Comparative characterization of the infant gut microbiome and their maternal lineage by a multi-omics approach.

The human gut microbiome establishes and matures during infancy, and dysregulation at this stage may lead to pathologies later in life. We conducted a multi-omics study comprising three generations of family members to investigate the early development of the gut microbiota. Fecal samples from 200 individuals, including infants (0-12 months old; 55% females, 45% males) and their respective mothers and grandmothers, were analyzed using two independent metabolomics platforms and metagenomics. For metabolomics, gas chromatography and capillary electrophoresis coupled to mass spectrometry were applied. For metagenomics, both 16S rRNA gene and shotgun sequencing were performed. Here we show that infants greatly vary from their elders in fecal microbiota populations, function, and metabolome. Infants have a less diverse microbiota than adults and present differences in several metabolite classes, such as short- and branched-chain fatty acids, which are associated with shifts in bacterial populations. These findings provide innovative biochemical insights into the shaping of the gut microbiome within the same generational line that could be beneficial in improving childhood health outcomes.

Three-year clinical outcomes after transcatheter aortic valve implantation in patients with bicuspid aortic disease: Comparison between self-expanding and balloon-expandable valves.

INTRODUCTION: Bicuspid aortic valve (BAV) stenosis is a complex anatomical scenario for transcatheter aortic valve implantation (TAVI). Favorable short-term clinical outcomes have been reported with TAVI in this setting, but long-term data are scarce. METHODS: We retrospectively included, in a single-center registry, patients with BAV stenosis who underwent TAVI before 2020. We compared patients treated with self-expanding valves (SEV) versus balloon-expandable valves (BEV). The primary endpoint was a composite of all-cause mortality, stroke and need for aortic valve (AV) reintervention at 3 years. Secondary endpoints included each component of the primary endpoint, cardiovascular mortality, permanent pacemaker implantation (PPI) rate, mean gradient and ≥moderate paravalvular leak (PVL) rate. RESULTS: A total of 150 consecutive patients (SEV = 83, BEV = 67) were included. No significant differences were reported between SEV and BEV groups for the primary composite endpoint (SEV 35.9% vs. BEV 32%, p = 0.66), neither for clinical secondary endpoints (all-cause mortality SEV 28.1% vs. BEV 28%, p = 0.988; cardiovascular mortality SEV 14.1% vs. BEV 20%, p = 0.399; stroke SEV 12.5% vs. BEV 6%, p = 0.342; need for AV reintervention SEV 0% vs. BEV 0%; PPI SEV 28.1% vs. BEV 24%, p = 0.620). A lower mean gradient persisted up to 3 years in the SEV group (SEV 8.8 ± 3.8 mmHg vs. BEV 10.7 ± 3.2 mmHg, p = 0.063), while no significant difference was found in the rate of ≥ moderate PVL (SEV 3/30 vs. BEV 0/25, p = 0.242). CONCLUSIONS: In this single center registry, we observed favorable 3-year clinical outcomes in nonselected BAV patients treated with different generation devices, without significant differences between patients receiving SEV or BEV.

Incidence, predictors, and prognostic significance of impaired functional status early after transcatheter aortic valve replacement.

