Biology subjects

Conine, C. C.

Publications and source records attributed to Conine, C. C..

13 recordsLinked to original sources

bioRxiv2026

Comparative profiling of sperm small RNAs in cnidarians highlights potential for paternal inheritance

Small RNAs in mature sperm are crucial regulators of gene expression following fertilization during early development in bilaterians. However, the presence and functions of sperm-derived small RNAs remain poorly understood in early-diverging (i.e., non-bilaterian) metazoans. In this study, we characterize three classes of small RNAs-microRNAs (miRNAs), piwi-interacting RNAs (piRNAs), and tRNA-derived RNAs (tDRs)-in sperm from three phylogenetically and ecologically distinct species from the early-diverging phylum Cnidaria: the sea anemone Nematostella vectensis, the temperate stony coral Astrangia poculata, and the reef-building tropical coral Acropora hyacinthus. All three classes of small RNAs examined were present in sperm from these cnidarians. Most miRNAs were species-specific; however, target prediction analysis revealed enriched putative targets involved in conserved developmental and signaling pathways. piRNAs overlapped with transposable elements and genic regions, indicating a potential dual role in genomic maintenance and gene expression regulation. Characterization of tDRs demonstrated that 5 prime-terminal fragments constituted the primary structural type across the three species, driven by high accumulation of specific isoacceptors including 5 prime-tDR-GlyGCC. Together, these findings provide the first evidence that cnidarian sperm carry a rich repertoire of small RNAs, including both conserved and species-specific types. Their presence suggests that paternal small RNAs contribute important regulatory information to the zygote in cnidarians, implicating paternal small RNAs as possible mediators of epigenetic inheritance in this early-branching metazoan phylum.

molecular biology↗

bioRxiv2026

Multi-omic profiling of early pregnancy small and large plasma extracellular vesicles reveals placental, metabolic, and structural adaptation signatures

Early human pregnancy is a critical period characterized by rapid growth and extensive maternal-fetal communication that influence maternal and fetal outcomes. Circulating extracellular vesicles (EVs) have the capacity to capture cargo that reflect these processes in real-time; however, signatures of EV subtypes during early pregnancy are poorly defined. Here we quantified mitochondrial DNA (mtDNA) and performed transcriptomic and proteomic profiling of small ([~]100 nm) and large ([~]200 nm) plasma EVs from n=10 normal pregnancies (11-15 weeks) to define subtype-specific molecular signatures. mtDNA and mitochondrial protein content were more abundant in large EVs (lEVs). lEVs also contained a more complex set of long RNAs enriched for placental, immune, and mitochondrial-related transcripts compared with small EVs (sEVs). Proteomic profiling showed enrichment of canonical EV markers and extracellular matrix proteins in sEVs, whereas lEVs were preferentially associated with pregnancy-specific proteins, including proteins related to placental hormone production. MicroRNAs (miRNAs) accounted for [~]25% of small RNAs in both EV subtypes with miR-223 and miR-16 enriched in lEVs and miR-639 enriched in sEVs. These data together, support a model where small and large plasma EVs have distinct, yet complementary signatures reporting systemic adaptations during the critical 11-15 week transition period. This work establishes a foundational framework for future studies linking EV signatures to placental dysfunction and adverse outcomes.

physiology↗

bioRxiv2026

Acute Paternal Immune Activation Shapes Embryonic Development and Protects Offspring from Viral Infection

The evolutionary arms race between host and pathogen is traditionally framed as a multigenerational paradigm, where beneficial host immune adaptations accrue in a population over time through DNA sequence variation. If pathogens exposure alters epigenetic information inherited by offspring through paternal gametes, advantageous immune traits could emerge within a single generation. We show that acute paternal immune activation (PIA) induces immune signaling in the male reproductive tract and remodels the sperm small RNA profile. Using IVF, we show that altered sperm small RNAs from immune-activated males reprograms gene expression in preimplantation embryos, eliciting sex-specific transcriptional responses. This transcriptional program is reflected in a striking inherited phenotype, as male offspring sired by PIA males exhibit enhanced survival following lethal viral challenge. These findings identify sperm small RNAs as a molecular mechanism that links PIA to embryonic development and offspring immunity, providing a comprehensive framework for how PIA shapes inherited pathogen protection.

immunology↗

bioRxiv2026

Physiologic variation in sperm miRNAs tune embryonic gene regulatory programs and developmental outcomes

