Primary intracranial sarcoma, DICER-1 mutant, with hemorrhagic presentation: A case report.

Background: Primary intracranial sarcomas (PIS) are rare tumors with mesenchymal origins. These tumors have a heterogeneous clinical presentation and are associated with a poor prognosis. Case Description: This report highlights the complexities associated with PIS by focusing on a 26-year-old male with recurrent tumor growth facing unique challenges regarding diagnosis and treatment options . A high-grade spindle-celled neoplasm with sarcomatous features characterized the patient's tumor. There were additional morphologic changes, including multinucleated giant cells and rare foci with eosinophilic spheroids. Genomic analysis revealed a DICER1-associated PIS. Treatment involved endovascular embolization, multiple surgical interventions, intrathecal etoposide injections, and oral pazopanib with adjuvant radiation therapy. Conclusion: This case additionally highlights an unusual association between PIS and anomalous hypervascularity, refractory hemorrhage, and subdural effusions, a presentation that is increasingly being reported in this type of tumor.

Exomic and epigenomic analysis of pulmonary blastoma.

OBJECTIVE: Pulmonary blastoma is a rare, biphasic, adult-onset lung tumor. In this study, we investigate whether DICER1 pathogenic variants are a feature of pulmonary blastomas through in-depth analysis of the molecular events defining them. METHODS: We performed exome-wide sequencing and DNA methylation profiling of 8 pulmonary blastomas from 6 affected persons. RESULTS: We identified biallelic somatic DICER1 pathogenic variants in 7 of 8 cases. The remaining case had a solitary missense pathogenic variant in the RNase IIIb domain of DICER1. Six of 8 cases carried a CTNNB1 hotspot variant and 4 of 8 had a somatic pathogenic variant in TP53. Methylation analysis showed that the pulmonary blastomas clustered with other DICER1-mutated tumors and not with other more common types of lung cancer. CONCLUSION: We conclude somatic DICER1 pathogenic variants are the major driver of pulmonary blastoma and are likely to act in conjunction with CTNNB1 hotspot variants that are often present.

Structural determinants in the miRNA/miRNA* duplex and the DCL1 PAZ domain for precise and efficient plant miRNA processing.

The accessory proteins Hyponastic-like 1 (HYL1) and Serrated (SE) enhance the precise and efficient processing of miRNAs by Dicer-like 1 (DCL1), which is important for proper miRNA function. However, other factors determining the precision and efficiency of miRNA biogenesis are not well-known. Here, we found that an asymmetric bulge (AB) at the 3' end of miR-5p (produced from the 5' arm of the pre-miRNA) reduced the precision of the second cleavage, whereas an AB at other sites of miR-5p mainly affected the accumulation level of miR-5p in transient expression in Nicotiana benthamiana. In contrast, many ABs in miR-3p (produced from the 3' arm of the pre-miRNA) impose strong negative impact on the processing precision and the accumulation level of miR-5p in N. benthamiana. Arabidopsis DCL1/SE/HYL1 complex-mediated miRNA processing was reconstituted in Saccharomyces cerevisiae to further investigate AB-mediated interference with DCL1 processing. With this system, the positional effect of AB on miRNA processing was tested. The results showed that ABs on the middle of miR-5p have less of an impact on DCL1 cleavage efficiency and precision, whereas those on miR-3p or near the ends of miR-5p strongly reduce DCL1 cleavage activity, precision or both. Studies using the yeast miRNA processing system and transgenic Arabidopsis also revealed the importance of the interaction between the 2-nt 3' overhang of pre-miRNA and the 3' overhang binding pocket (3'BP) on the precision of the second cleavage reaction for many endogenous miRNAs. These findings provide new insights into the mechanism of miRNA biogenesis.

Increased viral tolerance mediates by antiviral RNA interference in bat cells.