INTRODUCTION AND OBJECTIVES: There are scarce data on the factors associated with impaired functional status after transcatheter aortic valve replacement (TAVR) and its clinical impact. This study aimed to determine the incidence, predictors, and prognostic implications of impaired functional class (NYHA class III-IV) following TAVR. METHODS: This multicenter study included 3462 transarterial TAVR patients receiving newer generation devices. The patients were compared according to their NYHA class at 1 month of follow-up (NYHA I-II vs NYHA III-IV). A multivariate logistic regression was performed to identify the predictors of 30-day NYHA class III-IV. Patient survival was compared with the Kaplan-Meier method and factors associated with decreased survival were identified with Cox regression analysis. RESULTS: The mean age of the study population was 80.3±7.3 years, with 47% of women, and a median Society of Thoracic Surgeons score of 3.8% [IQR, 2.5-5.8]. A total of 208 patients (6%) were in NYHA class III-IV 1 month after TAVR. Predictors of 30-day NYHA class III-IV were baseline NYHA class III-IV (OR, 1.76; 95%CI, 1.08-2.89; P=.02), chronic pulmonary obstructive disease (OR, 1.80; 95%CI, 1.13-2.83; P=.01), and post-TAVR severe mitral regurgitation (OR, 2.00; 95%CI, 1.21-3.31; P<.01). Patients in NYHA class III-IV 1 month after TAVR were at higher risk of death (HR, 3.68; 95%CI, 2.39-5.70; P<.01) and heart failure-related hospitalization (HR, 6.00; 95%CI, 3.76-9.60; P<.01) at 1-year follow-up. CONCLUSIONS: Up to 6% of contemporary TAVR patients exhibited an impaired functional status following TAVR. Worse baseline NYHA class, chronic pulmonary obstructive disease, and severe mitral regurgitation predicted 30-day NYHA class III/IV, and this determined a higher risk of mortality and heart failure hospitalization at 1-year follow-up. Further studies on the prevention and treatment optimization of patients with impaired functional status after TAVR are needed.

Cardiac Death After Transcatheter Aortic Valve Replacement With Contemporary Devices.

BACKGROUND: The burden of cardiac death after transcatheter aortic valve replacement (TAVR), particularly from advanced heart failure (HF) and sudden cardiac death (SCD), remains largely unknown. OBJECTIVES: This study sought to evaluate the incidence and predictors of SCD and HF-related death in TAVR recipients treated with newer-generation devices. METHODS: This study included a total of 5,421 consecutive patients who underwent TAVR with newer-generation devices using balloon (75.7%) or self-expandable (24.3%) valves. RESULTS: After a median follow-up of 2 (IQR: 1-3) years, 976 (18.0%) patients had died, 50.8% from cardiovascular causes. Advanced HF and SCD accounted for 11.6% and 7.5% of deaths, respectively. Independent predictors of HF-related death were atrial fibrillation (HR: 2.17; 95% CI: 1.47-3.22; P < 0.001), prior pacemaker (HR: 1.79; 95% CI: 1.10-2.92; P = 0.01), reduced left ventricular ejection fraction (HR: 1.08 per 5% decrease; 95% CI: 1.01-1.14; P = 0.02), transthoracic approach (HR: 2.50; 95% CI: 1.37-4.55; P = 0.003), and new-onset persistent left bundle branch block (HR: 1.85; 95% CI: 1.14-3.02; P = 0.01). Two baseline characteristics (diabetes, HR: 1.81; 95% CI: 1.13-2.89; P = 0.01; and chronic kidney disease, HR: 1.72; 95% CI: 1.02-2.90; P = 0.04) and 3 procedural findings (valve in valve, HR: 2.17; 95% CI: 1.01-4.64; P = 0.04; transarterial nontransfemoral approach, HR: 2.23; 95% CI: 1.23-4.48; P = 0.01; and periprocedural ventricular arrhythmia, HR: 7.19; 95% CI: 2.61-19.76; P < 0.001) were associated with an increased risk of SCD after TAVR. CONCLUSIONS: Advanced HF and SCD accounted for a fifth of deaths after TAVR in contemporary practice. Potentially treatable factors leading to increased risk of HF deaths and SCD were identified, such as arrhythmia/dyssynchrony factors for HF and valve-in-valve TAVR or periprocedural ventricular arrhythmias for SCD.

Regulation of miRNA expression by α4ß1 integrin-dependent multiple myeloma cell adhesion.

The α4ß1 integrin regulates the trafficking of multiple myeloma (MM) cells and contributes to MM disease progression. MicroRNAs (miRNAs) can have both tumor suppressor and oncogenic roles and thus are key controllers of tumor evolution, and have been associated with different phases of MM pathogenesis. Using small RNAseq analysis, we show here that α4ß1-dependent MM cell adhesion regulates the expression of forty different miRNAs, therefore expanding our current view of the α4ß1 involvement in MM cell biology. Specific upregulation of miR-324-5p and miR-331-3p in cells attached to α4ß1 ligands was confirmed upon silencing the α4 integrin subunit, and their increased levels found to be dependent on Erk1/2- and PI3K-Akt-, but not Src-dependent signaling. Enhanced miR-324-5p expression upon α4ß1-mediated MM cell adhesion aimed the hedgehog (Hh) component SMO, revealing that the miR-324-5p-SMO module represents a α4ß1-regulated pathway that could control Hh-dependent cellular responses in myeloma. Our results open new therapy research avenues around the α4ß1 contribution to MM progression that deserve to be investigated.