Small RNAs delivered by sperm can transmit environmentally regulated, epigenetically inherited phenotypes to offspring, yet the mechanisms by which modest changes in sperm microRNA abundance overcome dilution within the much larger egg to influence embryonic development remain unresolved. Here, we show that physiologically relevant variation in individual sperm miRNAs is sufficient to quantitatively program embryonic gene expression and developmental outcomes. Using parthenogenetic and fertilized embryos, we show that as few as 200 molecules of miR-200c-3p or miR-465c-3p induces reproducible, dose-dependent gene expression responses across defined developmental windows. Parthenogenetic embryos faithfully recapitulate early miRNA-driven gene expression changes observed in fertilized embryos, validating their use for isolating early regulatory mechanisms. We further developed AGO2-REMORA, an RNA adenosine base editor fused to Argonaute2 to map miRNA-mRNA interactions in embryos, revealing that early mRNA repression reflects direct miRNA targeting, while transcriptional changes at later stages arise as secondary consequences of these initial interactions. Furthermore, we show that modest elevation of miR-200c-3p during early development is sufficient to induce transcriptional alterations through early development and produce craniofacial phenotypes in late-stage embryos, recapitulating features of fetal alcohol syndrome associated with paternal alcohol consumption. Together, these findings establish a generalizable framework by which small perturbations in sperm miRNA content quantitatively modulate early gene regulatory programs, triggering cascades that persist throughout development and influence offspring phenotype.

developmental biology↗

bioRxiv2025

Pseudouridine selects RNAs for extracellular transport

RNAs move through the extracellular space to transmit information between cells, including mammalian neurons, yet how specific RNAs are channeled into these extracellular routes is unknown. Using genome-wide CRISPR screening, proteomics, and high-sensitivity transcriptomics in a neuronal model system, we identify domesticated retroviral proteins and RNA-modifying enzymes that regulate RNA loading into and transportation via extracellular vesicles. We show that the pseudouridine synthase PUS1 is a key determinant of RNA trafficking, and that its catalytic product in RNA, pseudouridine, is enriched in extracellular RNAs from transformed and primary neurons. Furthermore, the presence of pseudouridine on select RNAs is both necessary and sufficient for their extracellular export. Finally, we show that myosin light chain 6 (MYL6) is a pseudouridine-binding protein required for secretion of synthetic and endogenous RNAs. These findings reveal a biochemical code linking chemical RNA modification to extracellular transport, and establish a framework to study the function of extracellular RNAs in the nervous system and beyond.

molecular biology↗

bioRxiv2025

Epididymal extracellular vesicles harbor and convey mRNA to sperm for transfer to zygotes

The epididymis plays a critical role in sperm maturation, including remodeling the sperm RNA payload. While small RNAs have been extensively studied in this context, the epididymal contribution to larger sperm RNAs, such as mRNAs, remains underexplored. mRNAs were among the first RNA species identified in sperm, yet their functional relevance has remained elusive, largely due to the translational quiescence of mature spermatozoa and the hypothesis that these RNAs are residual by-products of spermatogenesis. Yet, mRNAs carried by sperm have been detected in the zygote, indicating they could play a role beyond fertilization. However, if the soma, the epididymis, actively contributes mRNAs to sperm as it does small RNAs, has not been experimentally assessed. To investigate this, here, we provide a comprehensive analysis of the mRNA landscape of mouse sperm, epithelial cells and extracellular vesicles (EVs) isolated from the proximal (caput) and distal (cauda) epididymis. Through this analysis and sperm-EV co-incubation experiments we demonstrate the transfer of mRNAs from epididymal EVs to sperm. Further, through sperm RNA microinjection into zygotes we uncover gene regulation in the early embryo driven by the introduction of sperm RNAs, specific to large RNA species. These findings ultimately reveal the dynamic mRNA profile of sperm that is delivered to the egg and demonstrate that RNA species beyond small RNAs are capable of influencing preimplantation embryo gene expression.

cell biology↗

bioRxiv2025

tRNA-derived RNA processing in sperm transmits non-genetically inherited phenotypes to offspring in C. elegans

The environment encountered by an organism can modulate epigenetic information in gametes to transmit non-genetically inherited phenotypes to offspring. In mouse models, the diet of males regulates specific tRNA-derived RNAs (tDRs) in sperm. After fertilization, tDRs regulate embryonic gene expression and generate metabolic phenotypes in adult offspring through uncharacterized changes during development. Here we demonstrate that in C. elegans, tDRs also accumulate in sperm and can similarly transmit epigenetically inherited phenotypes to offspring. We identify the RNaseT2 enzyme, rnst-2, as a regulator of C. elegans tDR accumulation. RNST- 2 processes or degrades tRNA-halves, leading to short <30 nt fragments. This rnst-2 dependent regulation of tDR length distribution modulates specific tDRs in sperm which, after fertilization, regulate early embryonic and developmental gene expression, leading to adaptive phenotypes in progeny. Our findings establish tDRs as a deeply conserved carrier of intergenerational epigenetic information and position the worm as a model for dissecting paternal non-genetic inheritance mechanistically.

