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Methylation of histone H3K36 in higher eukaryotes is mediated by multiple methyltransferases. Set2-related H3K36 methyltransferases are targeted to genes by association with RNA Polymerase II and are involved in preventing aberrant transcription initiation within the body of genes. The targeting and roles of the NSD family of mammalian H3K36 methyltransferases, known to be involved in human developmental disorders and oncogenesis, are not known. We used genome-wide chromatin immunoprecipitation (ChIP) to investigate the targeting and roles of the Caenorhabditis elegans NSD homolog MES-4, which is maternally provided to progeny and is required for the survival of nascent germ cells. ChIP analysis in early C. elegans embryos revealed that, consistent with immunostaining results, MES-4 binding sites are concentrated on the autosomes and the leftmost approximately 2% (300 kb) of the X chromosome. MES-4 overlies the coding regions of approximately 5,000 genes, with a modest elevation in the 5' regions of gene bodies. Although MES-4 is generally found over Pol II-bound genes, analysis of gene sets with different temporal-spatial patterns of expression revealed that Pol II association with genes is neither necessary nor sufficient to recruit MES-4. In early embryos, MES-4 associates with genes that were previously expressed in the maternal germ line, an interaction that does not require continued association of Pol II with those loci. Conversely, Pol II association with genes newly expressed in embryos does not lead to recruitment of MES-4 to those genes. These and other findings suggest that MES-4, and perhaps the related mammalian NSD proteins, provide an epigenetic function for H3K36 methylation that is novel and likely to be unrelated to ongoing transcription. We propose that MES-4 transmits the memory of gene expression in the parental germ line to offspring and that this memory role is critical for the PGCs to execute a proper germline program.

The p53 tumor suppressor promotes apoptosis in response to DNA damage. Here we describe the Caenorhabditis elegans gene ced-13 , which encodes a conserved BH3-only protein. We show that ced-13 mRNA accumulates following DNA damage and that this accumulation is dependent on an intact C . elegans cep-1 / p53 gene. We demonstrate that CED-13 protein physically interacts with the antiapoptotic Bcl-2-related protein CED-9. Furthermore, overexpression of ced-13 in somatic cells leads to the death of cells that normally survive and this death requires the core apoptotic pathway of C. elegans . Recent studies have implicated two BH3-only proteins, Noxa and PUMA, in p53-induced apoptosis in mammals. Our studies suggest that in addition to the BH3-only protein EGL-1, CED-13 might also promote apoptosis in the C. elegans germ line in response to p53 activation. We propose that an evolutionarily conserved pathway exists in which p53 promotes cell death by inducing expression of two BH3-only genes.

Single-cell RNA sequencing provides the power to identify the developmental trajectory of an organism. However, identifying the temporal lineage of cell development can be difficult without large-scale analyses. Packer et al. sequenced more than 80,000 cells from embryos of the roundworm Caenorhabditis elegans to determine the expression of genes directing the development of terminal cell types. Because all somatic cells in a C. elegans individual have been mapped, the authors are able to connect gene expression with cell lineages over time during development, noting stark transitions in some cases. Science , this issue p. eaax1971 Single-cell transcriptomes from Caenorhabditis elegans embryos reconstruct cell lineages and terminal cell differentiation. Caenorhabditis elegans is an animal with few cells but a wide diversity of cell types. In this study, we characterize the molecular basis for their specification by profiling the transcriptomes of 86,024 single embryonic cells. We identify 502 terminal and preterminal cell types, mapping most single-cell transcriptomes to their exact position in C. elegans ’ invariant lineage. Using these annotations, we find that (i) the correlation between a cell’s lineage and its transcriptome increases from middle to late gastrulation, then falls substantially as cells in the nervous system and pharynx adopt their terminal fates; (ii) multilineage priming contributes to the differentiation of sister cells at dozens of lineage branches; and (iii) most distinct lineages that produce the same anatomical cell type converge to a homogenous transcriptomic state.