Optimal Degree of Balloon-Expandable Transcatheter Valve Oversizing in Patients With Borderline Aortic Annulus Measurements: Insights From a Multicenter Real-World Experience.

BACKGROUND: The potential benefit of using larger or smaller transcatheter heart valves (THV) in patients with borderline aortic annulus measurement (BAM) remains uncertain. The objective of this study was to evaluate the clinical outcomes associated with the selection of larger or smaller THV in the context of BAM. METHODS: This was a multicenter observational study including patients who underwent transcatheter aortic valve replacement with the SAPIEN 3 or SAPIEN 3 Ultra-valve systems (Edwards Lifesciences, Irvine, CA) from April 2014 to June 2021. BAM was defined according to the manufacturer sizing chart and included the following annulus areas: 314 to 346, 400 to 430, 500 to 546 mm2. A 1:1 propensity score matching was used to compare outcomes of patients with larger or smaller THV. RESULTS: From a total of 2467 patients, BAM was identified in 852 patients (34.5%). A larger and smaller THV was selected in 338 (39.7%) and 514 patients (60.3%) patients, respectively. The choice of a larger THV was associated (before and after propensity matching) with a higher risk of new-onset left bundle branch block (HR, 2.25 [95% CI, 1.39-3.65; P=0.001) and permanent pacemaker implantation (HR, 1.86 [95% CI, 1.11-3.09]; P=0.016) without any impact on gradients or the risk of moderate or severe paravalvular regurgitation at discharge (HR, 0.78 [95% CI, 0.41-1.45]; P=0.427). The risk of periprocedural complications such as aortic rupture and tamponade was low (<1%) and similar between groups. CONCLUSIONS: In patient with BAM, selecting a larger SAPIEN 3/Ultra THV increased the risk of conduction disturbances without any benefit on valve hemodynamics and clinical outcomes.

Cusp-overlap technique during TAVI using the self-expanding Portico FlexNav system.

INTRODUCTION AND OBJECTIVES: The cusp overlap technique (COT) has been proposed to reduce conduction disturbances (CD) after transcatheter aortic valve implantation (TAVI) with self-expanding supra-annular devices, but there are scarce data on COT with intra-annular valves. The aim of this study was to determine whether the use of the COT during Portico implantation results in higher valve implantation and lower rates of CD. METHODS: We included 85 patients undergoing TAVI with the Portico FlexNav system: 43 retrospective patients using the standard 3-cusp view and 42 prospective patients with the COT. Primary endpoints were implantation depth and new-onset CD (composite outcome of new-onset left bundle branch block and new permanent pacemaker implantation). RESULTS: COT resulted in a higher implantation depth (noncoronary cusp: 4.9±3.9 vs 7.4±3.0; P=.005) and lower new-onset CD (31.0% vs 58.1%; P=.012), with a tendency toward a lower need for permanent pacemaker implantation (14.3% vs 30.2%, P=.078; 7.7% vs 31.0%; P=.011 in patients without pre-existing right bundle branch block). Transvalvular aortic gradients were slightly lower with COT (8.7±3.7 vs 11.0±6.1; P=.044). There were no differences in technical success or major procedure-related complications. On multivariate analysis, COT use was associated with a lower risk of new-onset CD. CONCLUSIONS: Use of the COT during Portico implantation is feasible and facilitates a higher valve implant, which in turn may help to reduce rates of new-onset CD.

Aortic Sinus Contrast Retention During TAVR: A Warning Sign Preceding a Potential Thrombotic Complication.

