Frank Schnorrer

Bio: Frank Schnorrer is an academic researcher from Aix-Marseille University. The author has contributed to research in topics: Sarcomere & Myofibril. The author has an hindex of 30, co-authored 59 publications receiving 5335 citations. Previous affiliations of Frank Schnorrer include Research Institute of Molecular Pathology & Institute of Molecular Biotechnology.

A genome-wide transgenic RNAi library for conditional gene inactivation in Drosophila

Abstract: Forward genetic screens in model organisms have provided important insights into numerous aspects of development, physiology and pathology. With the availability of complete genome sequences and the introduction of RNA-mediated gene interference (RNAi), systematic reverse genetic screens are now also possible. Until now, such genome-wide RNAi screens have mostly been restricted to cultured cells and ubiquitous gene inactivation in Caenorhabditis elegans. This powerful approach has not yet been applied in a tissue-specific manner. Here we report the generation and validation of a genome-wide library of Drosophila melanogaster RNAi transgenes, enabling the conditional inactivation of gene function in specific tissues of the intact organism. Our RNAi transgenes consist of short gene fragments cloned as inverted repeats and expressed using the binary GAL4/UAS system. We generated 22,270 transgenic lines, covering 88% of the predicted protein-coding genes in the Drosophila genome. Molecular and phenotypic assays indicate that the majority of these transgenes are functional. Our transgenic RNAi library thus opens up the prospect of systematically analysing gene functions in any tissue and at any stage of the Drosophila lifespan.

A genome-wide resource for the analysis of protein localisation in Drosophila.

Abstract: The Drosophila genome contains >13000 protein-coding genes, the majority of which remain poorly investigated. Important reasons include the lack of antibodies or reporter constructs to visualise these proteins. Here, we present a genome-wide fosmid library of 10000 GFP-tagged clones, comprising tagged genes and most of their regulatory information. For 880 tagged proteins, we created transgenic lines, and for a total of 207 lines, we assessed protein expression and localisation in ovaries, embryos, pupae or adults by stainings and live imaging approaches. Importantly, we visualised many proteins at endogenous expression levels and found a large fraction of them localising to subcellular compartments. By applying genetic complementation tests, we estimate that about two-thirds of the tagged proteins are functional. Moreover, these tagged proteins enable interaction proteomics from developing pupae and adult flies. Taken together, this resource will boost systematic analysis of protein expression and localisation in various cellular and developmental contexts.

Systematic genetic analysis of muscle morphogenesis and function in Drosophila

Abstract: A genome-wide RNAi screen has been used to define the genetic basis for the formation and function of Drosophila muscle. A total of 2,785 genes were recognized as having a role in muscle, many of then assigned to specific functions in the organization of muscles, myofibrils or sarcomeres. Many of these genes are phylogenetically conserved, and the list of conserved and functionally validated muscle genes includes a strong enrichment for genes implicated in mammalian sarcomere organization and human muscle diseases. A genome-wide RNA interference screen to systematically test the genetic basis for formation and function of the Drosophila muscle is described. A role in muscle for 2,785 genes is identified; many of these genes are phylogenetically conserved. Systematic genetic approaches have provided deep insight into the molecular and cellular mechanisms that operate in simple unicellular organisms. For multicellular organisms, however, the pleiotropy of gene function has largely restricted such approaches to the study of early embryogenesis. With the availability of genome-wide transgenic RNA interference (RNAi) libraries in Drosophila1,2, it is now possible to perform a systematic genetic dissection of any cell or tissue type at any stage of the lifespan. Here we apply these methods to define the genetic basis for formation and function of the Drosophila muscle. We identify a role in muscle for 2,785 genes, many of which we assign to specific functions in the organization of muscles, myofibrils or sarcomeres. Many of these genes are phylogenetically conserved, including genes implicated in mammalian sarcomere organization and human muscle diseases.

The molecular motor dynein is involved in targeting swallow and bicoid RNA to the anterior pole of Drosophila oocytes.

