Identification of protein features encoded by alternative exons using Exon Ontology.

Transcriptomic genome-wide analyses demonstrate massive variation of alternative splicing in many physiological and pathological situations. One major challenge is now to establish the biological contribution of alternative splicing variation in physiological- or pathological-associated cellular phenotypes. Toward this end, we developed a computational approach, named "Exon Ontology," based on terms corresponding to well-characterized protein features organized in an ontology tree. Exon Ontology is conceptually similar to Gene Ontology-based approaches but focuses on exon-encoded protein features instead of gene level functional annotations. Exon Ontology describes the protein features encoded by a selected list of exons and looks for potential Exon Ontology term enrichment. By applying this strategy to exons that are differentially spliced between epithelial and mesenchymal cells and after extensive experimental validation, we demonstrate that Exon Ontology provides support to discover specific protein features regulated by alternative splicing. We also show that Exon Ontology helps to unravel biological processes that depend on suites of coregulated alternative exons, as we uncovered a role of epithelial cell-enriched splicing factors in the AKT signaling pathway and of mesenchymal cell-enriched splicing factors in driving splicing events impacting on autophagy. Freely available on the web, Exon Ontology is the first computational resource that allows getting a quick insight into the protein features encoded by alternative exons and investigating whether coregulated exons contain the same biological information.

Chemical Inhomogeneity in Iron Oxide@CoO Core-Shell Nanoparticles: A Local Probe Study Using Zero-Field and In-Field 57Fe Mössbauer Spectrometry.

Core-shell magnetic nanoparticles type Fe3-xO4 (≈10 nm)@CoO (≈3 nm) prepared by forced hydrolysis in polyol medium have been investigated through the combined use of dc magnetization measurements by SQUID and local analysis by 57Fe Mössbauer spectrometry. A shift of the hysteresis loop along the field axis was observed, which highlights the presence of exchange bias coupling in these nanoparticles. This exchange bias coupling is accompanied by an unexpected high value of coercive field in hysteresis loop in mode ZFC. A local probe study using both zero-field and in-field Mössbauer spectrometry reveals complex hyperfine structures. Great attention is paid to the fitting procedure: it is concluded that Fe3-xO4@CoO nanoparticles result from a mixture of magnetite and maghemite phases in addition to the formation of an interface-like layer close to a cobalt ferrite. Such a structure explains both the high coercive field value observed by SQUID and the presence of exchange bias coupling.

Endothelial, epithelial, and fibroblast cells exhibit specific splicing programs independently of their tissue of origin.

Alternative splicing is the main mechanism of increasing the proteome diversity coded by a limited number of genes. It is well established that different tissues or organs express different splicing variants. However, organs are composed of common major cell types, including fibroblasts, epithelial, and endothelial cells. By analyzing large-scale data sets generated by The ENCODE Project Consortium and after extensive RT-PCR validation, we demonstrate that each of the three major cell types expresses a specific splicing program independently of its organ origin. Furthermore, by analyzing splicing factor expression across samples, publicly available splicing factor binding site data sets (CLIP-seq), and exon array data sets after splicing factor depletion, we identified several splicing factors, including ESRP1 and 2, MBNL1, NOVA1, PTBP1, and RBFOX2, that contribute to establishing these cell type-specific splicing programs. All of the analyzed data sets are freely available in a user-friendly web interface named FasterDB, which describes all known splicing variants of human and mouse genes and their splicing patterns across several dozens of normal and cancer cells as well as across tissues. Information regarding splicing factors that potentially contribute to individual exon regulation is also provided via a dedicated CLIP-seq and exon array data visualization interface. To the best of our knowledge, FasterDB is the first database integrating such a variety of large-scale data sets to enable functional genomics analyses at exon-level resolution.

HTLV-1-infected CD4+ T-cells display alternative exon usages that culminate in adult T-cell leukemia.