Sinus contrast material retention after transcatheter aortic valve replacement (TAVR) is a rare phenomenon that may reflect an increased risk for thrombotic complications. We present 3 cases of persistent contrast agent retention in the sinus of Valsalva during the TAVR procedure that portend the occurrence of embolic stroke or bioprosthetic valve thrombosis. (Level of Difficulty: Advanced.).

Prognostic Implication of Non-Obstructive Coronary Lesions: A New Classification in Different Settings.

The clinical significance of non-obstructive coronary artery disease is the subject of debate. Our objective was to evaluate the long-term cardiovascular prognosis associated with non-obstructive coronary artery disease in patients undergoing coronary angiography, and to conduct a stratification by sex, diabetes, and clinical indication. We designed a multi-centre retrospective longitudinal observational study of 3265 patients that were classified into three groups: normal coronary arteries (lesion <20%, 1426 patients), non-obstructive coronary artery disease (20-50%, 643 patients), and obstructive coronary artery disease (>70%, 1196 patients). During a mean follow-up of 43 months, we evaluated a combined cardiovascular event: acute myocardial infarction, stroke, hospitalization for heart failure, or cardiovascular death. Multivariable-adjusted Cox proportional hazard models showed a worse prognosis in patients with non-obstructive coronary artery disease, in comparison with patients of normal coronary arteries group, in the total population (hazard ratio 1.72, 95% confidence interval 1.23-2.39; p for trend <0.001), in non-diabetics (hazard ratio 2.12, 95% confidence interval: 1.40-3.22), in women (hazard ratio 1.75, 95% confidence interval 1.10-2.77), and after acute coronary syndrome (hazard ratio 2.07, 95% confidence interval 1.25-3.44). In conclusion, non-obstructive coronary artery disease is associated with an impaired long-term cardiovascular prognosis. This association held for non-diabetics, women, and after acute coronary syndrome.

A multilayered post-GWAS assessment on genetic susceptibility to pancreatic cancer.

BACKGROUND: Pancreatic cancer (PC) is a complex disease in which both non-genetic and genetic factors interplay. To date, 40 GWAS hits have been associated with PC risk in individuals of European descent, explaining 4.1% of the phenotypic variance. METHODS: We complemented a new conventional PC GWAS (1D) with genome spatial autocorrelation analysis (2D) permitting to prioritize low frequency variants not detected by GWAS. These were further expanded via Hi-C map (3D) interactions to gain additional insight into the inherited basis of PC. In silico functional analysis of public genomic information allowed prioritization of potentially relevant candidate variants. RESULTS: We identified several new variants located in genes for which there is experimental evidence of their implication in the biology and function of pancreatic acinar cells. Among them is a novel independent variant in NR5A2 (rs3790840) with a meta-analysis p value = 5.91E-06 in 1D approach and a Local Moran's Index (LMI) = 7.76 in 2D approach. We also identified a multi-hit region in CASC8-a lncRNA associated with pancreatic carcinogenesis-with a lowest p value = 6.91E-05. Importantly, two new PC loci were identified both by 2D and 3D approaches: SIAH3 (LMI = 18.24), CTRB2/BCAR1 (LMI = 6.03), in addition to a chromatin interacting region in XBP1-a major regulator of the ER stress and unfolded protein responses in acinar cells-identified by 3D; all of them with a strong in silico functional support. CONCLUSIONS: This multi-step strategy, combined with an in-depth in silico functional analysis, offers a comprehensive approach to advance the study of PC genetic susceptibility and could be applied to other diseases.

DNA Methylation-Derived Immune Cell Profiles, CpG Markers of Inflammation, and Pancreatic Cancer Risk.