Bats harbor highly virulent viruses that can infect other mammals, including humans, posing questions about their immune tolerance mechanisms. Bat cells employ multiple strategies to limit virus replication and virus-induced immunopathology, but the coexistence of bats and fatal viruses remains poorly understood. Here, we investigate the antiviral RNA interference pathway in bat cells and discover that they have an enhanced antiviral RNAi response, producing canonical viral small interfering RNAs upon Sindbis virus infection that are missing in human cells. Disruption of Dicer function results in increased viral load for three different RNA viruses in bat cells, indicating an interferon-independent antiviral pathway. Furthermore, our findings reveal the simultaneous engagement of Dicer and pattern-recognition receptors, such as retinoic acid-inducible gene I, with double-stranded RNA, suggesting that Dicer attenuates the interferon response initiation in bat cells. These insights advance our comprehension of the distinctive strategies bats employ to coexist with viruses.

DICER1-sarcomas of GYN tract: Expanding on an emerging entity.

Tumors with pathogenic DICER1 mutation are rare and encompass sporadic or hereditary benign, intermediate and malignant tumors. DICER1-associated sarcomas are heterogeneous; however, the prototypical ones in the GYN-tract include embryonal rhabdomyosarcoma, adenosarcoma and moderately to poorly differentiated Sertoli-Leydig tumor. In this report, we present three unique uterine sarcomas with DICER1 mutation and remarkable diffuse round/spindle cell morphology. The tumors occurred in cervix (n = 1), and uterine corpus (n = 2). The patient ages were 30, 37 and 59 years with tumor size of 8.8, 10 and 8.6 cm, respectively. Morphologically all three tumors were characterized by distinct spindle/round cell morphology and various amounts of neuroectodermal differentiation (yolk sac-like tubules, blastomatous areas and rosette formation). Other morphologic features of DICER1-sarcoma reported in the literature including cambium layer, focal or diffuse anaplasia, solid and cystic architecture, and chondroid/osteoid areas were absent. All three sarcomas were positive for SALL4 and had variable neuroendocrine marker expression. Whole genome methylation analysis was performed on one of the uterine sarcomas, which clustered the tumor with embryonal tumor with multilayered rosettes. Follow up information was available on all three cases. Two patients were alive with no evidence of disease 13 and 14 months post operation, while one patient had imaging evidence of local recurrence 4 months post operation. In summary, we describe three unique DICER1-sarcomas and expand the phenotypic spectrum of this emerging entity, particularly with GYN-tract origin.

A genome-first approach to characterize DICER1 pathogenic variant prevalence, penetrance and cancer, thyroid, and other phenotypes in 2 population-scale cohorts.

Purpose: Population-scale, exome-sequenced cohorts with linked electronic health records (EHR) permit genome-first exploration of phenotype. Phenotype and cancer risk are well-characterized in children with a pathogenic DICER1 (HGNC ID:17098) variant. Here, the prevalence, penetrance and phenotype of pathogenic germline DICER1 variants in adults was investigated in two population-scale cohorts. Methods: Variant pathogenicity was classified using published DICER1 ClinGen criteria in the UK Biobank (469,787 exomes; unrelated: 437,663) and Geisinger (170,503 exomes; unrelated: 109,789) cohorts. In the UK Biobank cohort, cancer diagnoses in the EHR, cancer and death registry were queried. For the Geisinger cohort, the Geisinger Cancer Registry and EHR were queried. Results: In the UK Biobank, there were 46 unique pathogenic DICER1 variants in 57 individuals (1:8,242;95%CI:1:6,362-1:10,677). In Geisinger, there were 16 unique pathogenic DICER1 variants (including one microdeletion) in 21 individuals (1:8,119;95%CI:1:5,310-1:12,412). Cohorts were well-powered to find larger effect sizes for common cancers. Cancers were not significantly enriched in DICER1 heterozygotes; however, there was a ~4-fold increased risk for thyroid disease in both cohorts. There were multiple ICD10 codes enriched >2-fold in both cohorts. Conclusion: Estimates of pathogenic germline DICER1 prevalence, thyroid disease penetrance and cancer phenotype from genomically ascertained adults are determined in two large cohorts.