BACKGROUND: Reprogramming cellular gene transcription sustains HTLV-1 viral persistence that ultimately leads to the development of adult T-cell leukemia/lymphoma (ATLL). We hypothesized that besides these quantitative transcriptional effects, HTLV-1 qualitatively modifies the pattern of cellular gene expression. RESULTS: Exon expression analysis shows that patients' untransformed and malignant HTLV-1(+) CD4(+) T-cells exhibit multiple alternate exon usage (AEU) events. These affect either transcriptionally modified or unmodified genes, culminate in ATLL, and unveil new functional pathways involved in cancer and cell cycle. Unsupervised hierarchical clustering of array data permitted to isolate exon expression patterns of 3977 exons that discriminate uninfected, infected, and transformed CD4(+) T-cells. Furthermore, untransformed infected CD4+ clones and ATLL samples shared 486 exon modifications distributed in 320 genes, thereby indicating a role of AEUs in HTLV-1 leukemogenesis. Exposing cells to splicing modulators revealed that Sudemycin E reduces cell viability of HTLV-1 transformed cells without affecting primary control CD4+ cells and HTLV-1 negative cell lines, suggesting that the huge excess of AEU might provide news targets for treating ATLL. CONCLUSIONS: Taken together, these data reveal that HTLV-1 significantly modifies the structure of cellular transcripts and unmask new putative leukemogenic pathways and possible therapeutic targets.

Assessment of Feeding Behaviors and Parents' Frustrations of Children with Autism Spectrum Disorder in Lebanon: A Case-Control Study.

Children with autism spectrum disorder (ASD) exhibit restrictive and repetitive behaviors that affect their eating habits. The purpose of this study is to identify the behavioral feeding problems and eating habits among ASD children compared to typically developed (TD) children age/gender-matched controls, along with their parents'/caregivers' strategies for dealing with them. It included 43 ASD children and 43 TD children aged two to eleven years. The analysis was performed based on two valid questionnaires: the Behavior Pediatrics Feeding Assessment Scale (BPFA) and "My Child Eating Habits" (MCEH). The BPFA and MCEH scores conceded three manifestations that fall into food selectivity and problematic mealtime behavior in both groups of children. Compared to TD children, children with ASD exhibited higher BPFA scores, which indicated food-related behavioral and skill-based problems (p = 0.004). Children with ASD were less likely to consume fruits, vegetables, and milk than TD children, which may lead to nutritional deficiencies (p = 0.003, p = 0.003, and p = 0.010, respectively). Parents of ASD children were concerned about their behavioral problems and expressed their intention of an early intervention. These findings highlight the importance of nutritional clinical routines that incorporate the evaluation of the nutritional status and feeding behaviors of ASD children.

COVID-19 home isolation and food consumption patterns: Investigating the correlates of poor dietary diversity in Lebanon: a cross-sectional study.

Background: The unfurling COVID-19 pandemic has uncovered the defenselessness of the Lebanese food system leading to serious implication in maintaining a healthy sustainable lifestyle. Aim: The main purpose of this study is to examine the impact of the COVID-19 pandemic on food consumption patterns and dietary diversity of the Lebanese people. Methods: The online survey, completed between April and June 2020, consisted of a cross-sectional study on 2282 Lebanese participants (mean age: 29.36±12.221, 80.9% women) that was part of a survey across 38 different countries conducted by De Backer, C. et al. A food frequency questionnaire was used to investigate the consumption patterns along with the calculation of the Food Consumption Score (FCS), a proxy indicator of dietary diversity. Data collected on cooking attitudes, shopping, and food stock identify the community mitigation measures. Results: Home isolation due to COVID-19 induced an increase in the consumption of legumes and pulses (3.2%, p-value=0.001) and whole wheat groups (2.8%, p-value=0.03). In contrast, a decrease of 5.4%, 6.9%, 5.8%, 5.1%, 3.1%, 3.4% and 2.8% was observed in the consumption of fruits (p-value=0), vegetables (p-value=0), processed meats, poultry, and fish (p-value=0), other dairy products (p-value=0), sweet snacks (p-value=0.001), sugared beverages (p-value=0), fats and oils (p-value=0.001), respectively. The FCS decreased by 4.6%. As food-related behaviors, most cooking attitudes, and practices (10 out of 13) showed an amelioration during the lockdown and the proportions of food stocked have been changing since the start of the pandemic seeing higher amounts of pasta, rice or other grains, flour, and legumes/pulses stocked. Conclusion: To conclude, the hostile home isolation strategy followed to prevent the COVID-19 spread in Lebanon, came at a high nutritional cost, driving poor dietary diversity.

