Decoupled evolution of the Sex Peptide gene family and Sex Peptide Receptor in Drosophilidae.

Across internally fertilising species, males transfer ejaculate proteins that trigger wide-ranging changes in female behaviour and physiology. Much theory has been developed to explore the drivers of ejaculate protein evolution. The accelerating availability of high-quality genomes now allows us to test how these proteins are evolving at fine taxonomic scales. Here, we use genomes from 264 species to chart the evolutionary history of Sex Peptide (SP), a potent regulator of female post-mating responses in Drosophila melanogaster. We infer that SP first evolved in the Drosophilinae subfamily and has since followed markedly different evolutionary trajectories in different lineages. Outside of the Sophophora-Lordiphosa, SP exists largely as a single-copy gene with independent losses in several lineages. Within the Sophophora-Lordiphosa, the SP gene family has repeatedly and independently expanded. Up to seven copies, collectively displaying extensive sequence variation, are present in some species. Despite these changes, SP expression remains restricted to the male reproductive tract. Alongside, we document considerable interspecific variation in the presence and morphology of seminal microcarriers that, despite the critical role SP plays in microcarrier assembly in D. melanogaster, appears to be independent of changes in the presence/absence or sequence of SP. We end by providing evidence that SP's evolution is decoupled from that of its receptor, Sex Peptide Receptor, in which we detect no evidence of correlated diversifying selection. Collectively, our work describes the divergent evolutionary trajectories that a novel gene has taken following its origin and finds a surprisingly weak coevolutionary signal between a supposedly sexually antagonistic protein and its receptor.

A single-cell atlas of the sexually dimorphic Drosophila foreleg and its sensory organs during development.

To respond to the world around them, animals rely on the input of a network of sensory organs distributed throughout the body. Distinct classes of sensory organs are specialized for the detection of specific stimuli such as strain, pressure, or taste. The features that underlie this specialization relate both to the neurons that innervate sensory organs and the accessory cells they comprise. To understand the genetic basis of this diversity of cell types, both within and between sensory organs, we performed single-cell RNA sequencing on the first tarsal segment of the male Drosophila melanogaster foreleg during pupal development. This tissue displays a wide variety of functionally and structurally distinct sensory organs, including campaniform sensilla, mechanosensory bristles, and chemosensory taste bristles, as well as the sex comb, a recently evolved male-specific structure. In this study, we characterize the cellular landscape in which the sensory organs reside, identify a novel cell type that contributes to the construction of the neural lamella, and resolve the transcriptomic differences among support cells within and between sensory organs. We identify the genes that distinguish between mechanosensory and chemosensory neurons, resolve a combinatorial transcription factor code that defines 4 distinct classes of gustatory neurons and several types of mechanosensory neurons, and match the expression of sensory receptor genes to specific neuron classes. Collectively, our work identifies core genetic features of a variety of sensory organs and provides a rich, annotated resource for studying their development and function.

Drosophila Sex Peptide controls the assembly of lipid microcarriers in seminal fluid.

Seminal fluid plays an essential role in promoting male reproductive success and modulating female physiology and behavior. In the fruit fly, Drosophila melanogaster, Sex Peptide (SP) is the best-characterized protein mediator of these effects. It is secreted from the paired male accessory glands (AGs), which, like the mammalian prostate and seminal vesicles, generate most of the seminal fluid contents. After mating, SP binds to spermatozoa and is retained in the female sperm storage organs. It is gradually released by proteolytic cleavage and induces several long-term postmating responses, including increased ovulation, elevated feeding, and reduced receptivity to remating, primarily signaling through the SP receptor (SPR). Here, we demonstrate a previously unsuspected SPR-independent function for SP. We show that, in the AG lumen, SP and secreted proteins with membrane-binding anchors are carried on abundant, large neutral lipid-containing microcarriers, also found in other SP-expressing Drosophila species. These microcarriers are transferred to females during mating where they rapidly disassemble. Remarkably, SP is a key microcarrier assembly and disassembly factor. Its absence leads to major changes in the seminal proteome transferred to females upon mating. Males expressing nonfunctional SP mutant proteins that affect SP's binding to and release from sperm in females also do not produce normal microcarriers, suggesting that this male-specific defect contributes to the resulting widespread abnormalities in ejaculate function. Our data therefore reveal a role for SP in formation of seminal macromolecular assemblies, which may explain the presence of SP in Drosophila species that lack the signaling functions seen in Dmelanogaster.

