Genes — Open Access Journal of Genetics & Genomics

In South Italy durum wheat (Triticum durum Desf.) has a long-time tradition of growing and breeding. Accessions collected and now preserved ex situ are a valuable genetic resource, but their effective use in agriculture and breeding programs remains very low. In this study, a small number (44) of simple sequence repeats (SSR) molecular markers were used to detect pattern of diversity for 136 accessions collected in South Italy over time, to identify the genepool of origin, and establish similarities with 28 Italian varieties with known pedigree grown in Italy over the same time-period. Phenotyping was conducted for 12 morphophysiological characters of agronomic interest. Based on discriminant analysis of principal components (DAPC) and STRUCTURE analysis six groups were identified, the assignment of varieties reflected the genetic basis and breeding strategies involved in their development. Some “old” varieties grown today are the result of evolution through natural hybridization and conservative pure line selection. A small number of molecular markers and little phenotyping coupled with powerful statistical analysis and comparison to pedigreed varieties can provide enough information on the genetic structure of durum wheat germplasm for a quick screening of the germplasm collection able to identify accessions for breeding or introduction in low input agriculture. Full article

The Asian seabass (Lates calcarifer) is a bony fish from the Latidae family, which is widely distributed in the tropical Indo-West Pacific region. The karyotype of the Asian seabass contains 24 pairs of A chromosomes and a variable number of AT- and GC-rich B chromosomes (Bchrs or Bs). Dot-like shaped and nucleolus-associated AT-rich Bs were microdissected and sequenced earlier. Here we analyzed DNA fragments from Bs to determine their repeat and gene contents using the Asian seabass genome as a reference. Fragments of 75 genes, including an 18S rRNA gene, were found in the Bs; repeats represented 2% of the Bchr assembly. The 18S rDNA of the standard genome and Bs were similar and enriched with fragments of transposable elements. A higher nuclei DNA content in the male gonad and somatic tissue, compared to the female gonad, was demonstrated by flow cytometry. This variation in DNA content could be associated with the intra-individual variation in the number of Bs. A comparison between the copy number variation among the B-related fragments from whole genome resequencing data of Asian seabass individuals identified similar profiles between those from the South-East Asian/Philippines and Indian region but not the Australian ones. Our results suggest that Bs might cause variations in the genome among the individuals and populations of Asian seabass. A personalized copy number approach for segmental duplication detection offers a suitable tool for population-level analysis across specimens with low coverage genome sequencing. Full article

The maintenance of genome integrity is crucial in seeds, due to the constant challenge of several endogenous and exogenous factors. The knowledge concerning DNA damage response and chromatin remodeling during seed development is still scarce, especially in Phaseolus vulgaris L. A transcriptomic profiling of the expression of genes related to DNA damage response/chromatin remodeling mechanisms was performed in P. vulgaris seeds at four distinct developmental stages, spanning from late embryogenesis to seed desiccation. Of the 14,001 expressed genes identified using massive analysis of cDNA ends, 301 belong to the DNA MapMan category. In late embryogenesis, a high expression of genes related to DNA damage sensing and repair suggests there is a tight control of DNA integrity. At the end of filling and the onset of seed dehydration, the upregulation of genes implicated in sensing of DNA double-strand breaks suggests that genome integrity is challenged. The expression of chromatin remodelers seems to imply a concomitant action of chromatin remodeling with DNA repair machinery, maintaining genome stability. The expression of genes related to nucleotide excision repair and chromatin structure is evidenced during the desiccation stage. An overview of the genes involved in DNA damage response and chromatin remodeling during P. vulgaris seed development is presented, providing insights into the mechanisms used by developing seeds to cope with DNA damage. Full article

Zinc homeostasis is essential for all organisms. The Zap1 transcriptional activator regulates these processes in the yeast Saccharomyces cerevisiae. During zinc deficiency, Zap1 increases expression of zinc transporters and proteins involved in adapting to the stress of zinc deficiency. Transcriptional activation by Zap1 can also repress expression of some genes, e.g., RTC4. In zinc-replete cells, RTC4 mRNA is produced with a short transcript leader that is efficiently translated. During deficiency, Zap1-dependent expression of an RNA with a longer transcript leader represses the RTC4 promoter. This long leader transcript (LLT) is not translated due to the presence of small open reading frames upstream of the RTC4 coding region. In this study, we show that the RTC4 LLT RNA also plays a second function, i.e., repression of the adjacent GIS2 gene. In generating the LLT transcript, RNA polymerase II transcribes RTC4 through the GIS2 promoter. Production of the LLT RNA correlates with the decreased expression of GIS2 mRNA and mutations that prevent synthesis of the LLT RNA or terminate it before the GIS2 promoter renders GIS2 mRNA expression and Gis2 protein accumulation constitutive. Thus, we have discovered an unusual regulatory mechanism that uses a bicistronic RNA to control two genes simultaneously. Full article