Dietary Diversity in the Eastern Mediterranean Region Before and During the COVID-19 Pandemic: Disparities, Challenges, and Mitigation Measures.

The COVID-19 pandemic has revealed the Eastern Mediterranean Region's food system's fragility posing severe challenges to maintaining healthy sustainable lifestyle. The aim of this cross-sectional study (N = 13,527 household's family members, mean age: 30.3 ±11.6, 80% women) is to examine the impact of the COVID-19 pandemic on food consumption patterns and household's dietary diversity in 10 Eastern Mediterranean countries. A food frequency questionnaire was used to investigate the consumption patterns along with the calculation of the Food Consumption Score (FCS), a proxy indicator of dietary diversity. Data collected on cooking attitudes, shopping and food stock explore the community mitigation measures. In the overall population, before and during the pandemic, most food groups were consumed less or equal to 4 times per week. As evident from our findings and considering that the pandemic may be better, but it's not over, small to moderate changes in food consumption patterns in relatively short time periods can become permanent and lead to substantial poor dietary diversity over time. While it is a priority to mitigate the immediate impact, one area of great concern is the long-term effects of this pandemic on dietary patterns and dietary diversity in Eastern Mediterranean households. To conclude, the COVID-19 crisis revealed the region's unpreparedness to deal with a pandemic. While the aggressive containment strategy was essential for most countries to help prevent the spread, it came at a high nutritional cost, driving poor dietary diversity.

Sex disparities in food consumption patterns, dietary diversity and determinants of self-reported body weight changes before and amid the COVID-19 pandemic in 10 Arab countries.

Background: The COVID-19 pandemic along with its confinement period boosted lifestyle modifications and impacted women and men differently which exacerbated existing gender inequalities. The main objective of this paper is to assess the gender-based differentials in food consumption patterns, dietary diversity and the determinants favoring weight change before and amid the COVID-19 pandemic among Arab men and women from 10 Arab countries. Methods: A cross-sectional study was conducted based on a convenience sample of 12,447 households' family members (mean age: 33.2 ± 12.9; 50.1% females) and information from participants aged 18 years and above was collected about periods before and during the pandemic. Results: Findings showed that, during the COVID-19 period, the dietary diversity, declined by 1.9% among females compared to males (0.4%) (p < 0.001) and by 1.5% among overweight participants (p < 0.001) compared to their counterparts. Conclusions: To conclude, gender-sensitive strategies and policies to address weight gain and dietary diversity during emergent shocks and pandemics are urgently needed in the region.

Impact of Bariatric Surgery on the Healthy Eating Index, Binge Eating Behavior and Food Craving in a Middle Eastern Population: A Lebanese Experience.

Even though bariatric surgeries (BS) are on the rise in Lebanon and the Middle East, the changes in diet quality, binge eating, and food cravings in this region are poorly studied peri-operatively. This cross-sectional study aimed to assess binge eating behaviors, food craving and the Healthy Eating Index (HEI) in Lebanese patients who underwent BS in a duration that exceeds 6 months. Evaluation included a dietary assessment of usual diet preoperatively and postoperatively. It included the collection of information on sociodemographic, anthropometric and surgical variables, as well as the administration of dietary recalls and questionnaires to calculate the HEI score, the Binge Eating Scale (BES) and the Food Craving Inventory (FCI). Participants (n = 60) were mostly females (85%) who had undergone sleeve gastrectomy (90%), with a mean duration since BS of 2.4 ± 1.8 years. Despite improvements in their HEI scores, 97% of the participants remained in the worst category. The frequency of participants in the severe BES category dropped markedly postoperatively from 78% to 5% (p < 0.01). Food craving followed a similar trend, with scores dropping from 50 ± 36 pre-surgery to 30 ± 25 post surgery (p < 0.01). Weight regain, prevalent among 40% of participants, was predicted by BES. Despite the improvement in BES and FCI, HEI improvement remained shy. Future interventions should validate findings in other countries and assess means for optimizing HEI scores among BS patients in the Middle East region.