The P. furiosus mre11/rad50 complex promotes 5' strand resection at a DNA double-strand break.

The Mre11/Rad50 complex has been implicated in the early steps of DNA double-strand break (DSB) repair through homologous recombination in several organisms. However, the enzymatic properties of this complex are incompatible with the generation of 3' single-stranded DNA for recombinase loading and strand exchange. In thermophilic archaea, the Mre11 and Rad50 genes cluster in an operon with genes encoding a helicase, HerA, and a 5' to 3' exonuclease, NurA, suggesting a common function. Here we show that purified Mre11 and Rad50 from Pyrococcus furiosus act cooperatively with HerA and NurA to resect the 5' strand at a DNA end under physiological conditions in vitro. The 3' single-stranded DNA generated by these enzymes can be utilized by the archaeal RecA homolog RadA to catalyze strand exchange. This work elucidates how the conserved Mre11/Rad50 complex promotes DNA end resection in archaea and may serve as a model for DSB processing in eukaryotes.

Male reproductive aging arises via multifaceted mating-dependent sperm and seminal proteome declines, but is postponable in Drosophila.

Declining ejaculate performance with male age is taxonomically widespread and has broad fitness consequences. Ejaculate success requires fully functional germline (sperm) and soma (seminal fluid) components. However, some aging theories predict that resources should be preferentially diverted to the germline at the expense of the soma, suggesting differential impacts of aging on sperm and seminal fluid and trade-offs between them or, more broadly, between reproduction and lifespan. While harmful effects of male age on sperm are well known, we do not know how much seminal fluid deteriorates in comparison. Moreover, given the predicted trade-offs, it remains unclear whether systemic lifespan-extending interventions could ameliorate the declining performance of the ejaculate as a whole. Here, we address these problems using Drosophila melanogaster. We demonstrate that seminal fluid deterioration contributes to male reproductive decline via mating-dependent mechanisms that include posttranslational modifications to seminal proteins and altered seminal proteome composition and transfer. Additionally, we find that sperm production declines chronologically with age, invariant to mating activity such that older multiply mated males become infertile principally via reduced sperm transfer and viability. Our data, therefore, support the idea that both germline and soma components of the ejaculate contribute to male reproductive aging but reveal a mismatch in their aging patterns. Our data do not generally support the idea that the germline is prioritized over soma, at least, within the ejaculate. Moreover, we find that lifespan-extending systemic down-regulation of insulin signaling results in improved late-life ejaculate performance, indicating simultaneous amelioration of both somatic and reproductive aging.

BMP signaling inhibition in Drosophila secondary cells remodels the seminal proteome and self and rival ejaculate functions.

Seminal fluid proteins (SFPs) exert potent effects on male and female fitness. Rapidly evolving and molecularly diverse, they derive from multiple male secretory cells and tissues. In Drosophila melanogaster, most SFPs are produced in the accessory glands, which are composed of ∼1,000 fertility-enhancing "main cells" and ∼40 more functionally cryptic "secondary cells." Inhibition of bone morphogenetic protein (BMP) signaling in secondary cells suppresses secretion, leading to a unique uncoupling of normal female postmating responses to the ejaculate: refractoriness stimulation is impaired, but offspring production is not. Secondary-cell secretions might therefore make highly specific contributions to the seminal proteome and ejaculate function; alternatively, they might regulate more global-but hitherto undiscovered-SFP functions and proteome composition. Here, we present data that support the latter model. We show that in addition to previously reported phenotypes, secondary-cell-specific BMP signaling inhibition compromises sperm storage and increases female sperm use efficiency. It also impacts second male sperm, tending to slow entry into storage and delay ejection. First male paternity is enhanced, which suggests a constraint on ejaculate evolution whereby high female refractoriness and sperm competitiveness are mutually exclusive. Using quantitative proteomics, we reveal changes to the seminal proteome that surprisingly encompass alterations to main-cell-derived proteins, indicating important cross-talk between classes of SFP-secreting cells. Our results demonstrate that ejaculate composition and function emerge from the integrated action of multiple secretory cell types, suggesting that modification to the cellular make-up of seminal-fluid-producing tissues is an important factor in ejaculate evolution.

