Biology subjects

Krsticevic, F.

Publications and source records attributed to Krsticevic, F..

9 recordsLinked to original sources

bioRxiv2026

The Y chromosome gene draupnir reveals constraints on engineering Y-linked sex-ratio distorters in malaria mosquitoes

Y chromosome sex distorters offer a powerful strategy for mosquito population suppression, but their implementation is constrained by the transcriptional silencing of Y-linked transgenes during spermatogenesis. To investigate how endogenous Y genes evade this repression, we characterized draupnir (formerly YG5), a multicopy Y-linked gene of Anopheles gambiae encoding a Zip3-like meiotic protein. Comparative genomic and phylogenetic analyses revealed that draupnir originated through duplication of the autosomal paralog skirnir, itself derived from the ancestral recombination factor vilya, followed by amplification into a tandem array on the Y chromosome. We show that draupnir is actively transcribed during meiosis, making it the only known Y-linked gene expressed during mosquito spermatogenesis. To test whether its regulatory region is sufficient to confer sperm-specific expression from the Y chromosome, we cloned the draupnir promoter to drive expression of an X-chromosome shredder. When inserted on an autosome, the construct drove meiotic expression and strong sex-ratio distortion in the progeny of transgenic males. In contrast, the same construct inserted on the Y chromosome was transcriptionally silent and produced balanced sex ratios. These results demonstrate that draupnir expression depends on its native genomic context within a multicopy Y-linked array and that its promoter alone cannot overcome transcription silencing. Our findings reveal a fundamental constraint on Y-chromosome-based genetic control strategies and point to future approaches for enabling transcription from otherwise repressed sex chromosomes.

genetics↗

bioRxiv2025

Post-Domestication selection of MKK3 Shaped Seed Dormancy and End-Use Traits in Barley

Anthropogenic selection of grain traits such as dormancy has shaped the developmental trajectories of crop plants (1). In cereals, shortening dormancy provides rapid and even post-harvest germination, but increases the risk of weather-induced pre-harvest sprouting (PHS) with harvest losses estimated at beyond 1 billion USD a year (2, 3, 4). Our understanding of how, why, when and where diversification of cereal dormancy arose is fragmentary. Here, we show in the founder cereal crop barley (Hordeum vulgare) that the Mitogen-Activated Protein Kinase (MAPK) pathway regulates dormancy primarily through a mosaic of locus haplotypes comprising copy-number variation and inherent kinase activity of Mitogen-activated protein kinase kinase 3 (MKK3). We provide evidence supporting the historical selection of specific MKK3 haplotypes that shape dormancy levels according to changing climatic pressures. Understanding the regulatory landscape of MKK3 provides a genetic framework to balance short grain dormancy and PHS-avoidance during a period of rapid climate change.

plant biology↗

bioRxiv2025

Comparative chemosensory mechanisms underlying larval foraging and competitive advantage in Aedes albopictus and Aedes aegypti

The invasive Asian tiger mosquito, Aedes albopictus (Skuse), and yellow fever mosquito, Aedes aegypti (L.) are known to compete for resources during the larval stage, often resulting in the ecological displacement of Ae. aegypti by Ae. albopictus. The chemosensory system plays a pivotal role in larval foraging behavior and may contribute to the competitive advantage. Here, we employed comparative transcriptomics and functional characterization of odorant receptors (ORs) to investigate species-specific differences in larval olfaction. Notably, we uncovered functional variation within the conserved olfactory indole receptor clade, indicating distinct ecological adaptations across species and life stages. We also developed a novel approach to functionally characterize the larval sensory cone and mapped its receptor neuron projections to two key brain regions: the antennal lobe and the subesophageal ganglion. This study provides new insights into the molecular and neural basis of chemosensory-driven behavior in mosquito larvae and highlights the potential role of olfaction in shaping interspecies competition and ecological success.

neuroscience↗

bioRxiv2025

Mosquito Sex Separation using Complementation of Selectable Traits and Engineered Neo-Sex Chromosomes

Effective and scalable sex separation remains a critical challenge for mosquito genetic control strategies. Genetic sexing strains (GSS) address this by genetically linking maleness with selectable traits, enabling efficient removal of females before release. Here, we describe a robust platform for the development of GSSs in the invasive Aedes albopictus mosquito by integrating a CRISPR-engineered selectable phenotype with sex conversion via nix, the male-determining factor. As a proof-of-concept, we disrupt the yellow gene to generate a vivid pigmentation marker, then rescue its function in males using nix-containing transgenes, creating a stable strain where all females are yellow and all engineered males are dark. The resulting GSS males are fertile, robust, and despite lacking the ancestral M locus, exhibit gene expression profiles closely resembling wild-type males. We benchmark sex separation based on pigmentation and discover that yellow mutant females exhibit slower larval development, enhancing protandry-based sorting. The GSS strain is compatible with existing size-based sex sorting systems, allowing for improved separation accuracy through the integration of natural and engineered sexually dimorphic traits. Additionally, we find that GSS females lay desiccation-sensitive eggs, reducing the risk of accidental female releases. Our approach is the first to engineer a sex-linked selectable trait by precisely targeting an endogenous gene and restoring its function in males, establishing a versatile platform for GSS development in Aedes mosquitoes.

synthetic biology↗

bioRxiv2023

Revisiting Y-chromosome detection methods: R-CQ and KAMY efficiently identify Y chromosome sequences in Tephritidae insect pests