Genes devoid of full-length transposable element insertions are involved in development and in the regulation of transcription in human and closely related species.

Transposable elements (TEs) are major components of mammalian genomes, and their impact on genome evolution is now well established. In recent years several findings have shown that they are associated with the expression level and function of genes. In this study, we analyze the relationships between human genes and full-length TE copies in terms of three factors (gene function, expression level, and selective pressure). We classified human genes according to their TE density, and found that TE-free genes are involved in important functions such as development, transcription, and the regulation of transcription, whereas TE-rich genes are involved in functions such as transport and metabolism. This trend is conserved through evolution. We show that this could be explained by a stronger selection pressure acting on both the coding and non-coding regions of TE-free genes than on those of TE-rich genes. The higher level of expression found for TE-rich genes in tumor and immune system tissues suggests that TEs play an important role in gene regulation.

Multispacer sequence typing of Coxiella burnetii from milk and hard tick samples from ruminant farms in Lebanon.

his study was carried out to detect and characterize Coxiella burnetii in ruminant milk samples and in different tick species from seropositive farms in four Lebanese regions. Milk and tick samples were screened for C. burnetii presence by quantitative real-time PCR (qPCR) targeting IS1111 region followed by multispacer sequence typing (MST). The overall positive percentages of 9.6% (27/282) and 95.45% (84/88) for C. burnetii were recorded in ruminant milk and tick samples, respectively. In detail, the C. burnetii DNA was recorded in 52/54 (96.3%) of Rhipicephalus annulatus, 20/21 (95.24%) of Rhipicephalus turanicus, 6/6 (100%) of Hyalomma anatolicum, 5/6 (83.3%) of Rhipicephalus sanguineus and 1/1 of Rhipicephalus bursa. After genotyping of some IS1111-positive samples (17/111), different MST genotypes were identified. Out of 15 positive ticks, 10 were infected with MST2 genotype, 4 were infected with MST7 genotype and 1 was infected with MST57. Moreover, genotypes MST20 and MST58 were found in one cow and one goat milk samples, respectively. The present study confirmed the high genetic diversity of C. burnetii in Lebanon.

The First Serological Study of Q Fever in Humans in Lebanon.

OBJECTIVE: The aim of this study was to estimate, for the first time, the human seroprevalence of Q fever in Lebanon, by assessing the presence of antibodies against the causative agent, Coxiella burnetii. A total number of 421 serum samples (226 females and 196 males) were collected in February 2015 from hospitals and laboratories dispersed in five Lebanese provinces: Akkar, Bekaa, Mount Lebanon, Nabatieh, and South Lebanon. METHODS: Serial testing approach was used. Samples were first screened for IgG phase II antibodies against C. burnetii by Enzyme Linked Immunosorbent Assay (ELISA) Kit. Then, both positive and inconclusive sera were reexamined by immunofluorescence assay (IFA) test with the aims to confirm and specify the infection status (past or probably acute infection) by detecting IgG (I/II) and IgM (I/II) in human sera. RESULTS: Screening of 421 samples was estimated to be 38.70% (95% CI 34-43.3) positive samples, 5.90% (95% CI 3.7-8.2) suspect samples (as doubtful results), and 55.40% (95% CI 50.7-60.1) negative samples. Furthermore, all positive and suspect samples by ELISA test were retested by immunofluorescence assay test (IFAT), and the prevalence of positive sample was 37% and the infection case was recorded: 23.75% (95% CI 19.7-27.8) samples resulted from past infection, 1.9% (95% CI 0.6-3.2) probably acute infection characterized by several dominance clinical symptoms as: fever, cough, headache, difficulty breathing, and atypical pneumonia, and 0.23% (95% CI 0-0.7) inconclusive sample accompanied by different symptoms as bone metastasis and lung cancer. CONCLUSION: The study records the exposition of 37% of 421 patients to C. burnetii distributed in five Lebanese provinces with the highest seroprevalence in Bekaa and Akkar provinces and the lowest reported in Mount Lebanon. This difference may be due to the presence of high density of livestock production and of major agricultural areas in these two provinces.

