Biology subjects

White-Cooper, H.

Publications and source records attributed to White-Cooper, H..

7 recordsLinked to original sources

bioRxiv2026

Single cell RNA sequencing of D. pseudoobscura testes reveals transcriptional signatures of heteromorphic spermatogenesis

Sperm heteromorphy, the production of multiple sperm morphs with distinct functions, has evolved repeatedly in animals but its developmental and molecular basis remains poorly understood. In the Drosophila obscura group, males produce fertilising eusperm and non-fertilising parasperm, yet when and how these lineages diverge is unclear. Here, we applied single-cell RNA sequencing to whole testes of D. pseudoobscura to resolve cell-type composition, developmental trajectories, and transcriptional differences between sperm morphs. Analysis of [~]6,500 cells across two replicates identified germline and somatic cell populations and reconstructed spermatogenesis from stem cells to elongated spermatids. We identified a bifurcation in the germline trajectory at the early spermatocyte stage, corresponding to eusperm and parasperm lineages, which was maintained through spermiogenesis. Euspermatocytes, destined to generate longer eusperm, exhibited higher transcriptional activity than paraspermatocytes. Differentially expressed genes included structural sperm tail components, chromatin condensation proteins, post-meiotically transcribed genes and gene duplications resulting in paralogues with reciprocal expression patterns. We also identified markers of somatic cyst cells, including head and tail cyst cells, but found no evidence of morph-specific specialisation within the cyst cell lineage. The transcriptional activities of the distinct germline trajectories corresponded with differential expression of orthologues of critical transcriptional regulators known in D. melanogaster. The transcriptional activator TGIF was enriched in euspermatocytes, while the transcriptional repressor kmg was upregulated in paraspermatocytes. GFP-tagged Kmg showed higher abundance and greater chromatin localisation in paraspermatocytes. These findings demonstrate that sperm morph identity is first apparent in early primary spermatocytes and is largely germline intrinsic. We propose that differential transcriptional provisioning and developmental timing underpin sperm length, providing a framework for understanding the evolution and regulation of sperm heteromorphy.

developmental biology↗

bioRxiv2025

CLUSTERING SPERM: A STATISTICAL APPROACH TO IDENTIFY SPERM MORPH NUMBERS IN THE DROSOPHILA OBSCURA SPECIES GROUP

The Drosophila obscura species group is sperm heteromorphic. Sperm heteromorphism is production of multiple sperm types, or morphs, within a single male. In the case of the obscura species, males produce two or three sperm morphs of different sizes, simultaneously, within the same testis, and throughout their lifetime. A long sperm morph, the eusperm is fertile, whereas shorter sperm morphs, the parasperm, are non-fertilising and protect the eusperm within the female reproductive tract. Many studies over the past 55 years have measured sperm length of eusperm and parasperm in obscura species. One recent study found that D. pseudoobscura produces three sperm morphs, two parasperm and one eusperm, in contrast to the two morphs described in older studies of this species. Thus, the number of morphs made by some species in this clade is still in dispute. Here, we review sperm length data from previous studies and re-measure sperm of eight species including D. pseudoobscura. We used two statistical cluster analysis approaches to identify sperm morphs based on sperm and nucleus length, to test whether multiple parasperm morphs are present in more species than previously identified. We confirmed the presence of two parasperm morphs in D. pseudoobscura, and found that two closely related species, D. persimilis and D. miranda, also produce two parasperm morphs. D. affinis, D. azteca, D. bifasciata, D. guanche and D. subobscura produce a single parasperm morph, indicating that the presence of two parasperm is a derived feature in the pseudoobscura species subgroup.

zoology↗

bioRxiv2023

Differential gene expression underpinning production of distinct sperm morphs in the wax moth Galleria mellonella.

Male Lepidoptera makes two distinct sperm types; each ejaculate contains both eupyrene sperm, which can fertilise the egg, and apyrene sperm, which are not fertilisation competent. These sperm have distinct morphologies, unique functions, and different proteomes. Their production is highly regulated, however very few genes with specific roles in production of one or other morph have been described. We present the first comparative transcriptomics study of precursors of eupyrene and apyrene sperm to identify genes potentially implicated in regulating or enacting the distinct differentiation programmes. Differentially expressed genes included genes with potential roles in transcriptional regulation, cell cycle and sperm morphology. We identified gene duplications generating paralogues with functions restricted to one or other morph. However phylogenetic analysis also revealed evolutionary flexibility in expression patterns of duplicated genes between different Lepidopteran species. Improved understanding of Lepidopteran reproduction will be vital in targeting prevalent pests in agriculture, and on the flip side, ensuring the fertility and thus survival of pollinator populations in response to environmental stress.

developmental biology↗

bioRxiv2023

Transgenic expression of cif genes from Wolbachia strain wAlbB recapitulates cytoplasmic incompatibility in Aedes aegypti.