Divergent allocation of sperm and the seminal proteome along a competition gradient in Drosophila melanogaster.

Sperm competition favors large, costly ejaculates, and theory predicts the evolution of allocation strategies that enable males to plastically tailor ejaculate expenditure to sperm competition threat. While greater sperm transfer in response to a perceived increase in the risk of sperm competition is well-supported, we have a poor understanding of whether males (i) respond to changes in perceived intensity of sperm competition, (ii) use the same allocation rules for sperm and seminal fluid, and (iii) experience changes in current and future reproductive performance as a result of ejaculate compositional changes. Combining quantitative proteomics with fluorescent sperm labeling, we show that Drosophila melanogaster males exercise independent control over the transfer of sperm and seminal fluid proteins (SFPs) under different levels of male-male competition. While sperm transfer peaks at low competition, consistent with some theoretical predictions based on sperm competition intensity, the abundance of transferred SFPs generally increases at high competition levels. However, we find that clusters of SFPs vary in the directionality and sensitivity of their response to competition, promoting compositional change in seminal fluid. By tracking the degree of decline in male mating probability and offspring production across successive matings, we provide evidence that ejaculate compositional change represents an adaptive response to current sperm competition, but one that comes at a cost to future mating performance. Our work reveals a previously unknown divergence in ejaculate component allocation rules, exposes downstream costs of elevated ejaculate investment, and ultimately suggests a central role for ejaculate compositional plasticity in sexual selection.

Quantitative Proteomics Identification of Seminal Fluid Proteins in Male Drosophila melanogaster.

Seminal fluid contains some of the fastest evolving proteins currently known. These seminal fluid proteins (Sfps) play crucial roles in reproduction, such as supporting sperm function, and particularly in insects, modifying female physiology and behavior. Identification of Sfps in small animals is challenging, and often relies on samples taken from the female reproductive tract after mating. A key pitfall of this method is that it might miss Sfps that are of low abundance because of dilution in the female-derived sample or rapid processing in females. Here we present a new and complementary method, which provides added sensitivity to Sfp identification. We applied label-free quantitative proteomics to Drosophila melanogaster, male reproductive tissue - where Sfps are unprocessed, and highly abundant - and quantified Sfps before and immediately after mating, to infer those transferred during copulation. We also analyzed female reproductive tracts immediately before and after copulation to confirm the presence and abundance of known and candidate Sfps, where possible. Results were cross-referenced with transcriptomic and sequence databases to improve confidence in Sfp detection. Our data were consistent with 125 previously reported Sfps. We found nine high-confidence novel candidate Sfps, which were both depleted in mated versus, unmated males and identified within the reproductive tract of mated but not virgin females. We also identified 42 more candidates that are likely Sfps based on their abundance, known expression and predicted characteristics, and revealed that four proteins previously identified as Sfps are at best minor contributors to the ejaculate. The estimated copy numbers for our candidate Sfps were lower than for previously identified Sfps, supporting the idea that our technique provides a deeper analysis of the Sfp proteome than previous studies. Our results demonstrate a novel, high-sensitivity approach to the analysis of seminal fluid proteomes, whose application will further our understanding of reproductive biology.

Survival factor NFIL3 restricts FOXO-induced gene expression in cancer.

Depending on the circumstance, FOXO (Forkhead O) (FOXO1, FOXO3, and FOXO4) transcription factors activate the expression of markedly different sets of genes to produce different phenotypic effects. For example, distinct FOXO-regulated transcriptional programs stimulate cell death or enhance organism life span. To gain insight into how FOXOs select specific genes for regulation, we performed a screen for genes that modify FOXO activation of TRAIL, a death receptor ligand capable of inducing extrinsic apoptosis. We discovered that the bZIP transcriptional repressor NFIL3 (nuclear factor interleukin 3-regulated) hindered FOXO transcription factor access to chromatin at the TRAIL promoter by binding to nearby DNA and recruiting histone deacetylase-2 (HDAC2) to reduce histone acetylation. In the same manner, NFIL3 repressed expression of certain FOXO targets--e.g., FAS, GADD45α (growth arrest and DNA damage-inducible, α), and GADD45ß--but not others. NFIL3, which we found to be overexpressed in different cancers, supported tumor cell survival largely through repression of TRAIL and antagonized hydrogen peroxide-induced cell death. Moreover, its expression in cancer was associated with lower patient survival. Therefore, NFIL3 alters cancer cell behavior and FOXO function by acting on chromatin to restrict the menu of FOXO target genes. Targeting of NFIL3 could be of therapeutic benefit for cancer patients.