Diversity and distribution of ticks from domestic ruminants in Lebanon.

Ticks (Acari: Ixodidae) are ectoparasites infesting livestock in every geographic area in the world and they are vectors of several viral, bacterial, and protozoan pathogens to animals and humans worldwide. A deep knowledge of the geographical distribution of these arthropods would have a key role in the control of tick-borne diseases. Few data are available about tick presence in domestic ruminants in Lebanon. The study aimed at providing an analysis of tick presence and distribution in Lebanon. Ticks were collected from cattle, sheep, and goats farms distributed in 6 Lebanese provinces between June and September 2014. A total of 272 adult hard ticks were randomly collected from domestic ruminants (cattle, sheep, and goats) located at 37 Lebanese farms, distributed among 30 villages. Ticks belonged to 4 Ixodidae genera: Rhipicephalus (72.4%), Haemaphysalis (11.4%), Dermacentor (8.1%), and Hyalomma (8.1%). They included the following species: Rhipicephalus annulatus (50.7%), Rhipicephalus turanicus (18.8%), Hyalomma anatolicum (8.1%), Haemaphylasis punctata (11.4%), Dermacentor marginatus (8.1%), Rhipicephalus sanguineus (2.5%), and Rhipicephalus bursa (0.4%). Rhipicephalus turanicus and H. anatolicum were found on cattle, sheep, and goats, R. annulatus on cattle and sheep, R. sanguineus, D. marginatus and Hea. punctata on sheep and goats, while R. bursa was collected only on sheep. Tick species involved in pathogen transmission were found and some of the identi ed species were recorded in Lebanon for the rst time.

TET2 exon 2 skipping is an independent favorable prognostic factor for cytogenetically normal acute myelogenous leukemia (AML): TET2 exon 2 skipping in AML.

In AML, approximately one-third of expressed genes are abnormally spliced, including aberrant TET2 exon 2 expression. In a discovery cohort (n=99), TET2 exon 2 skipping (TET2E2S) was found positively associated with a significant reduction in the cumulative incidence of relapse (CIR). Age, cytogenetics, and TET2E2S were independent prognostic factors for disease-free survival (DFS), and favorable effects on outcomes predominated in cytogenetic normal (CN)-AML and younger patients. Using the same cutoff in a validation cohort of 86 CN-AML patients, TET2E2Shigh patients were found to be younger than TET2low patients without a difference in the rate of complete remission. However, TET2E2Shigh patients exhibited a significantly lower CIR (p<10-4). TET2E2S and FLT3-ITD, but not age or NPM1 mutation status were independent prognostic factors for DFS and event-free survival (EFS), while TET2E2S was the sole prognostic factor that we identified for overall survival (OS). In both the intermediate-1 and favorable ELN genetic categories, TET2E2S remained significantly associated with prolonged survival. There was no correlation between TET2E2S status and outcomes in 34 additional AML patients who were unfit for IC. Therefore our results suggest that assessments of TET2 exon 2 splicing status might improve risk stratification in CN-AML patients treated with IC.

Oncogene- and drug resistance-associated alternative exon usage in acute myeloid leukemia (AML).

In addition to spliceosome gene mutations, oncogene expression and drug resistance in AML might influence exon expression. We performed exon-array analysis and exon-specific PCR (ESPCR) to identify specific landscapes of exon expression that are associated with DEK and WT1 oncogene expression and the resistance of AML cells to AraC, doxorubicin or azacitidine. Data were obtained for these five conditions through exon-array analysis of 17 cell lines and 24 patient samples and were extended through qESPCR of samples from 152 additional AML cases. More than 70% of AEUs identified by exon-array were technically validated through ESPCR. In vitro, 1,130 to 5,868 exon events distinguished the 5 conditions from their respective controls while in vivo 6,560 and 9,378 events distinguished chemosensitive and chemoresistant AML, respectively, from normal bone marrow. Whatever the cause of this effect, 30 to 80% of mis-spliced mRNAs involved genes unmodified at the whole transcriptional level. These AEUs unmasked new functional pathways that are distinct from those generated by transcriptional deregulation. These results also identified new putative pathways that could help increase the understanding of the effects mediated by DEK or WT1, which may allow the targeting of these pathways to prevent resistance of AML cells to chemotherapeutic agents.