The detection and characterization of sex chromosome sequences is particularly important for major pest families, like the Tephritidae, whereas alternative pest management approaches, mainly involving male-only release programs, rely on the ability to target and manipulate sex-specific genomic regions, particularly those of the Y chromosome. However, resolving and detecting X and Y chromosome sequences at the chromosome level requires careful consideration of algorithmic outputs, especially in species where extensive sex chromosome markers are not available. Here, we present R-CQ and KAMY, two computational methods developed for the detection of sex chromosome-linked sequences. We evaluate their performance on newly generated chromosome-level assemblies of four important Tephritid pest species: Ceratitis capitata, Bactrocera dorsalis, Bactrocera zonata and Anastrepha ludens. By combining algorithmic predictions with a manual curation process, we assess the strengths and limitations of each method and provide a robust dataset of curated X- and Y-linked sequences. Overall, our results establish a framework for studying poorly characterized sex chromosome lineages and identifying sex-specific genomic regions, supporting the broader development of sex chromosome-based pest managements systems.

genomics↗

bioRxiv2023

Targeting mosquito X-chromosomes reveals complex transmission dynamics of sex ratio distorting gene drives

Engineered sex ratio distorters (SRDs) have been proposed as a powerful component of genetic control strategies designed to suppress harmful insect pests. Two types of CRISPR-based SRD mechanisms have been proposed: X-shredding eliminates X-bearing sperm, while X-poisoning eliminates daughters inheriting disrupted X-chromosomes. These differences can have a profound impact on the population dynamics of SRDs when linked to the Y-chromosome: an X-shredder is invasive constituting a classical meiotic Y-drive, whereas X-poisoning is self-limiting, unable to invade but also insulated from selection. Here, we established the first X-poisoning strains in the malaria vector Anopheles gambiae targeting three X-linked genes during spermatogenesis resulting in male bias. Surprisingly, we found that sex distortion was primarily driven by a loss of X-bearing sperm with limited evidence for postzygotic lethality of daughters. By leveraging a Drosophila melanogaster model, we show unambiguously that engineered SRD traits can operate differently in these two insects. Unlike X-shredding, X-poisoning could theoretically operate at early stages of spermatogenesis. We therefore explored premeiotic Cas9 expression to target the mosquito X-chromosome. We find that, by pre-empting the onset of meiotic sex chromosome inactivation, this approach may enable the development of Y-linked SRDs if mutagenesis of spermatogenesis-essential genes is functionally balanced.

synthetic biology↗

bioRxiv2023

Gene drive and genetic sex conversion in the global agricultural pest Ceratitis capitata

Homing-based gene drives are novel interventions promising the area-wide, species-specific genetic control of harmful insect populations. Here we characterise a first set of gene drives in a tephritid agricultural pest species, the Mediterranean fruit fly Ceratitis capitata (medfly). Our results show that the medfly is highly amenable to homing-based gene drive strategies. By targeting the medfly transformer gene, we also demonstrate two different mechanisms by which CRISPR-Cas9 gene drive can be coupled to sex conversion, whereby genetic females are transformed into fertile and harmless XX males. Given this unique malleability of sex determination, we modelled gene drive interventions that couple sex conversion and female sterility and found that such approaches could be effective and tolerant of resistant allele selection in the target population. Our results open the door for developing gene drive strains for the population suppression of the medfly and related tephritid pests by co-targeting female reproduction and shifting the reproductive sex ratio towards males. They demonstrate the untapped potential for gene drives to tackle agricultural pests in an environmentally friendly and economical way.

synthetic biology↗

bioRxiv2020

Engineered sex distortion in the global agricultural pest Ceratitis capitata

Genetic sex ratio distorters have potential for the area-wide control of harmful insect populations. Endonucleases targeting the X-chromosome and whose activity is restricted to male gametogenesis have recently been pioneered as a means to engineer such traits. Here we enabled endogenous CRISPR/Cas9 and CRISPR/Cas12a activity during spermatogenesis of the Mediterranean fruit fly Ceratitis capitata, a worldwide agricultural pest of extensive economic significance. In the absence of a chromosome-level assembly, we analysed long and short-read genome sequencing data from males and females to identify two clusters of abundant and X-chromosome specific sequence repeats. When targeted by gRNAs in conjunction with Cas9 they yielded a significant and consistent distortion of the sex ratio in independent transgenic strains and a combination of distorters induced a strong bias towards males (~80%). Our results demonstrate the design of sex distorters in a non-model organism and suggest that strains with characteristics suitable for field application could be developed for a range of medically or agriculturally relevant insect species.

synthetic biology↗

bioRxiv2020

Improved reference genome of the arboviral vector Aedes albopictus

The Asian tiger mosquito Aedes albopictus is globally expanding and has become the main vector for human arboviruses in Europe. Here we present AalbF2, a dramatically improved assembly of the Ae. albopictus genome that has revealed widespread viral insertions, novel microRNAs and piRNA clusters, the sex determining locus, new immunity genes, and has enabled genome-wide studies of geographically diverse Ae. albopictus populations and analyses of the developmental and stage-dependent network of expression data. Additionally, we built the first physical map for this species with 75% of the assembled genome anchored to the chromosomes. These up-to-date resources of the genome provide a foundation to improve understanding of the adaptation potential and the epidemiological relevance of this species and foster the development of innovative control measures. One Sentence SummaryLong-read and Hi-C-based de novo assembly of the arboviral vector Aedes albopictus genome fosters deeper understanding of its biological features.

genomics↗