Fertility-sparing strategy in a rare case of highly malignant Dicer-1-associated sarcoma of the cervix.

INTRODUCTION: We present the rare case of an 18-year-old patient with a Dicer-1 mutation-associated sarcoma of the cervix uteri. CASE: The patient presented with irregular vaginal bleeding in July 2022. The clinical examination showed an exophytic tumor of the cervix, uterus and ovaries were normal in sonogram. The tumor of the cervix was resected, followed by a diagnostic hysteroscopy and abrasion of the uterine cervix and cavity. Hysteroscopy showed normal findings of the cervix and uterus. After diagnosis of a highly malignant Dicer-1 mutation-associated sarcoma of the cervix, cryopreservation of oocytes was realized. Based on the principle of obtaining maximum oncological safety while preserving fertility in this 18-year-old patient, we recommended chemotherapy rather than radiation with its far severe implications on the patient´s reproductive organs. 4 cycles of chemotherapy consisting of doxorubicin and ifosfamide were applied until December 2022. After re-staging in December 2022 via CT scan and MRI, the abdomen and pelvis as well as control hysteroscopy and abrasion were unremarkable. Until now, the patient is tumor free. DISCUSSION: Primary sarcomas of the cervix are very rare. Recent literature hints towards a distinct DICER-1 sarcoma entity characterized by specific mutational clusters. Limited follow-up data suggested that DICER1-mutant tumors might exhibit a less aggressive clinical course than DICER1-wild-type tumors. CONCLUSION: Decision-making in case of rare histological entities with sparse recommendations in the literature poses a challenge to the treating physician. Treatment strategies should consider oncological safety as well as options of preserving fertility. Gonadotoxic potential of different strategies should be taken into consideration and discussed in detail with the affected patient.

Double-stranded RNA induces antiviral transcriptional response through the Dicer-2/Ampk/FoxO axis in an arthropod.

Invertebrates mainly rely on sequence-specific RNA interference (RNAi) to resist viral infections. Increasing studies show that double-stranded RNA (dsRNA) can induce sequence-independent protection and that Dicer-2, the key RNAi player that cleaves long dsRNA into small interfering RNA (siRNA), is necessary for this protection. However, how this protection occurs remains unknown. Herein, we report that it is caused by adenosine triphosphate (ATP)-hydrolysis accompanying the dsRNA-cleavage. Dicer-2 helicase domain is ATP-dependent; therefore, the cleavage consumes ATP. ATP depletion activates adenosine monophosphate-activated protein kinase (Ampk) and induces nuclear localization of Fork head box O (FoxO), a key transcriptional factor for dsRNA-induced genes. siRNAs that do not require processing cannot activate the transcriptional response. This study reveals a unique nonspecific antiviral mechanism other than the specific RNAi in shrimp. This mechanism is functionally similar to, but mechanistically different from, the dsRNA-activated antiviral response in vertebrates and suggests an interesting evolution of innate antiviral immunity.

Non-C19MC-altered embryonal tumor with multilayered rosettes in a young woman with DICER1 syndrome: case report and review of the literature.

Embryonal tumors with multilayered rosettes (ETMR) are highly aggressive and therapy-resistant pediatric central nervous system (CNS) tumors that have three histological patters: embryonal tumor with abundant neuropil and true rosettes, ependymoblastoma, and medulloepithelioma. We present a case of ETMR in an 18-year-old woman with DICER1 syndrome. This report confirms the important role of DNA-methylation analysis in the classification of CNS embryonal tumors and the importance of investigating somatic and germline DICER1 mutations in all CNS embryonal tumors.

