Extracellular vesicles (EVs) are a heterogeneous group of membrane‐enclosed structures produced by prokaryotic and eukaryotic cells. EVs carry a range of biological cargoes, including RNA, protein, and lipids, which may have both metabolic significance and signalling potential. EV release has been suggested to play a critical role in maintaining intracellular homeostasis by eliminating unnecessary biological material from EV producing cells, and as a delivery system to enable cellular communication between both neighbouring and distant cells without physical contact. In this review, we give an overview of what is known about the relative enrichment of the different types of RNA that have been associated with EVs in the most recent research efforts. We then examine the selective and non‐selective incorporation of these different RNA biotypes into EVs, the molecular systems of RNA sorting into EVs that have been elucidated so far, and the role of this process in EV‐producing cells. Finally, we also discuss the model systems providing evidence for EV‐mediated delivery of RNA to recipient cells, and the implications of this evidence for the relevance of this RNA delivery process in both physiological and pathological scenarios.

Key Points: Comprehensive genome-wide association study of eGFR in diabetes, accounting for diabetes duration, kidney disease, and known modifiers, identified novel genetic effects. Incorporation of various kidney multi-omics data provides supporting evidence for the role of novel genome-wide association study loci in diabetic kidney disease. Background: Diabetic kidney disease (DKD) is a serious diabetes complication caused by both environmental and genetic risk factors. Previous genome-wide association studies (GWAS) have identified several loci associated with kidney function and kidney disease in the general population and, to a lesser extent, in diabetes. Methods: To uncover the genetic factors driving diabetes-induced kidney function, we conducted a series of GWAS meta-analyses of eGFR in 17,267 individuals with type 1 diabetes and 35,264 with type 2 diabetes (52,531 total), using multiple well-characterized cohorts of type 1 diabetes DKD and data from the UK Biobank and SUrrogate markers for Micro- and Macrovascular hard end points for Innovative diabetes Tools (SUMMIT) consortium. We further accounted for DKD case/control status, diabetes duration and subtype, body mass index, glycated hemoglobin levels, and the relationship between eGFR and albuminuria. Results: GWAS identified 13 loci associated with eGFR ( P < 5×10 ⁻⁸ ), with five loci (candidate genes: HIPK3 , TRIM5 , RORA , ERBB4 , and BCL6 / LPP ) not associated with or were in opposite directions as compared with eGFR in the general population. Four candidate genes ( HIPK3 , BCL6, LPP , and RORA ) demonstrated evidence of differential expression in kidney compartments and cells among subgroups with DKD or diabetes versus controls. Lead single-nucleotide polymorphisms rs8027829 ( RORA ) and rs76300256 ( BCL6/LPP ) were methylation quantitative trait loci in whole blood and kidney tissue, respectively, and rs76300256 and its related CpGs all cluster in a kidney enhancer. Conclusions: Our integrated approach identified candidate genes with diabetes-specific effects on kidney function.

Cell‐cell communication is an essential aspect of multicellular life, key for coordinating cell proliferation, growth, and death in response to environmental changes. Whilst caspases are well‐known for facilitating apoptotic and pyroptotic cell death, several recent investigations are uncovering new roles for these enzymes in biological scenarios requiring long‐range intercellular signalling mediated by extracellular vesicles (EVs). EVs are small membrane‐bound nanoparticles released from cells that may carry and deliver cargo between distant cells, thus helping to coordinate their behaviour. Intriguingly, there is emerging evidence indicating a key contribution of caspases in the biogenesis of EVs, the selection of their cargo content, and EV uptake/function in recipient cells. Here, we discuss the latest findings supporting the interplay between caspases and EVs, and the biological relevance of this molecular convergence for cellular signalling, principally in non‐apoptotic scenarios.

The liver fluke, Fasciola hepatica, is a global burden on the wellbeing and productivity of farmed ruminants, and a zoonotic threat to human health. Despite the clear need for accelerated discovery of new drug and vaccine treatments for this pathogen, we still have a relatively limited understanding of liver fluke biology and host interactions. Noncoding RNAs, including micro (mi)RNAs, are key to transcriptional regulation in all eukaryotes, such that an understanding of miRNA biology can shed light on organismal function at a systems level. Four previous publications have reported up to 89 mature miRNA sequences from F. hepatica, but our data show that this does not represent a full account of this species miRNome. We have expanded on previous studies by sequencing, for the first time, miRNAs from multiple life stages (adult, newly excysted juvenile (NEJ), metacercariae and adult-derived extracellular vesicles (EVs)). These experiments detected an additional 61 high-confidence miRNAs, most of which have not been described in any other species, expanding the F. hepatica miRNome to 150 mature sequences. We used quantitative (q)PCR assays to provide the first developmental profile of miRNA expression across metacercariae, NEJ, adult and adult-derived Evs. The majority of miRNAs were expressed most highly in metacercariae, with at least six distinct expression clusters apparent across life stages. Intracellular miRNAs were functionally analyzed to identify target mRNAs with inversely correlated expression in F. hepatica tissue transcriptomes, highlighting regulatory interactions with key virulence transcripts including cathepsin proteases, and neuromuscular genes that control parasite growth, development and motility. We also linked 28 adult-derived EV miRNAs with downregulation of 397 host genes in F. hepatica-infected transcriptomes from ruminant lymph node, peripheral blood mononuclear cell (PBMC) and liver tissue transcriptomes. These included genes involved in signal transduction, immune and metabolic pathways, adding to the evidence for miRNA-based immunosuppression during fasciolosis. These data expand our understanding of the F. hepatica miRNome, provide the first data on developmental miRNA regulation in this species, and provide a set of testable hypotheses for functional genomics interrogations of liver fluke miRNA biology.

The existence of three independent binary systems for conditional gene expression (Gal4/UAS; LexA/LexAop; QF/QUAS) has greatly expanded versatile genetic analyses in the Drosophila melanogaster; however, the experimental application of these tools is limited by the need to generate multiple collections of noninterchangeable transgenic fly strains for each inducible gene expression system. To address this practical limitation, we developed a modular vector that contains the regulatory elements from all three binary systems, enabling Gal4-, LexA- or QF-dependent expression of transgenes. Our methods also incorporate DNA elements that facilitate independent site-specific recombination and elimination of regulatory UAS, LexAop or QUAS modules with spatial and temporal control, thus offering unprecedented possibilities and logistical advantages for in vivo genetic modulation and efficient interconversion of overexpression transgenic fly lines.

Telomeres are protective structures at the end of eukaryotic chromosomes that are strongly implicated in ageing and ill health. They attrition upon every cellular reproductive cycle. Evidence suggests that short telomeres trigger DNA damage responses that lead to cellular senescence. Accurate methods for measuring telomeres are required to fully investigate the roles that shortening telomeres play in the biology of disease and human ageing. The last two decades have brought forth several techniques that are used for measuring telomeres. This editorial highlights strengths and limitations of traditional and emerging techniques, guiding researchers to choose the most appropriate methodology for their research needs. These methods include Quantitative Polymerase Chain Reaction (qPCR), Omega qPCR (Ω-qPCR), Terminal Restriction Fragment analysis (TRF), Single Telomere Absolute-length Rapid (STAR) assays, Single TElomere Length Analysis (STELA), TElomere Shortest Length Assays (TESLA), Telomere Combing Assays (TCA), and Long-Read Telomere Sequencing. Challenges include replicating telomere measurement within and across cohorts, measuring the length of telomeres on individual chromosomes, and standardised reporting for publications. Areas of current and future focus have been highlighted, with recent methodical advancements, such as long-read sequencing, providing significant scope to study telomeres at an individual chromosome level.