Fungal infections, such as candidiasis caused by Candida, pose a problem of growing medical concern. In developed countries, the incidence of Candida infections is increasing due to the higher survival of susceptible populations, such as immunocompromised patients or the elderly. Existing treatment options are limited to few antifungal drug families with efficacies that vary depending on the infecting species. In this context, the emergence and spread of resistant Candida isolates are being increasingly reported. Understanding how resistance can evolve within naturally susceptible species is key to developing novel, more effective treatment strategies. However, in contrast to the situation of antibiotic resistance in bacteria, few studies have focused on the evolutionary mechanisms leading to drug resistance in fungal species. In this review, we will survey and discuss current knowledge on the genetic bases of resistance to antifungal drugs in Candida opportunistic pathogens. We will do so from an evolutionary genomics perspective, focusing on the possible evolutionary paths that may lead to the emergence and selection of the resistant phenotype. Finally, we will discuss the potential of future studies enabled by current developments in sequencing technologies, in vitro evolution approaches, and the analysis of serial clinical isolates. Full article

We address the problem of observing personal diploid methylomes, CpG methylome pairs of homologous chromosomes that are distinguishable with respect to phased heterozygous variants (PHVs), which is challenging due to scarcity of PHVs in personal genomes. Single molecule real-time (SMRT) sequencing is promising as it outputs long reads with CpG methylation information, but a serious concern is whether reliable PHVs are available in erroneous SMRT reads with an error rate of ∼15%. To overcome the issue, we propose a statistical model that reduces the error rate of phasing CpG site to 1%, thereby calling CpG hypomethylation in each haplotype with >90% precision and sensitivity. Using our statistical model, we examined GNAS complex locus known for a combination of maternally, paternally, or biallelically expressed isoforms, and observed allele-specific methylation pattern almost perfectly reflecting their respective allele-specific expression status, demonstrating the merit of elucidating comprehensive personal diploid methylomes and transcriptomes. Full article

Long non-coding RNA (lncRNA) research in plants has recently gained momentum taking cues from studies in animals systems. The availability of next-generation sequencing has enabled genome-wide identification of lncRNA in several plant species. Some lncRNAs are inhibitors of microRNA expression and have a function known as target mimicry with the sequestered transcript known as an endogenous target mimic (eTM). The lncRNAs identified to date show diverse mechanisms of gene regulation, most of which remain poorly understood. In this review, we discuss the role of identified putative lncRNAs that may act as eTMs for nutrient-responsive microRNAs (miRNAs) in plants. If functionally validated, these putative lncRNAs would enhance current understanding of the role of lncRNAs in nutrient homeostasis in plants. Full article

It is estimated that 30% of all genes in the mammalian cells are regulated by microRNA (miRNAs). The most relevant miRNAs in a cellular context are not necessarily those with the greatest change in expression levels between healthy and diseased tissue. Differentially expressed (DE) miRNAs that modulate a large number of messenger RNA (mRNA) transcripts ultimately have a greater influence in determining phenotypic outcomes and are more important in a global biological context than miRNAs that modulate just a few mRNA transcripts. Here, we describe the development of a tool, “miRmapper”, which identifies the most dominant miRNAs in a miRNA–mRNA network and recognizes similarities between miRNAs based on commonly regulated mRNAs. Using a list of miRNA–target gene interactions and a list of DE transcripts, miRmapper provides several outputs: (1) an adjacency matrix that is used to calculate miRNA similarity utilizing the Jaccard distance; (2) a dendrogram and (3) an identity heatmap displaying miRNA clusters based on their effect on mRNA expression; (4) a miRNA impact table and (5) a barplot that provides a visual illustration of this impact. We tested this tool using nonmetastatic and metastatic bladder cancer cell lines and demonstrated that the most relevant miRNAs in a cellular context are not necessarily those with the greatest fold change. Additionally, by exploiting the Jaccard distance, we unraveled novel cooperative interactions between miRNAs from independent families in regulating common target mRNAs; i.e., five of the top 10 miRNAs act in synergy. Full article

