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Background: Longitudinal studies are needed to decipher mechanistic links between the gut microbiome and nonalcoholic steatohepatitis (NASH). We examined shifts in the gut microbiome in persons with NASH with improvement in liver stiffness measurement (LSM) by magnetic resonance (MR) elastography. Methods: Gut microbial profiling was performed at baseline and study completion (24 weeks) using 16 S rRNA gene sequencing in 69 adults with biopsy-confirmed NASH and significant fibrosis (stages 2–3) enrolled in a multi-center randomized controlled trial evaluating selonsertib alone or in combination with simtuzumab. Differential abundance of bacterial taxa at baseline and end of study were examined in participants with and without longitudinal improvement in LSM. Gut microbial shifts that correlated with secondary outcomes, including reduction in MR imaging-derived proton density fat faction (MRI-PDFF) and histologic fibrosis regression were evaluated. Fecal samples from 32 healthy adults were profiled and genus-level multidimensional scaling was used to determine if microbial shifts in persons with NASH improvement represented a shift toward a healthy gut microbiome. Results: Shifts in abundance of 36 bacterial taxa including Lactobacillus (log2FC = –4.51, FDR < 0.001), Enterococcus (log2FC = –6.72, FDR < 0.001), and Megasphaera (log2FC = 7.74, FDR < 0.001) were associated with improvement in LSM. Improvement in LSM was associated with microbial shifts toward healthy reference (p = 0.05). Significant shifts in 10 and 12 bacterial taxa were associated with improvement in LSM in addition to MRI-PDFF and fibrosis regression, respectively, indicating consistent taxonomic changes across multiple clinical endpoints. Conclusion: Longitudinal changes in the gut microbiota are observed in adults with NASH and clinical improvement and represent a shift toward a healthy microbiome. [ABSTRACT FROM AUTHOR]

Hepatitis B infection is a world-wide public health burden causing serious liver complications. Previous studies suggest that hepatitis B integration into the human genome plays a crucial role in triggering oncogenic process and may also constitutively produce viral antigens. Despite the progress in HBV biology and sequencing technology, our fundamental understanding of how many hepatocytes in the liver actually carry viral integrations is still lacking. Herein we provide evidence that the HBV virus integrates with a lower-bound frequency of 0.84 per diploid genome in hepatitis B positive hepatocellular cancer patients. Moreover, we calculate that integrated viral DNA generates ~80% of the HBsAg transcripts in these patients. These results underscore the need to re-evaluate the clinical end-point and treatment strategies for chronic hepatitis B patients. [ABSTRACT FROM AUTHOR]

Mechanisms that generate transcript diversity are of fundamental importance in eukaryotes. Although a large fraction of human protein-coding genes and lincRNAs produce more than one mRNA isoform each, the regulation of this phenomenon is still incompletely understood. Much progress has been made in deciphering the role of sequence-specific features as well as DNA-and RNA-binding proteins in alternative splicing. Recently, however, several experimental studies of individual genes have revealed a direct involvement of epigenetic factors in alternative splicing and transcription initiation. While histone modifications are generally correlated with overall gene expression levels, it remains unclear how histone modification enrichment affects relative isoform abundance. Therefore, we sought to investigate the associations between histone modifications and transcript diversity levels measured by the rates of transcription start-site switching and alternative splicing on a genome-wide scale across protein-coding genes and lincRNAs. We found that the relationship between enrichment levels of epigenetic marks and transcription start-site switching is similar for protein-coding genes and lincRNAs. Furthermore, we found associations between splicing rates and enrichment levels of H2az, H3K4me1, H3K4me2, H3K4me3, H3K9ac, H3K9me3, H3K27ac, H3K27me3, H3K36me3, H3K79me2, and H4K20me, marks traditionally associated with enhancers, transcription initiation, transcriptional repression, and others. These patterns were observed in both normal and cancer cell lines. Additionally, we developed a novel computational method that identified 840 epigenetically regulated candidate genes and predicted transcription start-site switching and alternative exon splicing with up to 92% accuracy based on epigenetic patterning alone. Our results suggest that the epigenetic regulation of transcript isoform diversity may be a relatively common genome-wide phenomenon representing an avenue of deregulation in tumor development. [ABSTRACT FROM AUTHOR]