Mitochondria are inherited maternally in most animals, but the mechanisms of selective paternal mitochondrial elimination (PME) are unknown. While examining fertilization in Caenorhabditis elegans, we observed that paternal mitochondria rapidly lose their inner membrane integrity. CPS-6, a mitochondrial endonuclease G, serves as a paternal mitochondrial factor that is critical for PME. We found that CPS-6 relocates from the intermembrane space of paternal mitochondria to the matrix after fertilization to degrade mitochondrial DNA. It acts with maternal autophagy and proteasome machineries to promote PME. Loss of cps-6 delays breakdown of mitochondrial inner membranes, autophagosome enclosure of paternal mitochondria, and PME. Delayed removal of paternal mitochondria causes increased embryonic lethality, demonstrating that PME is important for normal animal development. Thus, CPS-6 functions as a paternal mitochondrial degradation factor during animal development.

RNA granules are subcellular compartments that are proposed to form by liquid–liquid phase separation (LLPS), a thermodynamic process that partitions molecules between dilute liquid phases and condensed liquid phases. The mechanisms that localize liquid phases in cells, however, are not fully understood. P granules are RNA granules that form in the posterior of Caenorhabditis elegans embryos. Theoretical studies have suggested that spontaneous LLPS of the RNA-binding protein PGL-3 with RNA drives the assembly of P granules. We find that the PGL-3 phase is intrinsically labile and requires a second phase for stabilization in embryos. The second phase is formed by gel-like assemblies of the disordered protein MEG-3 that associate with liquid PGL-3 droplets in the embryo posterior. Co-assembly of gel phases and liquid phases confers local stability and long-range dynamics, both of which contribute to localized assembly of P granules. Our findings suggest that condensation of RNA granules can be regulated spatially by gel-like polymers that stimulate LLPS locally in the cytoplasm.P granules, which are cytoplasmic RNA granules that form via liquid–liquid phase separation in the posterior of Caenorhabditis elegans embryos, require gel and liquid phases for localized assembly and stability

Parental exposure to pathogens can prime offspring immunity in diverse organisms. The mechanisms by which this heritable priming occurs are largely unknown. Here we report that the soil bacteria Pseudomonas vranovensis is a natural pathogen of the nematode Caenorhabditis elegans and that parental exposure of animals to P. vranovensis promotes offspring resistance to infection. Furthermore, we demonstrate a multigenerational enhancement of progeny survival when three consecutive generations of animals are exposed to P. vranovensis . By investigating the mechanisms by which animals heritably adapt to P. vranovensis infection, we found that parental infection by P. vranovensis results in increased expression of the cysteine synthases cysl-1 and cysl-2 and the regulator of hypoxia inducible factor rhy-1 in progeny, and that these three genes are required for adaptation to P. vranovensis . These observations establish a CYSL-1, CYSL-2, and RHY-1 dependent mechanism by which animals heritably adapt to infection. Caenorhabditis elegans exhibits multigenerational adaptation to bacterial infection but the mechanisms remain unclear. Here, the authors show that C. elegans parental exposure to Pseudomonas vranovensis promotes offspring resistance to infection, a process mediated by the cysteine synthases CYSL-1 and CYSL-2.