Incautious design of shRNAs for stable overexpression of miRNAs could result in generation of undesired isomiRs.

shRNA-mediated strategy of miRNA overexpression based on RNA Polymerase III (Pol III) expression cassettes is widely used for miRNA functional studies. For some miRNAs, e.g., encoded in the genome as a part of a polycistronic miRNA cluster, it is most likely the only way for their individual stable overexpression. Here we have revealed that expression of miRNAs longer than 19 nt (e.g. 23 nt in length hsa-miR-93-5p) using such approach could be accompanied by undesired predominant generation of 5' end miRNA isoforms (5'-isomiRs). Extra U residues (up to five) added by Pol III at the 3' end of the transcribed shRNA during transcription termination could cause a shift in the Dicer cleavage position of the shRNA. This results in the formation of 5'-isomiRs, which have a significantly altered seed region compared to the initially encoded canonical hsa-miR-93-5p. We demonstrated that the commonly used qPCR method is insensitive to the formation of 5'-isomiRs and cannot be used to confirm miRNA overexpression. However, the predominant expression of 5'-isomiRs without three or four first nucleotides instead of the canonical isoform could be disclosed based on miRNA-Seq analysis. Moreover, mRNA sequencing data showed that the 5'-isomiRs of hsa-miR-93-5p presumably regulate their own mRNA targets. Thus, omitting miRNA-Seq analysis may lead to erroneous conclusions regarding revealed mRNA targets and possible molecular mechanisms in which studied miRNA is involved. Overall, the presented results show that structures of shRNAs for stable overexpression of miRNAs requires careful design to avoid generation of undesired 5'-isomiRs.

Liver-specific DICER1 syndrome model mice develop cystic liver tumors with defective primary cilia.

DICER1 syndrome is a tumor predisposition syndrome caused by familial genetic mutations in DICER1. Pathogenic variants of DICER1 have been discovered in many rare cancers, including cystic liver tumors. However, the molecular mechanisms underlying liver lesions induced by these variants remain unclear. In the present study, we sought to gain a better understanding of the pathogenesis of these variants by generating a mouse model of liver-specific DICER1 syndrome. The mouse model developed bile duct hyperplasia with fibrosis, similar to congenital hepatic fibrosis, as well as cystic liver tumors resembling those in Caroli's syndrome, intrahepatic cholangiocarcinoma, and hepatocellular carcinoma. Interestingly, the mouse model of DICER1 syndrome showed abnormal formation of primary cilia in the bile duct epithelium, which is a known cause of bile duct hyperplasia and cyst formation. These results indicated that DICER1 mutations contribute to cystic liver tumors by inducing defective primary cilia. The mouse model generated in this study will be useful for elucidating the potential mechanisms of tumorigenesis induced by DICER1 variants and for obtaining a comprehensive understanding of DICER1 syndrome. © 2024 The Pathological Society of Great Britain and Ireland.

Pleuropulmonary blastoma (PPB) with central nervous system metastasis: case report, imaging findings, and review of literature.

Pleuropulmonary blastoma (PPB) is a very rare tumor of the chest seen predominantly in young children with great heterogeneity and clinical, biochemical, and biological complexity and recognized, described, and classified as distinct from the pulmonary blastoma typically encountered in adults. Unfortunately, it has a poor and dismal prognosis and is mainly classified as cystic (type 1), mixed type (type 2), and solid (type 3). Herein, we present one case of PPB type 2 presenting clinically with a right pulmonary abscess, a rare clinical presentation of PPB, which was initially treated with surgery, and after approximately 1 year of follow-up, pulmonary rest-recurrence and central nervous system secondary deposits were detected. When a large pleural-based mass is identified in a young child, PPB should also be considered, especially in a patient with a positive oncological family history. Suggestive findings include the absence of chest wall invasion, presence of pleural fluid, right-sided location, and heterogeneous native (NECT) low attenuation with variable postcontrast enhancement. The authors believe that a modern therapeutic approach should consider these results for a better understanding of the genetic nature and complex mechanism and process of PPB disease development (both clinical and preclinical data concerning PPB pathophysiology are still lacking and are not completely understood) so that it would be possible to establish new possible therapeutic options (i.e. nuclear medicine theranostics in PPB treatment, developments and innovation in FLASH radiotherapy and proton therapy) and approaches, and so that, given the severity of the disease, it would be possible to indicate the importance of genetic testing and counseling of close relatives. In line with the previous, the rapid development of artificial intelligence could potentially bring the development of a novel fusion of radio mics and semantic features and MRI-based machine learning in distinguishing PPB from similar pathology.