The endosymbiotic bacteria Wolbachia can invade insect populations by modifying host reproduction through cytoplasmic incompatibility (CI), an effect that results in embryonic lethality when Wolbachia-carrying males mate with Wolbachia-free females. Here we describe a transgenic system for recreating CI in the major arbovirus vector Aedes aegypti using CI factor (cif) genes from wAlbB, a Wolbachia strain currently being deployed to reduce dengue transmission. CI-like sterility was induced when cifA and cifB were co-expressed in testes; this sterility could be rescued by maternal cifA expression, thereby reproducing the pattern of Wolbachia-induced CI. Expression of cifB alone was associated with extensive DNA damage and disrupted spermatogenesis. The strength of rescue by maternal cifA expression was dependent on the comparative levels of cifA/cifB expression in males. These findings are consistent with CifB acting as a toxin and CifA as an antitoxin, with CifA attenuating CifB toxicity in both the male germline and in developing embryos.

microbiology↗

bioRxiv2022

Emergent dynamics of adult stem cell lineages from single nucleus and single cell RNA-Seq of Drosophila testes

Proper differentiation of sperm from germline stem cells, essential for production of the next generation, requires dramatic changes in gene expression that drive remodeling of almost all cellular components, from chromatin to organelles to cell shape itself. Here we provide a single nucleus and single cell RNA-seq resource covering all of spermatogenesis in Drosophila starting from in-depth analysis of adult testis single nucleus RNA-seq (snRNA-seq) data from the Fly Cell Atlas (FCA) study (Li et al., 2022). With over 44,000 nuclei and 6,000 cells analyzed, the data provide identification of rare cell types, mapping of intermediate steps in differentiation, and the potential to identify new factors impacting fertility or controlling differentiation of germline and supporting somatic cells. We justify assignment of key germline and somatic cell types using combinations of known markers, in situ hybridization, and analysis of extant protein traps. Comparison of single cell and single nucleus datasets proved particularly revealing of dynamic developmental transitions in germline differentiation. To complement the web-based portals for data analysis hosted by the FCA, we provide datasets compatible with commonly used software such as Seurat and Monocle. The foundation provided here will enable communities studying spermatogenesis to interrogate the datasets to identify candidate genes to test for function in vivo.

developmental biology↗

bioRxiv2021

Fly Cell Atlas: a single-cell transcriptomic atlas of the adult fruit fly

The ability to obtain single cell transcriptomes for stable cell types and dynamic cell states is ushering in a new era for biology. We created the Tabula Drosophilae, a single cell atlas of the adult fruit fly which includes 580k cells from 15 individually dissected sexed tissues as well as the entire head and body. Over 100 researchers from the fly community contributed annotations to >250 distinct cell types across all tissues. We provide an in-depth analysis of cell type-related gene signatures and transcription factor markers, as well as sexual dimorphism, across the whole animal. Analysis of common cell types that are shared between tissues, such as blood and muscle cells, allowed the discovery of rare cell types and tissue-specific subtypes. This atlas provides a valuable resource for the entire Drosophila community and serves as a comprehensive reference to study genetic perturbations and disease models at single cell resolution.

genomics↗

bioRxiv2019

Loss of function of the RNA export factor, Nxt1, in Drosophila causes muscle degeneration and reduced expression of genes with long introns.

The RNA export pathway is essential for export-competent mRNAs to pass from the nucleus into the cytoplasm, and thus is essential for protein production and normal function of cells. Drosophila with partial loss of function of Nxt1, a core factor in the pathway, show reduced viability and male and female sterility. The male sterility has previously been shown to be caused by defects in testis-specific gene expression, particularly of genes without introns. Here we describe a specific defect in growth and maintenance of the larval muscles, leading to muscle degeneration in Nxt1 mutants. RNAseq revealed reduced expression of mRNAs of many genes in Nxt1 mutant muscles. Despite this, the degeneration was rescued by increased expression of a single gene, the costamere component tn (abba), in muscles. Genes under-expressed in the mutant typically have long introns, and most normally encode circular RNAs in addition to mRNAs. This is the first report of a specific role for the RNA export pathway gene Nxt1 in muscle integrity. Our data on Nxt1 links the mRNA export pathway to a global role in mRNA expression of genes that also produce circular RNAs, in vivo. Author summaryIn eukaryotic cells the DNA encoding instructions for protein synthesis is located in the nucleus, it is transcribed to generate pre-mRNA, which is processed at both ends and spliced to remove internal spacer regions (introns) to generate mRNA. This mRNA is then transported by the mRNA export pathway via nuclear pores to the cytoplasm for protein synthesis. We have previously shown that reduction in activity of a specific protein in the mRNA export pathway, Nxt1, has an additional role in testis-specific transcription. Here we describe a further role for this protein specifically in gene expression, particularly of genes with long introns, and in muscle maintenance. Drosophila larvae with reduced Nxt1 activity have normal muscle pattern when they are small, but show muscular atrophy and degeneration as they grow, resulting in significant defects in their movement speed. We discovered that expression of many genes is reduced in the mutant larvae, but that restoring the expression of just one of these, abba, the Drosophila homologue of Trim32 (a human gene involved in muscular dystrophy) is capable of preventing the muscle degeneration.

developmental biology↗