developmental biology↗

bioRxiv2024

The effect of intrauterine growth restriction on the developing pancreatic immune system

Immune cells in the pancreas are known to participate in organ development. However, the resident pancreatic immune system has yet to be fully defined. Immune cells also play a role in pathology and are implicated in diseases such as diabetes induced by intrauterine growth restriction (IUGR). We hypothesized that the resident immune system is established during neonatal development and disrupted by IUGR. Using single cell RNAseq and flow cytometry we identified many immune cell populations in the near-term fetus (at embryologic day 22) and neonatal (postnatal day 1, 7, &14) islets, non-endocrine pancreas, and the spleen in the rat. Using flow cytometry, we observed the resident immune system is established during neonatal development in the pancreas and spleen. We identified 9 distinct immune populations in the pancreatic islets and 8 distinct immune populations in the spleen by single cell RNAseq. There were no sex-specific differences in the relative proportion of immune cells in the pancreas or spleen. Finally, we tested if IUGR disrupted the neonatal immune system using bilateral uterine artery ligation. We found significant changes to the percentage of CD11B+ HIS48- and CD8+ T cells in the islets and non-endocrine pancreas and in the spleen. IUGR-induced alterations were influenced by the tissue environment and the sex of the offspring. Future research to define the role of these immune cells in pancreatic development may identify disrupted pathways that contribute to the development of diabetes following IUGR.

immunology↗

bioRxiv2024

Sub-chronic elevation in ambient temperature drives alterations to the sperm epigenome and accelerates early embryonic development in mice

Forecasted increases in the prevalence and severity of extreme weather events accompanying changes in climatic behavior pose potential risk to the reproductive capacity of humans and animals of ecological and agricultural significance. While several studies have revealed that heat stress induced by challenges such as testicular insulation can elicit a marked negative effect on the male reproductive system, and particularly the production of spermatozoa, less is known about the immediate impact on male reproductive function following sub-chronic whole-body exposure to elevated ambient temperature. To address this knowledge gap, we exposed unrestrained male mice to heat stress conditions that emulate a heat wave (daily cycle of 8_h at 35{degrees}C followed by 16 h at 25{degrees}C) for a period of seven days. Neither the testes or epididymides of heat exposed male mice exhibited evidence of gross histological change, and similarly, spermatozoa of exposed males retained their functionality and ability to support embryonic development. However, the embryos generated from heat exposed spermatozoa experienced pronounced changes in gene expression linked to acceleration of early embryo development, aberrant blastocyst hatching and increased fetal weight. Such changes were causally associated with an altered sperm small non-coding RNA (sncRNA) profile, such that these developmental phenotypes were recapitulated by microinjection of wild-type embryos sired by control spermatozoa with RNAs extracted from heat exposed spermatozoa. Such data highlight that even a relatively modest excursion in ambient temperature can affect male reproductive function and identify the sperm sncRNA profile as a particular point of vulnerability to this imposed environmental stress. Significance StatementThe fidelity of sperm production underpins successful reproduction yet is highly vulnerable to various forms of environmental challenge, including heat stress. Despite this knowledge, we lack a complete understanding of the immediate impact on male reproduction of whole-body exposure to elevated ambient temperatures such as those encountered during a heatwave. By experimentally emulating heatwave conditions, we demonstrate that the spermatozoa of exposed male mice accumulate changes in their small RNA profile that are causally linked to pronounced changes in embryonic gene expression, accelerated pre-implantation development, aberrant blastocyst hatching, and increased fetal weight. Such data highlight that even a relatively modest alteration in ambient temperature can affect male reproductive function, demonstrating the acute sensitivity of sperm small RNAs to environmental stress.

cell biology↗

bioRxiv2024

COVID-19 during pregnancy alters circulating extracellular vesicle cargo and their effects on trophoblast