Predictive Factors of Mortality in Pediatric Pleuropulmonary Blastoma: A Surveillance Epidemiology and End Results Database Analysis.

Purpose: Pleuropulmonary blastoma (PPB) is a rare malignancy associated with mutations in the DICER1 gene. Early-stage disease (PPB type I) mimics cystic lung malformations and develops in infants <1 year of age, and PPB type II and III arises in older children. The objective of this study was to analyze predictive factors of mortality in pediatric patients aged 0-19 years diagnosed with PPB between 2000 and 2019 in the USA. Methods: A retrospective analysis of pediatric patients (0-19 years) in the Surveillance Epidemiology and End Results database was conducted from 2000 to 2019 with a diagnosis of PPB using International Classification of Disease for Oncology, third edition code 8973/3 and rare tumor code 45. Demographics, incidence, staging, treatment, and mortality were extracted. A mortality risk predictive equation was developed using logistic regression. Statistical analysis was conducted through Microsoft Excel Analysis ToolPak and Solver. Results: There were a total of 71 new cases of PPB during the study period, with 16 (22%) deaths. The demographic analysis demonstrated that 40/71 (56.3%) patients were female, 57/71 (80.3%) were White, and 64/71 (90.1%) resided in metropolitan areas. Regression analysis demonstrated a statistically significant correlation between mortality and stage (P = 0.029), need for chemotherapy (P = 0.047), and female sex (P = 0.019). There was no significant correlation between mortality and need for radiation, race, or age at diagnosis. Multiple logistic regression analysis generated a predictive equation of mortality dependent on the stage of PPB, need for chemotherapy, and sex. This equation has an 82% accuracy, 81% sensitivity, and an 18% false positive rate. Conclusion: PPB is a rare disease. Distinguishing PPB from benign cystic lung malformations in infancy is important to avoid progression to Type II and III PPB. Advanced stages of PPB have a greater need for systemic chemotherapy and radiation with a poor prognosis.

Type II pleuropulmonary blastoma in a fetus: case report of a rare mesenchymal mediastinal tumor and literature review.

Mediastinal tumors are exceedingly rare during fetal development, presenting significant diagnostic challenges and potentially leading to severe outcomes such as stillbirth or metastatic disease if not promptly identified and managed. Pleuropulmonary blastomas are primitive mesenchymal tumors often linked to mutations in the DICER1 gene, indicating a hereditary pattern associated with other common adult neoplasms with dominant inheritance. This report describes a case involving a 20-year-old Caucasian woman whose pregnancy was complicated by a stillbirth in the second trimester. Initial suspicions of a mediastinal tumor arose from blood tests and ultrasound examinations during pregnancy surveillance. However, the definitive diagnosis of a type II pleuropulmonary blastoma was established through a pathological examination at autopsy. This case underscores the complexities of diagnosing fetal mediastinal tumors and contributes to the sparse literature on neonatal pleuropulmonary blastomas. Our comprehensive review of the differential diagnoses and literature emphasizes the unique characteristics of pleuropulmonary blastoma and its similarities to other soft tissue sarcomas, enhancing understanding of their clinical and genetic profiles.

microRNA-8514-5p regulates adipokinetic hormone/corazonin-related peptide receptor to affect development and reproduction of Plutella xylostella.

