\nAuthor SummaryWe have shown how DNA sequencing can be used to detect and characterise potentially pathogenic microorganisms carried by ticks. We surveyed hard ticks collected from domesticated animals across the West Bank territory of Palestine. All the ticks came from species that are also capable of feeding on humans. We detected several important pathogens, including two species of Rickettsia, the sheep pathogen Anaplasma ovis and canine parvovirus. These findings highlight the importance of hard ticks and the hazards they present for human and animal health in Palestine and the opportunities presented by high-throughput sequencing and bioinformatics analyses of DNA sequences in this setting.

microbiology↗

bioRxiv2018

A Type I Restriction-Modification System Associated with Enterococcus faecium Subspecies Separation

The gastrointestinal colonizer Enterococcus faecium is a leading cause of hospital-acquired infections. Multidrug-resistant (MDR) E. faecium are particularly concerning for infection treatment. Previous comparative genomic studies revealed that subspecies referred to as Clade A and Clade B exist within E. faecium. MDR E. faecium belong to Clade A, while Clade B consists of drug-susceptible fecal commensal E. faecium. Isolates from Clade A are further grouped into two sub-clades, A1 and A2. In general, Clade A1 isolates are hospital epidemic isolates whereas Clade A2 isolates are isolates from animals and sporadic human infections. Such phylogenetic separation indicates that reduced gene exchange occurs between the clades. We hypothesize that endogenous barriers to gene exchange exist between E. faecium clades. Restriction-modification (R-M) systems are such barriers in other microbes. We utilized bioinformatics analysis coupled with second generation and third generation deep sequencing platforms to characterize the methylome of two representative E. faecium strains, one from Clade A1 and one from Clade B. We identified a Type I R-M system that is Clade A1-specific, is active for DNA methylation, and significantly reduces transformability of Clade A1 E. faecium. Based on our results, we conclude that R-M systems act as barriers to horizontal gene exchange in E. faecium and propose that R-M systems contribute to E. faecium subspecies separation.\n\nIMPORTANCEEnterococcus faecium is a leading cause of hospital-acquired infections around the world. Rising antibiotic resistance in certain E. faecium lineages leaves fewer treatment options. The overarching aim of the attached work was to determine whether restriction-modification (R-M) systems contribute to the structure of the E. faecium species, wherein hospital-epidemic and non-hospital-epidemic isolates have distinct evolutionary histories and highly resolved clade structures. R-M provides bacteria with a type of innate immunity to horizontal gene transfer (HGT). We identified a Type I R-M system that is enriched in the hospital-epidemic clade and determined that it is active for DNA modification activity and significantly impacts HGT. Overall, this work is important because it provides a mechanism for the observed clade structure of E. faecium as well as a mechanism for facilitated gene exchange among hospital-epidemic E. faecium.

microbiology↗

bioRxiv2018

FUS controls the processing of snoRNAs into smaller RNA fragments that can regulate gene expression

FUS is a multifunctional protein involved in many steps of RNA metabolism, including transcription, splicing, miRNA processing and replication-dependent histone gene expression. In this paper, we show for the first time that FUS binds and negatively regulates the levels of a subset of snoRNAs in cells. Scanning of available human small RNA databases revealed the existence of smaller RNA fragments that can be processed from FUS-dependent snoRNAs. Therefore, we suggest that FUS mediates the biogenesis of snoRNA-derived small RNAs, called sdRNAs. Further in silico approaches enabled us to predict putative targets of selected FUS-dependent sdRNAs. Our results indicate that sdRNAs may bind to different regions of target mRNAs as well as to noncoding transcripts and influence the posttranscriptional level or translation of these targets. SIGNIFICANCE STATEMENTRNA metabolism is orchestrated by a complex network of RNA-protein interactions and involves various classes of RNA molecules. Small nucleolar RNAs (snoRNAs) are commonly considered essential components of the ribosome biogenesis pathway. However, recent studies have revealed that snoRNAs can also be fragmented into small entities called snoRNA-derived RNAs (sdRNAs), which have been linked to multiple cancer types and thus may serve as next-generation prognostic or diagnostic biomarkers. In this paper, a multifunctional protein, FUS, was shown to be involved in the biogenesis of snoRNA-derived fragments. Furthermore, we combined bioinformatic analyses with complementary experimental approaches to elucidate the role of FUS-dependent sdRNAs in gene expression regulation. Our findings reveal the considerable regulatory potential of this new class of small noncoding RNAs.