Human tumors result from an evolutionary process operating on somatic cells within tissues, whereby natural selection operates on the phenotypic variability generated by the accumulation of genetic, genomic and epigenetic alterations. This somatic evolution leads to adaptations such as increased proliferative, angiogenic, and invasive phenotypes. In this review we outline how cancer genomes are beginning to be investigated from an evolutionary perspective. We describe recent progress in the cataloging of somatic genetic and genomic alterations, and investigate the contributions of germline as well as epigenetic factors to cancer genome evolution. Finally, we outline the challenges facing researchers who investigate the processes driving the evolution of the cancer genome. [Copyright &y& Elsevier]

Vomeronasal sensitivity is important for detecting intraspecific pheromonal cues as well as environmental odorants and is involved in mating, social interaction, and other daily activities of many vertebrates. Two large families of seven-transmembrane G-protein–coupled receptors, V1rs and V2rs, bind to various ligands to initiate vomeronasal signal transduction. Although the macroevolution of V1r and V2r genes has been well characterized throughout vertebrates, especially mammals, little is known about their microevolutionary patterns, which hampers a clear understanding of the evolutionary forces behind the rapid evolutionary turnover of V1r and V2r genes and the great diversity in receptor repertoire across species. Furthermore, the role of divergent vomeronasal perception in enhancing premating isolation and maintaining species identity has not been evaluated. Here we sequenced 44 V1r genes and 25 presumably neutral noncoding regions in 14 wild-caught mice belonging to Mus musculus and M. domesticus, two closely related species with strong yet incomplete reproductive isolation. We found that nucleotide changes in V1rs are generally under weak purifying selection and that only ∼5% of V1rs may have been subject to positive selection that promotes nonsynonymous substitutions. Consistent with the low functional constraints on V1rs, 18 of the 44 V1rs have null alleles segregating in one or both species. Together, our results demonstrate that, despite occasional actions of positive selection, the evolution of V1rs is in a large part shaped by purifying selection and random drift. These findings have broad implications for understanding the driving forces of rapid gene turnovers that are often observed in the evolution of large gene families. [ABSTRACT FROM AUTHOR]

Although our knowledge of the genes and genomes of extinct organisms is improving as a result of progress in sequencing ancient DNA, the transcriptomes of extinct organisms remain inaccessible, owing to the rapid degradation of messenger RNA after death. We provide empirical evidence that gene expression levels in the reproductive tissues of mice and during early mouse development correlate highly with the rate of inherited retroposition: the source of processed pseudogenes in the genome. Thus, processed pseudogenes might serve as fossilized footprints of the expression of their parent genes, shedding light on ancient transcriptomes that could provide significant insights into the evolution of gene expression. [Copyright &y& Elsevier]

Do novel complex traits evolve when pre-existent complex developmental networks are recruited into new places in the body? Here we look closely at the genomic fingerprints that are produced as a result of gene network co-option. [ABSTRACT FROM AUTHOR]

Positive Darwinian selection on advantageous point substitutions has been demonstrated in many genes. We here provide empirical evidence, for the first time, that positive selection can also act on insertion/deletion (indel) substitutions in the evolution of a protein. CATSPER1 is a voltage-gated calcium channel found exclusively in the plasma membrane of the mammalian sperm tail and it is. essential for sperm motility. We determined the DNA sequences of the first exon of the CATSPER1 gene from 15 primates, which encodes the intracellular N terminus region of &ap;400 aa. These sequences exhibit an excessively high frequency of indels. However, all indels have lengths that are multiples of 3 nt (3n indels) and do not disrupt the ORF. The number of indel substitutions per site per year in CATSPER1 is five to eight times the corresponding rates calculated from two large-scale primate genomic comparisons, which represent the neutral rate of indel substitutions. Moreover, CATSPER1 indels are considerably longer than neutral indels. These observations strongly suggest that positive selection has been promoting the fixation of indel mutations in CATSPER1 exon 1. It has been shown in certain ion channels that the length of the N terminus region affects the rate of channel inactivation. This finding suggests that the selection detected may be related to the regulation of the CATSPER1 channel, which can affect sperm motility, an important determinant in sperm competition. [ABSTRACT FROM AUTHOR]