Multifold Enhancement of Third-Harmonic Generation in Dielectric Nanoparticles Driven by Magnetic Fano Resonances.

Strong Mie-type magnetic dipole resonances in all-dielectric nanostructures provide novel opportunities for enhancing nonlinear effects at the nanoscale due to the intense electric and magnetic fields trapped within the individual nanoparticles. Here we study third-harmonic generation from quadrumers of silicon nanodisks supporting high-quality collective modes associated with the magnetic Fano resonance. We observe nontrivial wavelength and angular dependencies of the generated harmonic signal featuring a multifold enhancement of the nonlinear response in oligomeric systems.

Hunk/Mak-v is a negative regulator of intestinal cell proliferation.

BACKGROUND: Conditional deletion of the tumour suppressor gene Apc within the murine intestine results in acute Wnt signalling activation. The associated over-expression of a myriad of Wnt signalling target genes yields phenotypic alterations that encompass many of the hallmarks of neoplasia. Previous transcriptomic analysis aimed at identifying genes that potentially play an important role in this process, inferred the Hormonally upregulated Neu-associated kinase (HUNK/Mak-v/Bstk1) gene as a possible candidate. Hunk is a SNF1 (sucrose non fermenting 1)-related serine/threonine kinase with a proposed association with many different tumour types, including colorectal cancer. METHODS: Here we describe the generation of a novel Hunk kinase deficient mouse which has been used to investigate the involvement of Hunk-kinase activity in intestinal homeostasis and tumourigenesis. RESULTS: We show that in the morphologically normal intestine, Hunk-kinase negatively regulates epithelial cell proliferation. However, the increase in cell proliferation observed in the Hunk kinase deficient intestine is counteracted by increased cell migration, thereby maintaining intestinal homeostasis. Using qRT-PCR, we further demonstrate that Hunk is significantly over-expressed in Apc deficient / Wnt-signalling activated intestinal tissue. Using the classical intestinal tumourigenesis Apc (Min) mouse model we show that loss of Hunk-kinase activity significantly reduced tumour initiation rates in the small intestine. However, an accompanying increase in the size of the tumours counteracts the impact this has on overall tumour burden or subsequently survival. CONCLUSIONS: In the intestinal setting we demonstrate that Hunk has a role in normal intestinal proliferation and homeostasis and, although it does not alter overall survival rates, activity of this kinase does impact on tumour initiation rates during the early stages in tumourigenesis in the small intestine.

Enhanced third-harmonic generation in silicon nanoparticles driven by magnetic response.

We observe enhanced third-harmonic generation from silicon nanodisks exhibiting both electric and magnetic dipolar resonances. Experimental characterization of the nonlinear optical response through third-harmonic microscopy and spectroscopy reveals that the third-harmonic generation is significantly enhanced in the vicinity of the magnetic dipole resonances. The field localization at the magnetic resonance results in two orders of magnitude enhancement of the harmonic intensity with respect to unstructured bulk silicon with the conversion efficiency limited only by the two-photon absorption in the substrate.

Observation of Fano resonances in all-dielectric nanoparticle oligomers.