BACKGROUND: Pancreatic cancer is projected to become the second most common cause of cancer-related death over the next 5 years. Because inflammation is thought to be a common trajectory for disease initiation, we sought to prospectively characterize immune profiles using DNA methylation markers and examine DNA methylation levels previously linked to inflammation biomarkers to evaluate whether these immune markers play a key role in pancreatic cancer. METHODS: In a nested case-control study pooling three U.S. prospective cohort studies, DNA methylation was measured in prediagnostic leukocytes of incident pancreatic cancer cases and matched controls using the Illumina MethylationEPIC array. Differentially methylated regions were used to predict immune cell types, and CpGs previously associated with inflammatory biomarkers were selected for the analysis. DNA methylation data from a retrospective case-control study conducted in Spain (PanGenEU) was used for independent replication. RESULTS: Immune cell proportions and ratio of cell proportions were not associated with pancreatic cancer risk in the nested case-control study. Methylation extent of CpGs residing in or near gene MNDA was significantly associated with pancreatic cancer risk in the nested case-control study and replicated in PanGenEU. Methylation level of a promoter CpG of gene PIM-1 was associated with survival in both studies. CONCLUSIONS: Using a targeted approach, we identified several CpGs that may play a role in pancreatic carcinogenesis in two large, independent studies with distinct study designs. IMPACT: These findings could provide insight into critical pathways that may help identify new markers of early disease and survival.

Outcomes of predefined optimisation criteria for intravascular ultrasound guidance of left main stenting.

AIMS: This study sought to investigate the prognostic effect of a protocol with optimisation targets for intravascular ultrasound (IVUS)-guided left main (LM) revascularisation. METHODS AND RESULTS: A protocol was prospectively applied for IVUS-guided LM revascularisation (IVUS-PRO group) including predefined optimisation targets. Using propensity score matching, we selected as control groups patients with angiography-guided PCI (ANGIO group) and IVUS-guided PCI (IVUS group) from a large multicentre registry. The primary endpoint was a composite of cardiac death, LM-related infarction and LM revascularisation at 12 months. In each group, 124 patients with comparable characteristics were included. The incidence of the primary outcome was significantly higher in the ANGIO group compared to the IVUS-PRO group (12.9% vs 4.8%, HR 0.35, 95% CI: 0.15 to 0.82, p=0.02), but not with respect to the IVUS group (12.9% vs 8%, HR 0.51, 95% CI: 0.20 to 1.22, p=0.1), driven by a lower rate of LM revascularisation (8% in the ANGIO group, 6.4% in the IVUS group and 3.2% in the IVUS-PRO group). IVUS-PRO resulted in being an independent risk predictor (HR 0.45, 95% CI: 0.15 to 0.98; p=0.041). CONCLUSIONS: IVUS guidance of LM stenting provides prognostic benefit with respect to the use of angiography alone, particularly when following a protocol with these predefined optimisation criteria.

Crosstalk between epigenetic silencing and infection by tobacco rattle virus in Arabidopsis.

DNA methylation is an important epigenetic mechanism for controlling innate immunity against microbial pathogens in plants. Little is known, however, about the manner in which viral infections interact with DNA methylation pathways. Here we investigate the crosstalk between epigenetic silencing and viral infections in Arabidopsis inflorescences. We found that tobacco rattle virus (TRV) causes changes in the expression of key transcriptional gene silencing factors with RNA-directed DNA methylation activities that coincide with changes in methylation at the whole genome level. Viral susceptibility/resistance was altered in DNA (de)methylation-deficient mutants, suggesting that DNA methylation is an important regulatory system controlling TRV proliferation. We further show that several transposable elements (TEs) underwent transcriptional activation during TRV infection, and that TE regulation likely involved both DNA methylation-dependent and -independent mechanisms. We identified a cluster of disease resistance genes regulated by DNA methylation in infected plants that were enriched for TEs in their promoters. Interestingly, TEs and nearby resistance genes were co-regulated in TRV-infected DNA (de)methylation mutants. Our study shows that DNA methylation contributes to modulate the outcome of viral infections in Arabidopsis, and opens up new possibilities for exploring the role of TE regulation in antiviral defence.

Challenges in the Integration of Omics and Non-Omics Data.