RIG-I-like receptors (retinoic acid-inducible gene-I-like receptors, or RLRs) are family of pattern-recognition receptors for RNA viruses, consisting of three members: retinoic acid-inducible gene I (RIG-I), melanoma differentiation-associated gene 5 (MDA5) and laboratory of genetics and physiology 2 (LGP2). To understand the role of RLRs in bird evolution, we performed molecular evolutionary analyses on the coding genes of avian RLRs using filtered predicted coding sequences from 62 bird species. Among the three RLRs, conservation score and dN/dS (ratio of nonsynonymous substitution rate over synonymous substitution rate) analyses indicate that avian MDA5 has the highest conservation level in the helicase domain but a lower level in the caspase recruitment domains (CARDs) region, which differs from mammals; LGP2, as a whole gene, has a lower conservation level than RIG-I or MDA5. We found evidence of positive selection across all bird lineages in RIG-I and MDA5 but only on the stem lineage of Galliformes in LGP2, which could be related to the loss of RIG-I in Galliformes. Analyses also suggest that selection relaxation may have occurred in LGP2 during the middle of bird evolution and the CARDs region of MDA5 contains many positively selected sites, which might explain its conservation level. Spearman’s correlation test indicates that species-to-ancestor dN/dS of RIG-I shows a negative correlation with endogenous retroviral abundance in bird genomes, suggesting the possibility of interaction between immunity and endogenous retroviruses during bird evolution. Full article

Unverricht-Lundborg disease (ULD) is a common form of progressive myoclonic epilepsy caused by mutations in the cystatin B gene (CSTB) that encodes an inhibitor of several lysosomal cathepsins. Presently, only pharmacological treatment and psychosocial support are available for ULD patients. To overcome the pathogenic effect of the ULD splicing mutation c.66G>A (exon 1), we investigated whether an antisense oligonucleotide therapeutic strategy could correct the defect in patient cells. A specific locked nucleic acid (LNA) antisense oligonucleotide was designed to block a cryptic 5′ss in intron 1. Overall, this approach allowed the restoration of the normal splicing pattern. Furthermore, the recovery was both sequence and dose-specific. In general, this work provides a proof of principle on the correction of a CSTB gene defect causing ULD through a mutation-specific antisense therapy. It adds evidence to the feasibility of this approach, joining the many studies that are paving the way for translating antisense technology into the clinical practice. The insights detailed herein make mutation-based therapy a clear candidate for personalized treatment of ULD patients, encouraging similar investigations into other genetic diseases. Full article

Congenital conotruncal heart defects (CCHD) are a subset of serious congenital heart defects (CHD) of the cardiac outflow tracts or great arteries. Its frequency is estimated in 1/1000 live births, accounting for approximately 10–30% of all CHD cases. Chromosomal abnormalities and copy number variants (CNVs) contribute to the disease risk in patients with syndromic and/or non-syndromic forms. Although largely studied in several populations, their frequencies are barely reported for Latin American countries. The aim of this study was to analyze chromosomal abnormalities, 22q11 deletions, and other genomic imbalances in a group of Argentinean patients with CCHD of unknown etiology. A cohort of 219 patients with isolated CCHD or associated with other major anomalies were referred from different provinces of Argentina. Cytogenetic studies, Multiplex-Ligation-Probe-Amplification (MLPA) and fluorescent in situ hybridization (FISH) analysis were performed. No cytogenetic abnormalities were found. 22q11 deletion was found in 23.5% of the patients from our cohort, 66% only had CHD with no other major anomalies. None of the patients with transposition of the great vessels (TGV) carried the 22q11 deletion. Other 4 clinically relevant CNVs were also observed: a distal low copy repeat (LCR)D-E 22q11 duplication, and 17p13.3, 4q35 and TBX1 deletions. In summary, 25.8% of CCHD patients presented imbalances associated with the disease. Full article

