L(3)mbt and the LINT complex safeguard cellular identity in the Drosophila ovary.

Maintenance of cellular identity is essential for tissue development and homeostasis. At the molecular level, cell identity is determined by the coordinated activation and repression of defined sets of genes. The tumor suppressor L(3)mbt has been shown to secure cellular identity in Drosophila larval brains by repressing germline-specific genes. Here, we interrogate the temporal and spatial requirements for L(3)mbt in the Drosophila ovary, and show that it safeguards the integrity of both somatic and germline tissues. l(3)mbt mutant ovaries exhibit multiple developmental defects, which we find to be largely caused by the inappropriate expression of a single gene, nanos, a key regulator of germline fate, in the somatic ovarian cells. In the female germline, we find that L(3)mbt represses testis-specific and neuronal genes. At the molecular level, we show that L(3)mbt function in the ovary is mediated through its co-factor Lint-1 but independently of the dREAM complex. Together, our work uncovers a more complex role for L(3)mbt than previously understood and demonstrates that L(3)mbt secures tissue identity by preventing the simultaneous expression of original identity markers and tissue-specific misexpression signatures

Temporal and Spatial Control of Germ-Plasm RNAs

SummaryIn many species, germ cells form in a specialized germ plasm, which contains localized maternal RNAs [1–5]. In the absence of active transcription in early germ cells, these maternal RNAs encode germ-cell components with critical functions in germ-cell specification, migration, and development [6, 7]. For several RNAs, localization has been correlated with release from translational repression, suggesting an important regulatory function linked to localization [3, 4, 8, 9]. To address the role of RNA localization and translational control more systematically, we assembled a comprehensive set of RNAs that are localized to polar granules, the characteristic germ-plasm organelles. We find that the 3′-untranslated regions (UTRs) of all RNAs tested control RNA localization and instruct distinct temporal patterns of translation of the localized RNAs. We demonstrate necessity for translational timing by swapping the 3′UTR of polar granule component (pgc), which controls translation in germ cells, with that of nanos, which is translated earlier. Translational activation of pgc is concurrent with extension of its poly(A) tail length but appears largely independent of the Drosophila CPEB homolog ORB. Our results demonstrate a role for 3′UTR mediated translational regulation in fine-tuning the temporal expression of localized RNA, and this may provide a paradigm for other RNAs that are found enriched at distinct cellular locations such as the leading edge of fibroblasts or the neuronal synapse

Chromatin accessibility and transcription factor binding through the perspective of mitosis

International audienceChromatin accessibility is generally perceived as a common property of active regulatory elements where transcription factors are recruited via DNA-specific interactions and other physico-chemical properties to regulate gene transcription. Recent work in the context of mitosis provides less trivial and potentially more interesting relationships than previously anticipated

piRNA-mediated regulation of transposon alternative splicing in the soma and germ line.

Transposable elements can drive genome evolution, but their enhanced activity is detrimental to the host and therefore must be tightly regulated. The Piwi-interacting small RNA (piRNA) pathway is vital for the regulation of transposable elements, by inducing transcriptional silencing or post-transcriptional decay of mRNAs. Here we show that piRNAs and piRNA biogenesis components regulate precursor mRNA splicing of P-transposable element transcripts in vivo, leading to the production of the non-transposase-encoding mature mRNA isoform in Drosophila germ cells. Unexpectedly, we show that the piRNA pathway components do not act to reduce transcript levels of the P-element transposon during P-M hybrid dysgenesis, a syndrome that affects germline development in Drosophila. Instead, splicing regulation is mechanistically achieved together with piRNA-mediated changes to repressive chromatin states, and relies on the function of the Piwi-piRNA complex proteins Asterix (also known as Gtsf1) and Panoramix (Silencio), as well as Heterochromatin protein 1a (HP1a; encoded by Su(var)205). Furthermore, we show that this machinery, together with the piRNA Flamenco cluster, not only controls the accumulation of Gypsy retrotransposon transcripts but also regulates the splicing of Gypsy mRNAs in cultured ovarian somatic cells, a process required for the production of infectious particles that can lead to heritable transposition events. Our findings identify splicing regulation as a new role and essential function for the Piwi pathway in protecting the genome against transposon mobility, and provide a model system for studying the role of chromatin structure in modulating alternative splicing during development

Molecular and epistatic interactions between pioneer transcription factors shape nucleosome dynamics and cell differentiation

Posted May 27, 2024 on bioRxiv.Pioneer transcription factors (TF) bind nucleosome-embedded DNA motifs to activate new regulatory elements and promote differentiation. However, the complexity, binding dependencies and temporal effects of their action remain unclear. Here, we dissect how the pioneer TF GATA6 triggers Primitive Endoderm (PrE) differentiation from pluripotent cells. We show that transient GATA6 binding exploits accessible regions to decommission active enhancers and promote pluripotency gene silencing. Simultaneously, GATA6 targets closed chromatin and initiates an extensive remodeling culminating in the establishment of fragile nucleosomes flanked by ordered nucleosome arrays and increased accessibility. This is directly enhanced by rapidly expressed PrE TFs (SOX17) and by pluripotency TFs repurposed for differentiation (OCT4/SOX2). Furthermore, GATA6 mediates the replacement of essential nuclear receptors for PrE differentiation, from ESRRB to ESRRA. Therefore, pioneer TFs orchestrate a complex gene regulatory network involving many if not all available pioneer TFs, including those required to support the original identity of differentiating cells

Nr5a2 is essential for morula development

Posted January 18, 2023 on bioRxiv.International audienceEarly embryogenesis is driven by transcription factors (TFs) that first activate the zygotic genome and then specify the lineages constituting the blastocyst. While the TFs specifying the blastocyst’s lineages are well characterised, those playing earlier roles are ill-defined. Using mouse models of the TF Nr5a2 , we show that Nr5a2 -/- embryos arrest at the early morula stage and exhibit overt phenotypical problems such as altered lineage specification, frequent mitotic failure and substantial chromosome segregation defects. Transcriptomic profiling shows that NR5A2 is a master regulator required for appropriate expression of thousands of genes at the 8-cells stage, including lineage-specifying TFs and genes involved in mitosis, telomere maintenance and DNA repair. We conclude that NR5A2 coordinates proliferation, genome stability and lineage specification to ensure proper morula development

Mitotic bookmarking redundancy by nuclear receptors in pluripotent cells

International audienceMitotic bookmarking transcription factors (TFs) are thought to mediate rapid and accurate reactivation after mitotic gene silencing. However, the loss of individual bookmarking TFs often leads to the deregulation of only a small proportion of their mitotic targets, raising doubts on the biological significance and importance of their bookmarking function. Here we used targeted proteomics of the mitotic bookmarking TF ESRRB, an orphan nuclear receptor, to discover a large redundancy in mitotic binding among members of the protein super-family of nuclear receptors. Focusing on the nuclear receptor NR5A2, which together with ESRRB is essential in maintaining pluripotency in mouse embryonic stem cells, we demonstrate conjoint bookmarking activity of both factors on promoters and enhancers of a large fraction of active genes, particularly those most efficiently reactivated in G1. Upon fast and simultaneous degradation of both factors during mitotic exit, hundreds of mitotic targets of ESRRB/NR5A2, including key players of the pluripotency network, display attenuated transcriptional reactivation. We propose that redundancy in mitotic bookmarking TFs, especially nuclear receptors, confers robustness to the reestablishment of gene regulatory networks after mitosis