Abstract: Localization of bicoid (bcd) messenger RNA to the anterior pole of the Drosophila oocyte requires the exuperantia ( exu), swallow (swa) and staufen (stau) genes. We show here that Swa protein transiently co-localizes with bcd RNA in mid-oogenesis. Swa also localizes to the anterior pole of the oocyte in the absence of bcd RNA. This localization does not require Exu, but depends on intact microtubules. In mutant ovaries with duplicated polarity of microtubules, Swa and bcd RNA are ectopically localized at the posterior pole, as well as being present at the anterior pole. We identify dynein light chain-1 (Ddlc-1), a component of the minus-end-directed microtubule motor cytoplasmic dynein, as a Swa-binding protein. We propose that Swa acts as an adaptor for the dynein complex and thereby enables dynein to transport bcd RNA along microtubules to their minus ends at the anterior pole of the oocyte.

Fly Cell Atlas: A single-nucleus transcriptomic atlas of the adult fruit fly

Abstract: For more than 100 years, the fruit fly Drosophila melanogaster has been one of the most studied model organisms. Here, we present a single-cell atlas of the adult fly, Tabula Drosophilae, that includes 580,000 nuclei from 15 individually dissected sexed tissues as well as the entire head and body, annotated to >250 distinct cell types. We provide an in-depth analysis of cell type–related gene signatures and transcription factor markers, as well as sexual dimorphism, across the whole animal. Analysis of common cell types between tissues, such as blood and muscle cells, reveals rare cell types and tissue-specific subtypes. This atlas provides a valuable resource for the Drosophila community and serves as a reference to study genetic perturbations and disease models at single-cell resolution. Description Cell type diversity in a whole fly The fruit fly Drosophila melanogaster has served as a premier model organism for discovering fundamental and evolutionarily conserved biological mechanisms. Combining recent advances in single-cell sequencing with powerful fly genetic tools holds great promise for making further discoveries. Li et al. present a single-cell atlas of the entire adult fly that includes 580,000 cells and more than 250 annotated cell types. Cells from the head and body recapitulated cell types from 15 dissected tissues. In-depth analyses revealed rare cell types, cell-type-specific gene signatures, and sexual dimorphism. This atlas provides a resource for the Drosophila community to study genetic perturbations and diseases at single-cell resolution. —BAP A single-nucleus transcriptomic map reveals more than 250 distinct cell types in the entire adult Drosophila melanogaster. INTRODUCTION Drosophila melanogaster has had a fruitful history in biological research because it has contributed to many key discoveries in genetics, development, and neurobiology. The fruit fly genome contains ~14,000 protein-coding genes, ~63% of which have human orthologs. Single-cell RNA-sequencing has recently been applied to multiple Drosophila tissues and developmental stages. However, these data have been generated by different laboratories on different genetic backgrounds with different dissociation protocols and sequencing platforms, which has hindered the systematic comparison of gene expression across cells and tissues. RATIONALE We aimed to establish a cell atlas for the entire adult Drosophila with the same genetic background, dissociation protocol, and sequencing platform to (i) obtain a comprehensive categorization of cell types, (ii) integrate single-cell transcriptome data with existing knowledge about gene expression and cell types, (iii) systematically compare gene expression across the entire organism and between males and females, and (iv) identify cell type–specific markers across the entire organism. We chose single-nucleus RNA-sequencing (snRNA-seq) to circumvent the difficulties of dissociating cells that are embedded in the cuticle (e.g., sensory neurons) or that are multinucleated (e.g., muscle cells). We took two complementary strategies: sequencing nuclei from dissected tissues to know the identity of the tissue source and sequencing nuclei from the entire head and body to ensure that all cells are sampled. Experts from 40 laboratories participated in crowd annotation to assign transcriptomic cell types with the best knowledge available. RESULTS We sequenced 570,000 cells using droplet-based 10x Genomics from 15 dissected tissues as well as whole heads and bodies, separately in females and males. We also sequenced 10,000 cells from dissected tissues using the plate-based Smart-seq2 platform, providing deeper coverage per cell. We developed reproducible analysis pipelines using NextFlow and implemented a distributed cell-type annotation system with controlled vocabularies in SCope. Crowd-based annotations of transcriptomes from dissected tissues identified 17 main cell categories and 251 detailed cell types linked to FlyBase ontologies. Many of these cell types are characterized for the first time, either because they emerged only after increasing cell coverage or because they reside in tissues that had not been previously subjected to scRNA-seq. The excellent correspondence of transcriptomic clusters from whole body and dissected tissues allowed us to transfer annotations and identify a few cuticular cell types not detected in individual tissues. Cross-tissue analysis revealed location-specific subdivisions of muscle cells and heterogeneity within blood cells. We then determined cell type–specific marker genes and transcription factors with different specificity levels, enabling the construction of gene regulatory networks. Finally, we explored sexual dimorphism, finding a link between sex-biased expression and the presence of doublesex, and investigated tissue dynamics through trajectory analyses. CONCLUSION Our Fly Cell Atlas (FCA) constitutes a valuable resource for the Drosophila community as a reference for studies of gene function at single-cell resolution. All the FCA data are freely available for further analysis through multiple portals and can be downloaded for custom analyses using other single-cell tools. The ability to annotate cell types by sequencing the entire head and body will facilitate the use of Drosophila in the study of biological processes and in modeling human diseases at a whole-organism level with cell-type resolution. All data with annotations can be accessed from www.flycellatlas.org, which provides links to SCope, ASAP, and cellxgene portals. Tabula Drosophilae. In this single-cell atlas of the adult fruit fly, 580,000 cells were sequenced and >250 cell types were annotated. They are from 15 individually dissected sexed tissues as well as the entire head and body. All data are freely available for visualization and download, with featured analyses shown at the bottom right.