molecular biology↗

bioRxiv2018

Genetic Determinants of Cortical Structure (Thickness, Surface Area and Volumes) among Disease Free Adults in the CHARGE Consortium

Cortical thickness, surface area and volumes (MRI cortical measures) vary with age and cognitive function, and in neurological and psychiatric diseases. We examined heritability, genetic correlations and genome-wide associations of cortical measures across the whole cortex, and in 34 anatomically predefined regions. Our discovery sample comprised 22,822 individuals from 20 cohorts within the Cohorts for Heart and Aging Research in Genomic Epidemiology (CHARGE) consortium and the United Kingdom Biobank. Significant associations were replicated in the Enhancing Neuroimaging Genetics through Meta-analysis (ENIGMA) consortium, and their biological implications explored using bioinformatic annotation and pathway analyses. We identified genetic heterogeneity between cortical measures and brain regions, and 161 genome-wide significant associations pointing to wnt/{beta}-catenin, TGF-{beta} and sonic hedgehog pathways. There was enrichment for genes involved in anthropometric traits, hindbrain development, vascular and neurodegenerative disease and psychiatric conditions. These data are a rich resource for studies of the biological mechanisms behind cortical development and aging.

genetics↗

bioRxiv2018

The functional circular RNA, ciRS-7 (CDR1as), is biosynthesized using back-splicing promoted by inverted mammalian-wide MIRs but not primate-specific Alus

Circular RNAs (circRNAs) are stable noncoding RNAs with a closed circular structure. One of the first and best studied circRNAs is ciRS-7 (CDR1as) that acts as a regulator of the microRNA miR-7, however, the biosynthesis pathway has remained an enigma. Here we delineate the biosynthesis pathway of ciRS-7. The back-splicing events that form circRNAs are often facilitated by flanking inverted repeats of the primate-specific Alu elements. ciRS-7 gene lacks these elements but, instead, we identified a set of flanking inverted elements belonging to the mammalian-wide interspersed repeat (MIR) family. Splicing reporter assays in HEK293 cells demonstrated that these inverted MIRs are required to generate ciRS-7 through a back-splicing and CRISPR/Cas9-mediated deletions confirmed the requirement of the endogenous MIR elements in SH-SY5Y cells. Using bioinformatics searches, we identified several other MIR-dependent circRNAs that we confirmed experimentally. We propose that MIR-mediated RNA circularization constitutes a new widespread biosynthesis principle for mammalian circRNAs.

molecular biology↗

bioRxiv2018

Genome-wide identification and expression specificity analysis of the DNA methyltransferase gene family under adversity stresses in cotton