Omics data integration is already a reality. However, few omics-based algorithms show enough predictive ability to be implemented into clinics or public health domains. Clinical/epidemiological data tend to explain most of the variation of health-related traits, and its joint modeling with omics data is crucial to increase the algorithm's predictive ability. Only a small number of published studies performed a "real" integration of omics and non-omics (OnO) data, mainly to predict cancer outcomes. Challenges in OnO data integration regard the nature and heterogeneity of non-omics data, the possibility of integrating large-scale non-omics data with high-throughput omics data, the relationship between OnO data (i.e., ascertainment bias), the presence of interactions, the fairness of the models, and the presence of subphenotypes. These challenges demand the development and application of new analysis strategies to integrate OnO data. In this contribution we discuss different attempts of OnO data integration in clinical and epidemiological studies. Most of the reviewed papers considered only one type of omics data set, mainly RNA expression data. All selected papers incorporated non-omics data in a low-dimensionality fashion. The integrative strategies used in the identified papers adopted three modeling methods: Independent, conditional, and joint modeling. This review presents, discusses, and proposes integrative analytical strategies towards OnO data integration.

miR-204-5p and miR-211-5p Contribute to BRAF Inhibitor Resistance in Melanoma.

Melanoma treatment with the BRAF V600E inhibitor vemurafenib provides therapeutic benefits but the common emergence of drug resistance remains a challenge. We generated A375 melanoma cells resistant to vemurafenib with the goal of investigating changes in miRNA expression patterns that might contribute to resistance. Increased expression of miR-204-5p and miR-211-5p occurring in vemurafenib-resistant cells was determined to impact vemurafenib response. Their expression was rapidly affected by vemurafenib treatment through RNA stabilization. Similar effects were elicited by MEK and ERK inhibitors but not AKT or Rac inhibitors. Ectopic expression of both miRNA in drug-naïve human melanoma cells was sufficient to confer vemurafenib resistance and more robust tumor growth in vivo Conversely, silencing their expression in resistant cells inhibited cell growth. Joint overexpression of miR-204-5p and miR-211-5p durably stimulated Ras and MAPK upregulation after vemurafenib exposure. Overall, our findings show how upregulation of miR-204-5p and miR-211-5p following vemurafenib treatment enables the emergence of resistance, with potential implications for mechanism-based strategies to improve vemurafenib responses.Significance: Identification of miRNAs that enable resistance to BRAF inhibitors in melanoma suggests a mechanism-based strategy to limit resistance and improve clinical outcomes. Cancer Res; 78(4); 1017-30. ©2017 AACR.

DoriTool: A Bioinformatics Integrative Tool for Post-Association Functional Annotation.

The emergence of high-throughput data in biology has increased the need for functional in silico analysis and prompted the development of integrative bioinformatics tools to facilitate the obtainment of biologically meaningful data. In this paper, we present DoriTool, a comprehensive, easy, and friendly pipeline integrating biological data from different functional tools. The tool was designed with the aim to maximize reproducibility and reduce the working time of the researchers, especially of those with limited bioinformatics skills, and to help them with the interpretation of the results. DoriTool is based upon an integrative strategy implemented following a modular design pattern. Using scripts written in Bash, Perl and R, it performs a functional in silico analysis annotation at mutation/variant level, gene level, pathway level and network level by combining up-to-date functional and genomic data and integrating also third-party bioinformatics tools in a pipeline. DoriTool uses GRCh37 human assembly and online mode. DoriTool provides nice visual reports including variant annotation, linkage disequilibrium proxies, gene annotation, gene ontology analysis, expression quantitative trait loci results from Genotype-Tissue Expression (GTEx) and coloured pathways. Here, we also show DoriTool functionalities by applying it to a dataset of 13 variants associated with prostate cancer. Project development, released code libraries, GitHub repository (https://github.com/doritool) and documentation are hosted at https://doritool.github.io/. DoriTool is, to our knowledge, the most complete bioinformatics tool offering functional in silico annotation of variants previously associated with a trait of interest, shedding light on the underlying biology and helping the researchers in the interpretation and discussion of the results.

Differential ß-glucosidase expression as a function of carbon source availability in Talaromyces amestolkiae: a genomic and proteomic approach.