Toll-like receptor signalling

Abstract: One of the mechanisms by which the innate immune system senses the invasion of pathogenic microorganisms is through the Toll-like receptors (TLRs), which recognize specific molecular patterns that are present in microbial components. Stimulation of different TLRs induces distinct patterns of gene expression, which not only leads to the activation of innate immunity but also instructs the development of antigen-specific acquired immunity. Here, we review the rapid progress that has recently improved our understanding of the molecular mechanisms that mediate TLR signalling.

Microtubule nucleation by γ-tubulin complexes

Abstract: Microtubule nucleation is regulated by the γ-tubulin ring complex (γTuRC) and related γ-tubulin complexes, providing spatial and temporal control over the initiation of microtubule growth. Recent structural work has shed light on the mechanism of γTuRC-based microtubule nucleation, confirming the long-standing hypothesis that the γTuRC functions as a microtubule template. The first crystallographic analysis of a non-γ-tubulin γTuRC component (γ-tubulin complex protein 4 (GCP4)) has resulted in a new appreciation of the relationships among all γTuRC proteins, leading to a refined model of their organization and function. The structures have also suggested an unexpected mechanism for regulating γTuRC activity via conformational modulation of the complex component GCP3. New experiments on γTuRC localization extend these insights, suggesting a direct link between its attachment at specific cellular sites and its activation.

Rapid SNP Discovery and Genetic Mapping Using Sequenced RAD Markers

Abstract: Single nucleotide polymorphism (SNP) discovery and genotyping are essential to genetic mapping. There remains a need for a simple, inexpensive platform that allows high-density SNP discovery and genotyping in large populations. Here we describe the sequencing of restriction-site associated DNA (RAD) tags, which identified more than 13,000 SNPs, and mapped three traits in two model organisms, using less than half the capacity of one Illumina sequencing run. We demonstrated that different marker densities can be attained by choice of restriction enzyme. Furthermore, we developed a barcoding system for sample multiplexing and fine mapped the genetic basis of lateral plate armor loss in threespine stickleback by identifying recombinant breakpoints in F2 individuals. Barcoding also facilitated mapping of a second trait, a reduction of pelvic structure, by in silico re-sorting of individuals. To further demonstrate the ease of the RAD sequencing approach we identified polymorphic markers and mapped an induced mutation in Neurospora crassa. Sequencing of RAD markers is an integrated platform for SNP discovery and genotyping. This approach should be widely applicable to genetic mapping in a variety of organisms.

Many roads to maturity: microRNA biogenesis pathways and their regulation

Abstract: MicroRNAs are important regulators of gene expression that control both physiological and pathological processes such as development and cancer. Although their mode of action has attracted great attention, the principles governing their expression and activity are only beginning to emerge. Recent studies have introduced a paradigm shift in our understanding of the microRNA biogenesis pathway, which was previously believed to be universal to all microRNAs. Maturation steps specific to individual microRNAs have been uncovered, and these offer a plethora of regulatory options after transcription with multiple proteins affecting microRNA processing efficiency. Here we review the recent advances in knowledge of the microRNA biosynthesis pathways and discuss their impact on post-transcriptional microRNA regulation during tumour development.