The World Health Organization plans to eliminate hepatitis B and C Infections by 2030. Therefore, there is a need to study and understand hepatitis B virus (HBV) epidemiology and viral evolution further, including evaluating occult (HBsAg-negative) HBV infection (OBI), given that such infections are frequently undiagnosed and rarely treated. We aimed to molecularly characterize HBV genomes from 108 individuals co-infected with human immunodeficiency virus (HIV) and chronic hepatitis B (CHB) or OBI identified from previous HIV studies conducted in Botswana from 2009 to 2012. Full-length (3.2 kb) and nearly full-length (~3 kb) genomes were amplified by nested polymerase chain reaction (PCR). Sequences from OBI participants were compared to sequences from CHB participants and GenBank references to identify OBI-unique mutations. HBV genomes from 50 (25 CHB and 25 OBI) individuals were successfully genotyped. Among OBI participants, subgenotype A1 was identified in 12 (48%), D3 in 12 (48%), and E in 1 (4%). A similar genotype distribution was observed in CHB participants. Whole HBV genome sequences from Botswana, representing OBI and CHB, were compared for the first time. There were 43 OBI-unique mutations, of which 26 were novel. Future studies using larger sample sizes and functional analysis of OBI-unique mutations are warranted. Full article

The derived human alcohol dehydrogenase (ADH)1B*48His allele of the ADH1B Arg48His polymorphism (rs1229984) has been identified as one component of an East Asian specific core haplotype that underwent recent positive selection. Our study has been extended to Southwest Asia and additional markers in East Asia. F_st values (Sewall Wright’s fixation index) and long-range haplotype analyses identify a strong signature of selection not only in East Asian but also in Southwest Asian populations. However, except for the ADH2B*48His allele, different core haplotypes occur in Southwest Asia compared to East Asia and the extended haplotypes also differ. Thus, the ADH1B*48His allele, as part of a core haplotype of 10 kb, has undergone recent rapid increases in frequency independently in the two regions after divergence of the respective populations. Emergence of agriculture may be the common factor underlying the evident selection. Full article

Phytochrome-interacting factors (PIFs), as the basic helix–loop–helix (bHLH) transcription factors, are the primary signaling partners for phytochromes (PHY) that play a key role in PHY-mediated light signal transduction. At present, there are few studies on PIFs in fruit trees. In order to clarify the status of PIFs in grapevines, we identified members of the grape PIFs family and conducted phylogenetic and expression analysis. We identified PIF1, PIF3, PIF4, and PIF7 in PIFs families of the grapevine (Vitis vinifera L.), which were distributed on four different chromosomes with similar gene structures. Except for the closer relationship with PIF1 of citrus, PIFs of grape were distant from the other fruit species such as apple, pear, peach, and strawberry. The VvPIFs (except VvPIF4) were located in the syntenic block with those from Arabidopsisthaliana, Solanum lycopersicum, or Citrus sinensis. In addition to PIF1, all PIFs in grapevines have conserved active PHYB binding (APB) sequences. VvPIF1 has a conserved PIF1-specific active PHYA binding (APA) sequence, while amino acid mutations occurred in the specific APA sequence in VvPIF3. Interestingly, two specific motifs were found in the PIF4 amino acid sequence. The photoreceptor-related elements in the VvPIFs promoter region were the most abundant. PIF1, LONG HYPOCOTYL 5 (HY5) and PIF3, PIF4, GIBBERELLIC ACID INSENSITIVE 1 (GAI1) may interact with each other and participate together in light signal transduction. The relative expression levels of the VvPIFs showed diverse patterns in the various organs at different developmental stages, of which PIF4 was most highly expressed. Prior to maturation, the expression of PIF4 and PIF7 in the skin of the different cultivars increased, while the expression of all PIFs in the flesh decreased. The transcription level of PIFs in grape leaves was sensitive to changes in lighting and shading. Shading treatment was beneficial for enhancing the transcription level of VvPIFs, but the effect on VvPIF3 and VvPIF4 was time-controlled. We concluded that PIFs in grapevines are both conservative and species-specific. The identification and analysis of grape PIFs could provide a theoretical foundation for the further construction of grape light regulation networks. Full article