DNA methylation is an important epigenetic mode of genomic DNA modification that is an important part of maintaining epigenetic content and regulating gene expression. DNA methyltransferases (MTases) are the key enzymes in the process of DNA methylation. Thus far, there has been no systematic analysis the DNA MTases found in cotton. In this study, the whole genome of cotton C5-Mtase coding genes was identified and analyzed using a bioinformatics method based on information from the cotton genome. In this study, 51 DNA MTase genes were identified, of which 8 belonged to G. raimondii (group D), 9 belonged to G. arboretum L. (group A), 16 belonged to G. hirsutum L. (group AD1) and 18 belonged to G. barbadebse L. (group AD2). Systematic evolutionary analysis divided the 51 genes into four subfamilies, including 7 MET homologous proteins, 25 CMT homologous proteins, 14 DRM homologous proteins and 5 DNMT2 homologous proteins. Further studies showed that the DNA MTases in cotton were more phylogenetically conserved. The comparison of their protein domains showed that the C-terminal functional domain of the 51 proteins had six conserved motifs involved in methylation modification, indicating that the protein has a basic catalytic methylation function and the difference in the N-terminal regulatory domains of the 51 proteins divided the proteins into four classes, MET, CMT, DRM and DNMT2, in which DNMT2 lacks an N-terminal regulatory domain. Gene expression in cotton is not the same under different stress treatments. Different expression patterns of DNA MTases show the functional diversity of the cotton DNA methyltransferase gene family. VIGS silenced Gossypium hirsutum l. in the cotton seedling of DNMT2 family gene GhDMT6, after stress treatment the growth condition was better than the control. The distribution of DNA MTases varies among cotton species. Different DNA MTase family members have different genetic structures, and the expression level changes with different stresses, showing tissue specificity. Under salt and drought stress, G. hirsutum L. TM-1 increased the number of genes more than G. raimondii and G. arboreum L. Shixiya 1. The resistance of Gossypium hirsutum L.TM-1 to cold, drought and salt stress was increased after the plants were silenced with GhDMT6 gene.

genomics↗

bioRxiv2018

Evolution of the highly repetitive PEVK region of titin across mammals

The protein titin plays a key role in vertebrate muscle where it acts like a giant molecular spring. Despite its importance and conservation over vertebrate evolution, a lack of high quality annotations in non-model species makes comparative evolutionary studies of titin challenging. The PEVK region of titin--named for its high proportion of Pro-Glu-Val-Lys amino acids--is particularly difficult to annotate due to its abundance of alternatively spliced isoforms and short, highly repetitive exons. To understand PEVK evolution across mammals, we first developed a bioinformatics tool, PEVK_Finder, to annotate PEVK exons from genomic sequences of titin and then applied it to a diverse set of mammals. PEVK_Finder consistently outperforms standard annotation tools across a broad range of conditions and improves annotations of the PEVK region in non-model mammalian species. We find that the PEVK region can be divided into two subregions (PEVK-N, PEVK-C) with distinct patterns of evolutionary constraint and divergence. The bipartite nature of the PEVK region has implications for titin diversification. In the PEVK-N region, certain exons are conserved and may be essential, but natural selection also acts on particular codons. This region is also rich in glutamate and may contribute to actin binding. In the PEVK-C, exons are more homogenous and length variation of the PEVK region may provide the raw material for evolutionary adaptation in titin function. Taken together, we find that the very complexity that makes titin a challenge for annotation tools may also promote evolutionary adaptation.

genomics↗

bioRxiv2018

Cross-talk between the receptor tyrosine kinases AXL and ERBB3 regulates invadopodia formation in melanoma cells

The invasive phenotype of metastatic cancer cells is accompanied by the formation of actin-rich invadopodia, which adhere to the extracellular matrix, and degrade it. In this study, we explored the role of the tyrosine kinome in the formation of invadopodia in metastatic melanoma cells. Using a microscopy-based siRNA screen, we identified novel invadopodia regulators, the knock-down of which either suppresses (e.g., TYK2, IGFR1, ERBB3, TYRO3, FES, ALK, PTK7) or enhances invadopodia formation and function (e.g., ABL2, AXL, CSK). Particularly intriguing was the discovery that the receptor tyrosine kinase AXL displays a dual regulatory function, manifested by enhancement of invadopodia function upon knock-down or long-term inhibition, as well as following its over-expression. We show here that this apparent contradiction may be attributed to the capacity of AXL to directly stimulate invadopodia; yet its suppression up-regulates the ERBB3 signaling pathway, which consequently activates core invadopodia regulators, and greatly enhances invadopodia function. Bioinformatic analysis of multiple melanoma cells points to an inverse expression pattern of AXL and ERBB3, with the apparent association of high-AXL melanomas, with high expression of invadopodia components and an invasive phenotype. The relevance of these results to melanoma metastasis in vivo, and to potential anti-invasion therapy, is discussed.

cancer biology↗