It is well-known that oligomers made of metallic nanoparticles are able to support sharp Fano resonances originating from the interference of two plasmonic resonant modes with different spectral width. While such plasmonic oligomers suffer from high dissipative losses, a new route for achieving Fano resonances in nanoparticle oligomers has opened up after the recent experimental observations of electric and magnetic resonances in low-loss dielectric nanoparticles. Here, light scattering by all-dielectric oligomers composed of silicon nanoparticles is studied experimentally for the first time. Pronounced Fano resonances are observed for a variety of lithographically-fabricated heptamer nanostructures consisting of a central particle of varying size, encircled by six nanoparticles of constant size. Based on a full collective mode analysis, the origin of the observed Fano resonances is revealed as a result of interference of the optically-induced magnetic dipole mode of the central particle with the collective mode of the nanoparticle structure. This allows for effective tuning of the Fano resonance to a desired spectral position by a controlled size variation of the central particle. Such optically-induced magnetic Fano resonances in all-dielectric oligomers offer new opportunities for sensing and nonlinear applications.

Health Insurance Coverage Projections For The US Population And Sources Of Coverage, By Age, 2024-34.

In the Congressional Budget Office's projections of health insurance coverage, 92.3 percent of the US population, or 316 million people, have coverage in 2024, and 7.7 percent, or 26 million, are uninsured. The uninsured share of the population will rise over the course of the next decade, before settling at 8.9 percent in 2034, largely as a result of the end of COVID-19 pandemic-related Medicaid policies, the expiration of enhanced subsidies available through the Affordable Care Act health insurance Marketplaces, and a surge in immigration that began in 2022. The largest increase in the uninsured population will be among adults ages 19-44. Employment-based coverage will be the predominant source of health insurance, and as the population ages, Medicare enrollment will grow significantly. After greater-than-expected enrollment in 2023, Marketplace enrollment is projected to reach an all-time high of twenty-three million people in 2025.

Decoupled evolution of the Sex Peptide gene family and Sex Peptide Receptor in Drosophilidae.

Across internally fertilising species, males transfer ejaculate proteins that trigger wide-ranging changes in female behaviour and physiology. Much theory has been developed to explore the drivers of ejaculate protein evolution. The accelerating availability of high-quality genomes now allows us to test how these proteins are evolving at fine taxonomic scales. Here, we use genomes from 264 species to chart the evolutionary history of Sex Peptide (SP), a potent regulator of female post-mating responses in Drosophila melanogaster. We infer that SP first evolved in the Drosophilinae subfamily and has followed markedly different evolutionary trajectories in different lineages. Outside of the Sophophora-Lordiphosa, SP exists largely as a single-copy gene with independent losses in several lineages. Within the Sophophora-Lordiphosa, the SP gene family has repeatedly and independently expanded. Up to seven copies, collectively displaying extensive sequence variation, are present in some species. Despite these changes, SP expression remains restricted to the male reproductive tract. Alongside, we document considerable interspecific variation in the presence and morphology of seminal microcarriers that, despite the critical role SP plays in microcarrier assembly in D. melanogaster, appear to be independent of changes in the presence/absence or sequence of SP. We end by providing evidence that SP's evolution is decoupled from that of its receptor, SPR, in which we detect no evidence of correlated diversifying selection. Collectively, our work describes the divergent evolutionary trajectories that a novel gene has taken following its origin and finds a surprisingly weak coevolutionary signal between a supposedly sexually antagonistic protein and its receptor.

The evolution of sex peptide: sexual conflict, cooperation, and coevolution.