Protein kinase B (Akt1) is a proto-oncogene that is overactive in most cancers. Akt1 activation requires phosphorylation at Thr308; phosphorylation at Ser473 further enhances catalytic activity. Akt1 activity is also regulated via interactions between the kinase domain and the N-terminal auto-inhibitory pleckstrin homology (PH) domain. As it was previously difficult to produce Akt1 in site-specific phosphorylated forms, the contribution of each activating phosphorylation site to auto-inhibition was unknown. Using a combination of genetic code expansion and in vivo enzymatic phosphorylation, we produced Akt1 variants containing programmed phosphorylation to probe the interplay between Akt1 phosphorylation status and the auto-inhibitory function of the PH domain. Deletion of the PH domain increased the enzyme activity for all three phosphorylated Akt1 variants. For the doubly phosphorylated enzyme, deletion of the PH domain relieved auto-inhibition by 295-fold. We next found that phosphorylation at Ser473 provided resistance to chemical inhibition by Akti-1/2 inhibitor VIII. The Akti-1/2 inhibitor was most effective against pAkt1^T308 and showed four-fold decreased potency with Akt1 variants phosphorylated at Ser473. The data highlight the need to design more potent Akt1 inhibitors that are effective against the doubly phosphorylated and most pathogenic form of Akt1. Full article

Tissue-specific gene expression has long been recognized as a crucial key for understanding tissue development and function. Efforts have been made in the past decade to identify tissue-specific expression profiles, such as the Human Proteome Atlas and FANTOM5. However, these studies mainly focused on “qualitatively tissue-specific expressed genes” which are highly enriched in one or a group of tissues but paid less attention to “quantitatively tissue-specific expressed genes”, which are expressed in all or most tissues but with differential expression levels. In this study, we applied machine learning algorithms to build a computational method for identifying “quantitatively tissue-specific expressed genes” capable of distinguishing 25 human tissues from their expression patterns. Our results uncovered the expression of 432 genes as optimal features for tissue classification, which were obtained with a Matthews Correlation Coefficient (MCC) of more than 0.99 yielded by a support vector machine (SVM). This constructed model was superior to the SVM model using tissue enriched genes and yielded MCC of 0.985 on an independent test dataset, indicating its good generalization ability. These 432 genes were proven to be widely expressed in multiple tissues and a literature review of the top 23 genes found that most of them support their discriminating powers. As a complement to previous studies, our discovery of these quantitatively tissue-specific genes provides insights into the detailed understanding of tissue development and function. Full article

Both type 1 and type 2 diabetes are conditions that are associated with the loss of insulin-producing β-cells within the pancreas. An active research therefore aims at regenerating these β-cells with the hope that they could restore euglycemia. The approaches classically used consist in mimicking embryonic development, making use of diverse cell sources or converting pre-existing pancreatic cells. Despite impressive progresses and promising successes, it appears that we still need to gain further insight into the molecular mechanisms underlying β-cell development. This becomes even more obvious with the emergence of a relatively new field of research, epigenetics. The current review therefore focuses on the latest advances in this field in the context of β-cell (neo-)genesis research. Full article

Candida albicans mutants deficient in homologous recombination (HR) are extremely sensitive to the alkylating agent methyl-methane-sulfonate (MMS). Here, we have investigated the role of HR genes in the protection and repair of C. albicans chromosomes by taking advantage of the heat-labile property (55 °C) of MMS-induced base damage. Acute MMS treatments of cycling cells caused chromosome fragmentation in vitro (55 °C) due to the generation of heat-dependent breaks (HDBs), but not in vivo (30 °C). Following removal of MMS wild type, cells regained the chromosome ladder regardless of whether they were transferred to yeast extract/peptone/dextrose (YPD) or to phosphate buffer saline (PBS); however, repair of HDB/chromosome restitution was faster in YPD, suggesting that it was accelerated by metabolic energy and further fueled by the subsequent overgrowth of survivors. Compared to wild type CAI4, chromosome restitution in YPD was not altered in a Carad59 isogenic derivative, whereas it was significantly delayed in Carad51 and Carad52 counterparts. However, when post-MMS incubation took place in PBS, chromosome restitution in wild type and HR mutants occurred with similar kinetics, suggesting that the exquisite sensitivity of Carad51 and Carad52 mutants to MMS is due to defective fork restart. Overall, our results demonstrate that repair of HDBs by resting cells of C. albicans is rather independent of CaRad51, CaRad52, and CaRad59, suggesting that it occurs mainly by base excision repair (BER). Full article