Heterogeneity of transposon expression and activation of the repressive network in human fetal germ cells.

Epigenetic resetting in germ cells during development de-represses transposable elements (TEs). piRNAs protect fetal germ cells by targeted mRNA destruction and deposition of repressive epigenetic marks. Here, we provide the first evidence for an active piRNA pathway and TE repression in germ cells of human fetal testis. We identify pre-pachytene piRNAs with features of secondary amplification that map most abundantly to the long interspersed element type 1 (L1) family of TEs. L1-ORF1p expression is heterogeneous in fetal germ cells, peaks at mid-gestation and declines concomitantly with increases in piRNAs, nuclear localization of HIWI2 and an increase in H3K9me3. Surprisingly, the same cells with accumulation of L1-ORF1p display highest levels of HIWI2 and H3K9me3. Conversely, the earliest germ cells with high levels of L1-ORF1p express low levels of the chaperone HSP90α. We propose that a subset of germ cells resists L1 expression, whereas L1-expressing germ cells activate the repression pathway that leads to epigenetic silencing of L1 via H3K9me3.

Cyclone: an accessible pipeline to analyze, evaluate and optimize multiparametric cytometry data.

In the past decade, high-dimensional single cell technologies have revolutionized basic and translational immunology research and are now a key element of the toolbox used by scientists to study the immune system. However, analysis of the data generated by these approaches often requires clustering algorithms and dimensionality reduction representation which are computationally intense and difficult to evaluate and optimize. Here we present Cyclone, an analysis pipeline integrating dimensionality reduction, clustering, evaluation and optimization of clustering resolution, and downstream visualization tools facilitating the analysis of a wide range of cytometry data. We benchmarked and validated Cyclone on mass cytometry (CyTOF), full spectrum fluorescence-based cytometry, and multiplexed immunofluorescence (IF) in a variety of biological contexts, including infectious diseases and cancer. In each instance, Cyclone not only recapitulates gold standard immune cell identification, but also enables the unsupervised identification of lymphocytes and mononuclear phagocytes subsets that are associated with distinct biological features. Altogether, the Cyclone pipeline is a versatile and accessible pipeline for performing, optimizing, and evaluating clustering on variety of cytometry datasets which will further power immunology research and provide a scaffold for biological discovery.

Cyclone: an accessible pipeline to analyze, evaluate, and optimize multiparametric cytometry data.

In the past decade, high-dimensional single-cell technologies have revolutionized basic and translational immunology research and are now a key element of the toolbox used by scientists to study the immune system. However, analysis of the data generated by these approaches often requires clustering algorithms and dimensionality reduction representation, which are computationally intense and difficult to evaluate and optimize. Here, we present Cytometry Clustering Optimization and Evaluation (Cyclone), an analysis pipeline integrating dimensionality reduction, clustering, evaluation, and optimization of clustering resolution, and downstream visualization tools facilitating the analysis of a wide range of cytometry data. We benchmarked and validated Cyclone on mass cytometry (CyTOF), full-spectrum fluorescence-based cytometry, and multiplexed immunofluorescence (IF) in a variety of biological contexts, including infectious diseases and cancer. In each instance, Cyclone not only recapitulates gold standard immune cell identification but also enables the unsupervised identification of lymphocytes and mononuclear phagocyte subsets that are associated with distinct biological features. Altogether, the Cyclone pipeline is a versatile and accessible pipeline for performing, optimizing, and evaluating clustering on a variety of cytometry datasets, which will further power immunology research and provide a scaffold for biological discovery.

Heterogeneity of primordial germ cells.

Primordial germ cells (PGCs) must complete a complex and dynamic developmental program during embryogenesis to establish the germline. This process is highly conserved and involves a diverse array of tasks required of PGCs, including migration, survival, sex differentiation, and extensive epigenetic reprogramming. A common theme across many organisms is that PGC success is heterogeneous: only a portion of all PGCs complete all these steps while many other PGCs are eliminated from further germline contribution. The differences that distinguish successful PGCs as a population are not well understood. Here, we examine variation that exists in PGCs as they navigate the many stages of this developmental journey. We explore potential sources of PGC heterogeneity and their potential implications in affecting germ cell behaviors. Lastly, we discuss the potential for PGC development to function as a multistage selection process that assesses heterogeneity in PGCs to refine germline quality.

Growth Coordination During Drosophila melanogaster Imaginal Disc Regeneration Is Mediated by Signaling Through the Relaxin Receptor Lgr3 in the Prothoracic Gland.

Damage to Drosophila melanogaster imaginal discs activates a regeneration checkpoint that (1) extends larval development and (2) coordinates the regeneration of the damaged disc with the growth of undamaged discs. These two systemic responses to damage are both mediated by Dilp8, a member of the insulin/insulin-like growth factor/relaxin family of peptide hormones, which is released by regenerating imaginal discs. Growth coordination between regenerating and undamaged imaginal discs is dependent on Dilp8 activation of nitric oxide synthase (NOS) in the prothoracic gland (PG), which slows the growth of undamaged discs by limiting ecdysone synthesis. Here we demonstrate that the Drosophila relaxin receptor homolog Lgr3, a leucine-rich repeat-containing G-protein-coupled receptor, is required for Dilp8-dependent growth coordination and developmental delay during the regeneration checkpoint. Lgr3 regulates these responses to damage via distinct mechanisms in different tissues. Using tissue-specific RNA-interference disruption of Lgr3 expression, we show that Lgr3 functions in the PG upstream of NOS, and is necessary for NOS activation and growth coordination during the regeneration checkpoint. When Lgr3 is depleted from neurons, imaginal disc damage no longer produces either developmental delay or growth inhibition. To reconcile these discrete tissue requirements for Lgr3 during regenerative growth coordination, we demonstrate that Lgr3 activity in both the CNS and PG is necessary for NOS activation in the PG following damage. Together, these results identify new roles for a relaxin receptor in mediating damage signaling to regulate growth and developmental timing.