BACKGROUND: Dicer1 plays a crucial role in regulating the development and reproduction of insects. Knockout of Dicer1 causes pupal deformity, low eclosion and low fecundity in Plutella xylostella, but the mechanism behind this phenomenon is not clear. This study aims to identify differentially-expressed genes and miRNAs in the Dicer1-knockout strain (ΔPxDcr-1) and assess their impact on the reproduction and development of P. xylostella. RESULTS: The knockout of Dicer1 affected the expression of genes including the adipokinetic hormone/corazonin-related peptide receptor (PxACPR). The expression of PxACPR was upregulated, and the expression of miR-8514-5p was downregulated in ΔPxDcr-1 of P. xylostella. The dual luciferase reporter assay and pull-down assay showed that miR-8514-5p bound to PxACPR in vitro and in vivo. The expression profiles demonstrated a negative correlation between PxACPR mRNA and miR-8514-5p in different developmental stages of the wild-type strain. Both the miR-8514-5p agomir and double-stranded RNA of ACPR (dsPxACPR) injected into the pre-pupae inhibited the mRNA level of PxACPR, causing high mortality and deformity of pupae, and low fecundity and hatching rate, which were consistent with the phenotype of ΔPxDcr-1. The injection of miR-8514-5p antagomir caused a similar phenotype to the injection of miR-8514-5p agomir. Additionally, the injection of miR-8514-5p antagomir significantly rescued the phenotype caused by dsPxACPR. CONCLUSION: These results indicate that miR-8514-5p affects the development and reproduction of P. xylostella by regulating PxACPR, and the homeostasis of PxACPR expression is essential for the development and reproduction of P. xylostella. © 2024 Society of Chemical Industry.

Argonaute-independent, Dicer-dependent antiviral defense against RNA viruses.

Antiviral RNA interference (RNAi) is conserved from yeasts to mammals. Dicer recognizes and cleaves virus-derived double-stranded RNA (dsRNA) and/or structured single-stranded RNA (ssRNA) into small-interfering RNAs, which guide effector Argonaute to homologous viral RNAs for digestion and inhibit virus replication. Thus, Argonaute is believed to be essential for antiviral RNAi. Here, we show Argonaute-independent, Dicer-dependent antiviral defense against dsRNA viruses using Cryphonectria parasitica (chestnut blight fungus), which is a model filamentous ascomycetous fungus and hosts a variety of viruses. The fungus has two dicer-like genes (dcl1 and dcl2) and four argonaute-like genes (agl1 to agl4). We prepared a suite of single to quadruple agl knockout mutants with or without dcl disruption. We tested these mutants for antiviral activities against diverse dsRNA viruses and ssRNA viruses. Although both DCL2 and AGL2 worked as antiviral players against some RNA viruses, DCL2 without argonaute was sufficient to block the replication of other RNA viruses. Overall, these results indicate the existence of a Dicer-alone defense and different degrees of susceptibility to it among RNA viruses. We discuss what determines the great difference in susceptibility to the Dicer-only defense.

Oocyte Maturation and miRNAs: Studying a Complicate Interaction to Reveal Possible Biomarkers for Female Infertility.

Cellular metabolism, apoptosis, fertilization, and proliferation of granulosa cells belong to a battery of processes where microRNAs can be detected and associated with infertility. The aim of the present review is to focus on mammalian oocyte maturation events and the association between oocyte growth and miRNA expression. PubMed/Medline, Google Scholar and Scopus databases were searched, and 33 studies were included. Regarding the correlation among miRNA expression and the regulation of granulosa cells and cumulus cells, the most important miRNAs were let-7b, let-7c and miR-21. Additionally, the loss of Dicer, an enzyme involved in miRNA biogenesis, is probably a crucial factor in oogenesis, oocyte maturation and embryogenesis. Furthermore, miRNAs interfere with different cellular mechanisms like apoptosis, steroidogenesis, genome integrity, angiogenesis, antioxidative response and, consequently, oocyte maturation. Hence, it is of major importance to clarify the role and mechanism of each miRNA as understanding its action may develop new tools and establish new diagnostic and treatment approaches for infertility and ovarian disorders.

Functional canonical RNAi in mice expressing a truncated Dicer isoform and long dsRNA.

