MiR-202 controls female fecundity by regulating medaka oogenesis.

Female gamete production relies on coordinated molecular and cellular processes that occur in the ovary throughout oogenesis. In fish, as in other vertebrates, these processes have been extensively studied both in terms of endocrine/paracrine regulation and protein expression and activity. The role of small non-coding RNAs in the regulation of animal reproduction remains however largely unknown and poorly investigated, despite a growing interest for the importance of miRNAs in a wide variety of biological processes. Here, we analyzed the role of miR-202, a miRNA predominantly expressed in male and female gonads in several vertebrate species. We studied its expression in the medaka ovary and generated a mutant line (using CRISPR/Cas9 genome editing) to determine its importance for reproductive success with special interest for egg production. Our results show that miR-202-5p is the most abundant mature form of the miRNA and that it is expressed in granulosa cells and in the unfertilized egg. The knock out (KO) of mir-202 gene resulted in a strong phenotype both in terms of number and quality of eggs produced. Mutant females exhibited either no egg production or produced a dramatically reduced number of eggs that could not be fertilized, ultimately leading to no reproductive success. We quantified the size distribution of the oocytes in the ovary of KO females and performed a large-scale transcriptomic analysis approach to identified dysregulated molecular pathways. Together, cellular and molecular analyses indicate that the lack of miR-202 impairs the early steps of oogenesis/folliculogenesis and decreases the number of large (i.e. vitellogenic) follicles, ultimately leading to dramatically reduced female fecundity. This study sheds new light on the regulatory mechanisms that control the early steps of follicular development, including possible targets of miR-202-5p, and provides the first in vivo functional evidence that a gonad-predominant microRNA may have a major role in female reproduction.

Sequencing DNA with nanopores: Troubles and biases.

Oxford Nanopore Technologies' (ONT) long read sequencers offer access to longer DNA fragments than previous sequencer generations, at the cost of a higher error rate. While many papers have studied read correction methods, few have addressed the detailed characterization of observed errors, a task complicated by frequent changes in chemistry and software in ONT technology. The MinION sequencer is now more stable and this paper proposes an up-to-date view of its error landscape, using the most mature flowcell and basecaller. We studied Nanopore sequencing error biases on both bacterial and human DNA reads. We found that, although Nanopore sequencing is expected not to suffer from GC bias, it is a crucial parameter with respect to errors. In particular, low-GC reads have fewer errors than high-GC reads (about 6% and 8% respectively). The error profile for homopolymeric regions or regions with short repeats, the source of about half of all sequencing errors, also depends on the GC rate and mainly shows deletions, although there are some reads with long insertions. Another interesting finding is that the quality measure, although over-estimated, offers valuable information to predict the error rate as well as the abundance of reads. We supplemented this study with an analysis of a rapeseed RNA read set and shown a higher level of errors with a higher level of deletion in these data. Finally, we have implemented an open source pipeline for long-term monitoring of the error profile, which enables users to easily compute various analysis presented in this work, including for future developments of the sequencing device. Overall, we hope this work will provide a basis for the design of better error-correction methods.