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection and resulting coronavirus disease (COVID-19) causes placental dysfunction, which increases the risk of adverse pregnancy outcomes. While abnormal placental pathology resulting from COVID-19 is common, direct infection of the placenta is rare. This suggests that pathophysiology associated with maternal COVID-19, rather than direct placental infection, is responsible for placental dysfunction and alteration of the placental transcriptome. We hypothesized that maternal circulating extracellular vesicles (EVs), altered by COVID-19 during pregnancy, contribute to placental dysfunction. To examine this hypothesis, we characterized maternal circulating EVs from pregnancies complicated by COVID-19 and tested their effects on trophoblast cell physiology in vitro. We found that the gestational timing of COVID-19 is a major determinant of circulating EV function and cargo. In vitro trophoblast exposure to EVs isolated from patients with an active infection at the time of delivery, but not EVs isolated from Controls, altered key trophoblast functions including hormone production and invasion. Thus, circulating EVs from participants with an active infection, both symptomatic and asymptomatic cases, can disrupt vital trophoblast functions. EV cargo differed between participants with COVID-19 and Controls, which may contribute to the disruption of the placental transcriptome and morphology. Our findings show that COVID-19 can have effects throughout pregnancy on circulating EVs and circulating EVs are likely to participate in placental dysfunction induced by COVID-19.

pathology↗

bioRxiv2023

Epididymal acquired sperm microRNAs modify post-fertilization embryonic gene expression

Sperm small RNAs have emerged as important non-genetic contributors to embryogenesis and offspring health. A subset of sperm small RNAs are thought to be acquired during epididymal transit. However, the transfer of RNAs from the somatic epididymis to sperm has been questioned, and the identity of the specific small RNAs transferred remains unclear. Here, we employ Cre/Lox genetics to generate germline- and epididymal-specific Dgcr8 conditional knockout mice to investigate the dynamics of sperm microRNAs and their function in the early embryo. Testicular sperm from germline specific Dgcr8 knockout mice have reduced levels of 98 microRNAs. Enthrallingly, following epididymal transit the abundance of 59% of these microRNAs are restored to control levels. Conversely, sperm from epididymal Dgcr8 knockouts displayed a reduction of > 3.4-fold in 25 miRNAs. This substantial loss of epididymal miRNAs in sperm was accompanied by transcriptomic changes in the embryo which was rescued by microinjection of epididymal miRNAs. These findings ultimately demonstrate the acquisition of miRNAs from the soma by sperm during epididymal transit and their subsequent regulation of post-fertilization embryonic gene expression.

developmental biology↗

bioRxiv2023

A ligation-independent sequencing method reveals tRNA-derived RNAs with blocked 3' termini

Despite the numerous sequencing methods available, the vast diversity in size and chemical modifications of RNA molecules makes the capture of the full spectrum of cellular RNAs a difficult task. By combining quasirandom hexamer priming with a custom template switching strategy, we developed a method to construct sequencing libraries from RNA molecules of any length and with any type of 3 terminal modification, allowing the sequencing and analysis of virtually all RNA species. Ligation-independent detection of all types of RNA (LIDAR) is a simple, effective tool to comprehensively characterize changes in small non-coding RNAs and mRNAs simultaneously, with performance comparable to separate dedicated methods. With LIDAR, we comprehensively characterized the coding and non- coding transcriptome of mouse embryonic stem cells, neural progenitor cells, and sperm. LIDAR detected a much larger variety of tRNA-derived RNAs (tDRs) compared to traditional ligation-dependent sequencing methods, and uncovered the presence of tDRs with blocked 3 ends that had previously escaped detection. Our findings highlight the potential of LIDAR to systematically detect all RNAs in a sample and uncover new RNA species with potential regulatory functions.

genomics↗

bioRxiv2023

The microbiota and immune system non-genetically affect offspring phenotypes transgenerationally

Summary paragraphThe host-microbiota relationship has evolved to shape mammalian processes, including immunity, metabolism, and development1-3. Host phenotypes change in direct response to microbial exposures by the individual. Here we show that the microbiota induces phenotypic change not only in the individual but also in their succeeding generations of progeny. We found that germ-free mice exhibit a robust sebum secretion defect and transcriptional changes in various organs, persisting across multiple generations despite microbial colonization and breeding with conventional mice. Host-microbe interactions could be involved in this process, since T cell-deficient mice, which display defective sebum secretion4, also transgenerationally transmit their phenotype to progeny. These phenotypes are inherited by progeny conceived during in vitro fertilization using germ-free sperm and eggs, demonstrating that epigenetic information in the gametes is required for phenotypic transmission. Accordingly, small non-coding RNAs that can regulate embryonic gene expression5 were strikingly and similarly altered in gametes of germ-free and T cell-deficient mice. Thus, we have uncovered a novel mechanism whereby the microbiota and immune system induce phenotypic changes in successive generations of offspring. This epigenetic form of inheritance could be advantageous for host adaptation to environmental perturbation, where phenotypic diversity can be introduced more rapidly than by genetic mutation.

microbiology↗