Canonical RNA interference (RNAi) is sequence-specific mRNA degradation guided by small interfering RNAs (siRNAs) made by RNase III Dicer from long double-stranded RNA (dsRNA). RNAi roles include gene regulation, antiviral immunity or defense against transposable elements. In mammals, RNAi is constrained by Dicer's adaptation to produce another small RNA class-microRNAs. However, a truncated Dicer isoform (ΔHEL1) supporting RNAi exists in mouse oocytes. A homozygous mutation to express only the truncated ΔHEL1 variant causes dysregulation of microRNAs and perinatal lethality in mice. Here, we report the phenotype and canonical RNAi activity in DicerΔHEL1/wt mice, which are viable, show minimal miRNome changes, but their endogenous siRNA levels are an order of magnitude higher. We show that siRNA production in vivo is limited by available dsRNA, but not by Protein kinase R, a dsRNA sensor of innate immunity. dsRNA expression from a transgene yields sufficient siRNA levels to induce efficient RNAi in heart and muscle. DicerΔHEL1/wt mice with enhanced canonical RNAi offer a platform for examining potential and limits of mammalian RNAi in vivo.

The Evolution and Characterization of the RNA Interference Pathways in Lophotrochozoa.

In animals, three main RNA interference mechanisms have been described so far, which respectively maturate three types of small noncoding RNAs (sncRNAs): miRNAs, piRNAs, and endo-siRNAs. The diversification of these mechanisms is deeply linked with the evolution of the Argonaute gene superfamily since each type of sncRNA is typically loaded by a specific Argonaute homolog. Moreover, other protein families play pivotal roles in the maturation of sncRNAs, like the DICER ribonuclease family, whose DICER1 and DICER2 paralogs maturate respectively miRNAs and endo-siRNAs. Within Metazoa, the distribution of these families has been only studied in major groups, and there are very few data for clades like Lophotrochozoa. Thus, we here inferred the evolutionary history of the animal Argonaute and DICER families including 43 lophotrochozoan species. Phylogenetic analyses along with newly sequenced sncRNA libraries suggested that in all Trochozoa, the proteins related to the endo-siRNA pathway have been lost, a part of them in some phyla (i.e. Nemertea, Bryozoa, Entoprocta), while all of them in all the others. On the contrary, early diverging phyla, Platyhelminthes and Syndermata, showed a complete endo-siRNA pathway. On the other hand, miRNAs were revealed the most conserved and ubiquitous mechanism of the metazoan RNA interference machinery, confirming their pivotal role in animal cell regulation.

The RNAi Machinery in the Fungus Fusarium fujikuroi Is Not Very Active in Synthetic Medium and Is Related to Transposable Elements.

Small RNAS (sRNAs) participate in regulatory RNA interference (RNAi) mechanisms in a wide range of eukaryotic organisms, including fungi. The fungus Fusarium fujikuroi, a model for the study of secondary metabolism, contains a complete set of genes for RNAi pathways. We have analyzed by high-throughput sequencing the content of sRNAs in total RNA samples of F. fujikuroi grown in synthetic medium in the dark or after 1 h of illumination, using libraries below 150 nt, covering sRNAs and their precursors. For comparison, a parallel analysis with Fusarium oxysporum was carried out. The sRNA reads showed a higher proportion of 5' uracil in the RNA samples of the expected sizes in both species, indicating the occurrence of genuine sRNAs, and putative miRNA-like sRNAs (milRNAS) were identified with prediction software. F. fujikuroi carries at least one transcriptionally expressed Ty1/copia-like retrotransposable element, in which sRNAs were found in both sense and antisense DNA strands, while in F. oxysporum skippy-like elements also show sRNA formation. The finding of sRNA in these mobile elements indicates an active sRNA-based RNAi pathway. Targeted deletion of dcl2, the only F. fujikuroi Dicer gene with significant expression under the conditions tested, did not produce appreciable phenotypic or transcriptomic alterations.