Neuropil is a fundamental form of tissue organization within the brain 1 , in which densely packed neurons synaptically interconnect into precise circuit architecture 2 , 3 . However, the structural and developmental principles that govern this nanoscale precision remain largely unknown 4 , 5 . Here we use an iterative data coarse-graining algorithm termed ‘diffusion condensation’ 6 to identify nested circuit structures within the Caenorhabditis elegans neuropil, which is known as the nerve ring. We show that the nerve ring neuropil is largely organized into four strata that are composed of related behavioural circuits. The stratified architecture of the neuropil is a geometrical representation of the functional segregation of sensory information and motor outputs, with specific sensory organs and muscle quadrants mapping onto particular neuropil strata. We identify groups of neurons with unique morphologies that integrate information across strata and that create neural structures that cage the strata within the nerve ring. We use high resolution light-sheet microscopy 7 , 8 coupled with lineage-tracing and cell-tracking algorithms 9 , 10 to resolve the developmental sequence and reveal principles of cell position, migration and outgrowth that guide stratified neuropil organization. Our results uncover conserved structural design principles that underlie the architecture and function of the nerve ring neuropil, and reveal a temporal progression of outgrowth—based on pioneer neurons—that guides the hierarchical development of the layered neuropil. Our findings provide a systematic blueprint for using structural and developmental approaches to understand neuropil organization within the brain. The C. elegans neuropil is shown to be organized into four strata composed of related behavioural circuits, and its design principles are linked to the developmental processes that underpin its assembly.

In sexually reproducing organisms, embryos specify germ cells, which ultimately generate sperm and eggs. In Caenorhabditis elegans, the first germ cell is established when RNA and protein-rich P granules localize to the posterior of the one-cell embryo. Localization of P granules and their physical nature remain poorly understood. Here we show that P granules exhibit liquid-like behaviors, including fusion, dripping, and wetting, which we used to estimate their viscosity and surface tension. As with other liquids, P granules rapidly dissolved and condensed. Localization occurred by a biased increase in P granule condensation at the posterior. This process reflects a classic phase transition, in which polarity proteins vary the condensation point across the cell. Such phase transitions may represent a fundamental physicochemical mechanism for structuring the cytoplasm.

Confocal microscopy 1 remains a major workhorse in biomedical optical microscopy owing to its reliability and flexibility in imaging various samples, but suffers from substantial point spread function anisotropy, diffraction-limited resolution, depth-dependent degradation in scattering samples and volumetric bleaching 2 . Here we address these problems, enhancing confocal microscopy performance from the sub-micrometre to millimetre spatial scale and the millisecond to hour temporal scale, improving both lateral and axial resolution more than twofold while simultaneously reducing phototoxicity. We achieve these gains using an integrated, four-pronged approach: (1) developing compact line scanners that enable sensitive, rapid, diffraction-limited imaging over large areas; (2) combining line-scanning with multiview imaging, developing reconstruction algorithms that improve resolution isotropy and recover signal otherwise lost to scattering; (3) adapting techniques from structured illumination microscopy, achieving super-resolution imaging in densely labelled, thick samples; (4) synergizing deep learning with these advances, further improving imaging speed, resolution and duration. We demonstrate these capabilities on more than 20 distinct fixed and live samples, including protein distributions in single cells; nuclei and developing neurons in Caenorhabditis elegans embryos, larvae and adults; myoblasts in imaginal disks of Drosophila wings; and mouse renal, oesophageal, cardiac and brain tissues. A combination of multiview imaging, structured illumination, reconstruction algorithms and deep-learning predictions realizes spatial- and temporal-resolution improvements in fluorescence microscopy to produce super-resolution images from diffraction-limited input images.

The mitochondrial genome is believed to be maternally inherited in many eukaryotes. Sperm-derived paternal mitochondria enter the oocyte cytoplasm upon fertilization and then normally disappear during early embryogenesis. However, the mechanism responsible for this clearance has been unknown. Here, we show that autophagy, which delivers cytosolic components to lysosomes for degradation, is required for the elimination of paternal mitochondria in Caenorhabditis elegans. Immediately after fertilization, sperm-derived components trigger the localized induction of autophagy around sperm mitochondria. Autophagosomes engulf paternal mitochondria, resulting in their lysosomal degradation during early embryogenesis. In autophagy-defective zygotes, paternal mitochondria and their genome remain even in the first larval stage. Thus, fertilization-triggered autophagy is required for selective degradation of paternal mitochondria and thereby maternal inheritance of mitochondrial DNA.