Evolutionary changes in the hepatitis B virus (HBV) genome could reflect its adaptation to host-induced selective pressure. Leveraging paired human exome and ultra-deep HBV genome-sequencing data from 567 affected individuals with chronic hepatitis B, we comprehensively searched for the signatures of this evolutionary process by conducting "genome-to-genome" association tests between all human genetic variants and viral mutations. We identified significant associations between an East Asian-specific missense variant in the gene encoding the HBV entry receptor NTCP (rs2296651, NTCP S267F) and mutations within the receptor-binding region of HBV preS1. Through in silico modeling and in vitro preS1-NTCP binding assays, we observed that the associated HBV mutations are in proximity to the NTCP variant when bound and together partially increase binding affinity to NTCP S267F. Furthermore, we identified significant associations between HLA-A variation and viral mutations in HLA-A-restricted T cell epitopes. We used in silico binding prediction tools to evaluate the impact of the associated HBV mutations on HLA presentation and observed that mutations that result in weaker binding affinities to their cognate HLA alleles were enriched. Overall, our results suggest the emergence of HBV escape mutations that might alter the interaction between HBV PreS1 and its cellular receptor NTCP during viral entry into hepatocytes and confirm the role of HLA class I restriction in inducing HBV epitope variations. [Display omitted] Host-induced selective pressure can shape viral evolution during chronic infections. By analyzing paired human and hepatitis B virus (HBV) genomic data, we pinpoint viral mutations associated with human NTCP and HLA-A variation, shedding light on HBV-evasion strategies that influence viral entry and HLA presentation. [ABSTRACT FROM AUTHOR]

Despite the high global prevalence of chronic hepatitis B (CHB) infection, datasets covering the whole hepatitis B viral genome from large patient cohorts are lacking, greatly limiting our understanding of the viral genetic factors involved in this deadly disease. We performed deep sequencing of viral samples from patients chronically infected with HBV to investigate the association between viral genome variation and patients' clinical characteristics. We discovered novel viral variants strongly associated with viral load and HBeAg status. Patients with viral variants C1817T and A1838G had viral loads nearly three orders of magnitude lower than patients without those variants. These patients consequently experienced earlier viral suppression while on treatment. Furthermore, we identified novel variants that either independently or in combination with precore mutation G1896A were associated with the transition from HBeAg positive to the negative phase of infection. These observations are consistent with the hypothesis that mutation of the HBeAg open reading frame is an important factor driving CHB patient's HBeAg status. This analysis provides a detailed picture of HBV genetic variation in the largest patient cohort to date and highlights the diversity of plausible molecular mechanisms through which viral variation affects clinical phenotype. [ABSTRACT FROM AUTHOR]

In insects, forewings and hindwings usually have different shapes, sizes, and color patterns. A variety of RNAi experiments across insect species have shown that the hox gene Ultrabithorax ( Ubx ) is necessary to promote hindwing identity. However, it remains unclear whether Ubx is sufficient to confer hindwing fate to forewings across insects. Here, we address this question by over-expressing Ubx in the butterfly Bicyclus anynana using a heat-shock promoter. Ubx whole-body over-expression during embryonic and larvae development led to body plan changes in larvae but to mere quantitative changes to adult morphology, respectively. Embryonic heat-shocks led to fused segments, loss of thoracic and abdominal limbs, and transformation of head limbs to larger appendages. Larval heat-shocks led to reduced eyespot size in the expected homeotic direction, but neither additional eyespots nor wing shape changes were observed in forewings as expected of a homeotic transformation. Interestingly, Ubx was found to be expressed in a novel, non-characteristic domain – in the hindwing eyespot centers. Furthermore, ectopic expression of Ubx on the pupal wing activated the eyespot-associated genes spalt and Distal-less , known to be directly repressed by Ubx in the fly׳s haltere and leg primordia, respectively, and led to the differentiation of black wing scales. These results suggest that Ubx has been co-opted into a novel eyespot gene regulatory network, and that it is capable of activating black pigmentation in butterflies. [ABSTRACT FROM AUTHOR]

Investigates the chromosomal speciation hypothesis for humans and chimpanzees. Chromosomal rearrangements; Gene flow; Percentric inversions; Genomic DNA sequence divergence; Divergence of protein coding sequences; Levels of divergence; Synonymous and nonsynonymous divergences among human, chimpanzee and gorilla genes; Implications and conclusions.