A recently evolved class of alternative 3'-terminal exons involved in cell cycle regulation by topoisomerase inhibitors.

Alternative 3'-terminal exons, which use intronic polyadenylation sites, are generally less conserved and expressed at lower levels than the last exon of genes. Here we discover a class of human genes, in which the last exon appeared recently during evolution, and the major gene product uses an alternative 3'-terminal exon corresponding to the ancestral last exon of the gene. This novel class of alternative 3'-terminal exons are downregulated on a large scale by doxorubicin, a cytostatic drug targeting topoisomerase II, and play a role in cell cycle regulation, including centromere-kinetochore assembly. The RNA-binding protein HuR/ELAVL1 is a major regulator of this specific set of alternative 3'-terminal exons. HuR binding to the alternative 3'-terminal exon in the pre-messenger RNA promotes its splicing, and is reduced by topoisomerase inhibitors. These findings provide new insights into the evolution, function and molecular regulation of alternative 3'-terminal exons.

RNA helicases DDX5 and DDX17 dynamically orchestrate transcription, miRNA, and splicing programs in cell differentiation.

The RNA helicases DDX5 and DDX17 are members of a large family of highly conserved proteins that are involved in gene-expression regulation; however, their in vivo targets and activities in biological processes such as cell differentiation, which requires reprogramming of gene-expression programs at multiple levels, are not well characterized. Here, we uncovered a mechanism by which DDX5 and DDX17 cooperate with heterogeneous nuclear ribonucleoprotein (hnRNP) H/F splicing factors to define epithelial- and myoblast-specific splicing subprograms. We then observed that downregulation of DDX5 and DDX17 protein expression during myogenesis and epithelial-to-mesenchymal transdifferentiation contributes to the switching of splicing programs during these processes. Remarkably, this downregulation is mediated by the production of miRNAs induced upon differentiation in a DDX5/DDX17-dependent manner. Since DDX5 and DDX17 also function as coregulators of master transcriptional regulators of differentiation, we propose to name these proteins "master orchestrators" of differentiation that dynamically orchestrate several layers of gene expression.

A comparative analysis of the amounts and dynamics of transposable elements in natural populations of Drosophila melanogaster and Drosophila simulans.

Genes are important in defining genetic variability, but they do not constitute the largest component of genomes, which in most organisms contain large amounts of various repeated sequences including transposable elements (TEs), which have been shown to account for most of the genome size. TEs contribute to genetic diversity by their mutational potential as a result of their ability to insert into genes or gene regulator regions, to promote chromosomal rearrangements, and to interfere with gene networks. Also, TEs may be activated by environmental stresses (such as temperature or radiation) that interfere with epigenetic regulation systems, and makes them powerful mutation agents in nature. To understand the relationship between genotype and phenotype, we need to analyze the portions of the genome corresponding to TEs in great detail, and to decipher their relationships with the genes. For this purpose, we carried out comparative analyses of various natural populations of the closely-related species Drosophila melanogaster and Drosophila simulans, which differ with regard to their TE amounts as well as their ecology and population size.

Splicing switch of an epigenetic regulator by RNA helicases promotes tumor-cell invasiveness.

Both epigenetic and splicing regulation contribute to tumor progression, but the potential links between these two levels of gene-expression regulation in pathogenesis are not well understood. Here, we report that the mouse and human RNA helicases Ddx17 and Ddx5 contribute to tumor-cell invasiveness by regulating alternative splicing of several DNA- and chromatin-binding factors, including the macroH2A1 histone. We show that macroH2A1 splicing isoforms differentially regulate the transcription of a set of genes involved in redox metabolism. In particular, the SOD3 gene that encodes the extracellular superoxide dismutase and plays a part in cell migration is regulated in an opposite manner by macroH2A1 splicing isoforms. These findings reveal a new regulatory pathway in which splicing factors control the expression of histone variant isoforms that in turn drive a transcription program to switch tumor cells to an invasive phenotype.