A central paradigm in evolutionary biology is that the fundamental divergence in the fitness interests of the sexes ('sexual conflict') can lead to both the evolution of sex-specific traits that reduce fitness for individuals of the opposite sex, and sexually antagonistic coevolution between the sexes. However, clear examples of traits that evolved in this way - where a single trait in one sex demonstrably depresses the fitness of members of the opposite sex, resulting in antagonistic coevolution - are rare. The Drosophila seminal protein 'sex peptide' (SP) is perhaps the most widely cited example of a trait that appears to harm females while benefitting males. Transferred in the ejaculate by males during mating, SP triggers profound and wide-ranging changes in female behaviour and physiology. Early studies reported that the transfer of SP enhances male fitness while depressing female fitness, providing the foundations for the widespread view that SP has evolved to manipulate females for male benefit. Here, we argue that this view is (i) a simplification of a wider body of contradictory empirical research, (ii) narrow with respect to theory describing the origin and maintenance of sexually selected traits, and (iii) hard to reconcile with what we know of the evolutionary history of SP's effects on females. We begin by charting the history of thought regarding SP, both at proximate (its production, function, and mechanism of action) and ultimate (its fitness consequences and evolutionary history) levels, reviewing how studies of SP were central to the development of the field of sexual conflict. We describe a prevailing paradigm for SP's evolution: that SP originated and continues to evolve to manipulate females for male benefit. In contrast to this view, we argue on three grounds that the weight of evidence does not support the view that receipt of SP decreases female fitness: (i) results from studies of SP's impact on female fitness are mixed and more often neutral or positive, with fitness costs emerging only under nutritional extremes; (ii) whether costs from SP are appreciable in wild-living populations remains untested; and (iii) recently described confounds in genetic manipulations of SP raise the possibility that measures of the costs and benefits of SP have been distorted. Beyond SP's fitness effects, comparative and genetic data are also difficult to square with the idea that females suffer fitness costs from SP. Instead, these data - from functional and evolutionary genetics and the neural circuitry of female responses to SP - suggest an evolutionary history involving the evolution of a dedicated SP-sensing apparatus in the female reproductive tract that is likely to have evolved because it benefits females, rather than harms them. We end by exploring theory and evidence that SP benefits females by functioning as a signal of male quality or of sperm receipt and storage (or both). The expanded view of the evolution of SP that we outline recognises the context-dependent and fluctuating roles played by both cooperative and antagonistic selection in the origin and maintenance of reproductive traits.

Evolution of sexual development and sexual dimorphism in insects.

Most animal species consist of two distinct sexes. At the morphological, physiological, and behavioral levels the differences between males and females are numerous and dramatic, yet at the genomic level they are often slight or absent. This disconnect is overcome because simple genetic differences or environmental signals are able to direct the sex-specific expression of a shared genome. A canonical picture of how this process works in insects emerged from decades of work on Drosophila. But recent years have seen an explosion of molecular-genetic and developmental work on a broad range of insects. Drawing these studies together, we describe the evolution of sexual dimorphism from a comparative perspective and argue that insect sex determination and differentiation systems are composites of rapidly evolving and highly conserved elements.

Structural variation in Drosophila melanogaster spermathecal ducts and its association with sperm competition dynamics.

The ability of female insects to retain and use sperm for days, months, or even years after mating requires specialized storage organs in the reproductive tract. In most orders, these organs include a pair of sclerotized capsules known as spermathecae. Here, we report that some Drosophila melanogaster females exhibit previously uncharacterized structures within the distal portion of the muscular duct that links a spermatheca to the uterus. We find that these 'spermathecal duct presences' (SDPs) may form in either or both ducts and can extend from the duct into the sperm-storing capsule itself. We further find that the incidence of SDPs varies significantly between genotypes, but does not change significantly with the age or mating status of females, the latter indicating that SDPs are not composed of or stimulated by sperm or male seminal proteins. We show that SDPs affect neither the number of first male sperm held in a spermatheca nor the number of offspring produced after a single mating. However, we find evidence that SDPs are associated with a lack of second male sperm in the spermathecae after females remate. This raises the possibility that SDPs provide a mechanism for variation in sperm competition outcome among females.

A Systematic Review of Complications Following Minimally Invasive Spine Surgery Including Transforaminal Lumbar Interbody Fusion.

PURPOSE OF REVIEW: To assess complications after minimally invasive spinal surgeries including transforaminal lumbar interbody fusion (MI-TLIF) by reviewing the most recent literature. RECENT FINDINGS: Current literature demonstrates that minimally invasive surgery (MIS) in spine has improved clinical outcomes and reduced complications when compared with open spinal procedures. Recent studies describing MI-TLIF primarily for degenerative disk disease, spondylolisthesis, and vertebral canal stenosis cite over 89 discrete complications, with the most common being radiculitis (ranging from 2.8 to 57.1%), screw malposition (0.3-12.7%), and incidental durotomy (0.3-8.6%). Minimally invasive spine surgery has a distinct set of complications in comparison with other spinal procedures. These complications vary based on the exact MIS procedure and indication. The most frequently documented MI-TLIF complications in current published literature were radiculitis, screw malposition, and incidental durotomy.

Seminal fluid.

Seminal fluid does more than transport sperm. Hopkins et al., describe the diverse features and functions of seminal fluid, and its role in evolution and medicine.