More systematic analysis of hepatitis B virus (HBV) genome diversity, linked with tenofovir disoproxil fumarate (TDF) treatment and HBeAg seroconversion, are needed. GS‐US‐174–0115 was a double‐blind, placebo‐controlled, Phase 3, 192‐week clinical trial that evaluated TDF in adolescents with chronic hepatitis B (CHB). HBV full‐genome deep sequencing was performed using Illumina MiSeq at baseline (BL; n = 85), Week 8 (W8; n = 80), Week 72 (W72; PBO only, n = 42), and treatment‐free follow‐up (TDF only, n = 25). The viral diversity was calculated using Shannon entropy and population nucleotide diversity with a 2% variant cutoff. Our data showed (i) a higher viral diversity in the X region at baseline than the core/polymerase/surface regions, (ii) higher core/surface viral diversity at baseline for patients with seroconversion, (iii) an expected reduction in viral diversity after 8 weeks of TDF treatment, and (iv) a drop in viral diversity at W72 for patients receiving placebo with a seroconversion (n = 7). The higher viral diversity in X was associated with higher baseline alanine aminotransferase (ALT) levels (p <.001). Patients with greater reduction of diversity at W8 of TDF treatment had higher baseline ALT levels. For placebo patients who seroconverted, the drop in viral diversity at W72 (p =.04) coincided with reduction of serum HBV DNA (average change from baseline = −4.10 log10 copies/ml) and unique combinations of variants were enriched in a patient's viral population post seroconversion. The basal core promoter (BCP) variants, A1762T and G1764A, and the pC variant, G1896A, were most often enriched at or after seroconversion. [ABSTRACT FROM AUTHOR]

Phylogenetic analyses suggest that violations of "Dollo's law"--that is, re-evolution of lost complex structures--do occur, albeit infrequently. However, the genetic basis of such reversals has not been examined. Here, we address this question using the Drosophila sex comb, a recently evolved, male-specific morphological structure composed of modified bristles. In some species, sex comb development involves only the modification of individual bristles, while other species have more complex "rotated" sex combs that are shaped by coordinated migration of epithelial tissues. Rotated sex combs were lost in the ananassae species subgroup and subsequently re-evolved, ~12 million years later, in Drosophila bipectinata and its sibling species. We examine the genetic basis of the differences in sex comb morphology between D. bipectinata and D. malerkotliana, a closely related species with a much simpler sex comb representing the ancestral condition. QTL mapping reveals that >50% of this difference is controlled by one chromosomal inversion that covers ~5% of the genome. Several other, larger inversions do not contribute appreciably to the phenotype. This genetic architecture suggests that rotating sex combs may have re-evolved through changes in relatively few genes. We discuss potential developmental mechanisms that may allow lost complex structures to be regained. [ABSTRACT FROM AUTHOR]

Background: Non-traditional model systems need new tools that will enable them to enter the field of functional genetics. These tools should enable the exploration of gene function, via knock-downs of endogenous genes, as well as overexpression and ectopic expression of transgenes. Methodology: We constructed a new vector called Pogostick that can be used to over-express or down-regulate genes in organisms amenable to germ line transformation by the piggyBac transposable element. Pogostick can be found at www. addgene.org, a non-profit plasmid repository. The vector currently uses the heat-shock promoter Hsp70 from Drosophila to drive transgene expression and, as such, will have immediate applicability to organisms that can correctly interpret this promotor sequence. We detail how to clone candidate genes into this vector and test its functionality in Drosophila by targeting a gene coding for the fluorescent protein DsRed. By cloning a single DsRed copy into the vector, and generating transgenic lines, we show that DsRed mRNA and protein levels are elevated following heat-shock. When cloning a second copy of DsRed in reverse orientation into a flanking site, and transforming flies constitutively expressing DsRed in the eyes, we show that endogenous mRNA and protein levels drop following heat-shock. We then test the over-expression vector, containing the complete cDNA of Ultrabithorax (Ubx) gene, in an emerging model system, Bicyclus anynana. We produce a transgenic line and show that levels of Ubx mRNA expression rise significantly following a heat-shock. Finally, we show how to obtain genomic sequence adjacent to the Pogostick insertion site and to estimate transgene copy number in genomes of transformed individuals. Significance: This new vector will allow emerging model systems to enter the field of functional genetics with few hurdles. [ABSTRACT FROM AUTHOR]

Identifies two genes (PRM2 and FOXP2) with significantly enhanced evolutionary rates in the hominid lineage, using comparative analysis of human, chimpanzee, and mouse protein sequences. PRM2 is a histone-like protein essential to spermatogenesis and was previously reported to be a likely target of sexual selection in humans and chimpanzees. FOXP2 is a transcription factor involved in speech and language development. Human FOXP2 experienced a >60-fold increase in substitution rate and incorporated two fixed amino acid changes in a broadly defined transcription suppression domain. A survey of a diverse group of placental mammals reveals the uniqueness of the human FOXP2 sequence and a population genetic analysis indicates possible adaptive selection behind the accelerated evolution. Taken together, our results suggest an important role that FOXP2 may have played in the origin of human speech and demonstrate a strategy for identifying candidate genes underlying the emergences of human-specific features.