BACKGROUND: Genomic and proteomic analysis are potent tools for metabolic characterization of microorganisms. Although cellulose usually triggers cellulase production in cellulolytic fungi, the secretion of the different enzymes involved in polymer conversion is subjected to different factors, depending on growth conditions. These enzymes are key factors in biomass exploitation for second generation bioethanol production. Although highly effective commercial cocktails are available, they are usually deficient for ß-glucosidase activity, and genera like Penicillium and Talaromyces are being explored for its production. RESULTS: This article presents the description of Talaromyces amestolkiae as a cellulase-producer fungus that secretes high levels of ß-glucosidase. ß-1,4-endoglucanase, exoglucanase, and ß-glucosidase activities were quantified in the presence of different carbon sources. Although the two first activities were only induced with cellulosic substrates, ß-glucosidase levels were similar in all carbon sources tested. Sequencing and analysis of the genome of this fungus revealed multiple genes encoding ß-glucosidases. Extracellular proteome analysis showed different induction patterns. In all conditions assayed, glycosyl hydrolases were the most abundant proteins in the supernatants, albeit the ratio of the diverse enzymes from this family depended on the carbon source. At least two different ß-glucosidases have been identified in this work: one is induced by cellulose and the other one is carbon source-independent. The crudes induced by Avicel and glucose were independently used as supplements for saccharification of slurry from acid-catalyzed steam-exploded wheat straw, obtaining the highest yields of fermentable glucose using crudes induced by cellulose. CONCLUSIONS: The genome of T. amestolkiae contains several genes encoding ß-glucosidases and the fungus secretes high levels of this activity, regardless of the carbon source availability, although its production is repressed by glucose. Two main different ß-glucosidases have been identified from proteomic shotgun analysis. One of them is produced under different carbon sources, while the other is induced in cellulosic substrates and is a good supplement to Celluclast in saccharification of pretreated wheat straw.

Diversity and functional convergence of small noncoding RNAs in male germ cell differentiation and fertilization.

The small noncoding RNAs (sncRNAs) are considered as post-transcriptional key regulators of male germ cell development. In addition to microRNAs (miRNAs) and PIWI-interacting RNAs (piRNAs), other sncRNAs generated from small nucleolar RNAs (snoRNAs), tRNAs, or rRNAs processing may also play important regulatory roles in spermatogenesis. By next-generation sequencing (NGS), we characterized the sncRNA populations detected at three milestone stages in male germ differentiation: primordial germ cells (PGCs), pubertal spermatogonia cells, and mature spermatozoa. To assess their potential transmission through the spermatozoa during fertilization, the sncRNAs of mouse oocytes and zygotes were also analyzed. Both, microRNAs and snoRNA-derived small RNAs are abundantly expressed in PGCs but transiently replaced by piRNAs in spermatozoa and endo-siRNAs in oocytes and zygotes. Exhaustive analysis of miRNA sequence variants also shows an increment of noncanonical microRNA forms along male germ cell differentiation. RNAs-derived from tRNAs and rRNAs interacting with PIWI proteins are not generated by the ping-pong pathway and could be a source of primary piRNAs. Moreover, our results strongly suggest that the small RNAs-derived from tRNAs and rRNAs are interacting with PIWI proteins, and specifically with MILI. Finally, computational analysis revealed their potential involvement in post-transcriptional regulation of mRNA transcripts suggesting functional convergence among different small RNA classes in germ cells and zygotes.

Global characterization and target identification of piRNAs and endo-siRNAs in mouse gametes and zygotes.

A set of small RNAs known as rasRNAs (repeat-associated small RNAs) have been related to the down-regulation of Transposable Elements (TEs) to safeguard genome integrity. Two key members of the rasRNAs group are piRNAs and endo-siRNAs. We have performed a comparative analysis of piRNAs and endo-siRNAs present in mouse oocytes, spermatozoa and zygotes, identified by deep sequencing and bioinformatic analysis. The detection of piRNAs and endo-siRNAs in the spermatozoa and revealed also in zygotes, hints to their potential delivery to oocytes during fertilization. However, a comparative assessment of the three cell types indicates that both piRNAs and endo-siRNAs are mainly maternally inherited. Finally, we have assessed the role of the different rasRNA molecules in connection with amplification processes by way of the "ping-pong cycle". Our results suggest that the ping-pong cycle can act on other rasRNAs, such as tRNA- and rRNA-derived fragments, thus not only being restricted to TEs during gametogenesis.