Biology subjects

Brill, J. A.

Publications and source records attributed to Brill, J. A..

6 recordsLinked to original sources

bioRxiv2026

In Step, the sole Drosophila TENT5 poly(A) polymerase, is required for spermatid elongation and individualization

Cytoplasmic poly(A) tail length is a key regulator of mRNA translation and stability, controlled in part by non-canonical poly(A) polymerases of the TENT5 family, which are required for gametogenesis in mice. We characterize the sole Drosophila melanogaster TENT5 ortholog, which we name in step (isep). Although identified as a candidate target of the eye specification factor Eyeless, isep mutants have normal eyes; instead, isep is essential for male fertility. The isep gene and its protein are expressed in spermatocytes and in elongating spermatids and are concentrated at the distal end of spermatid cysts. Mutant spermatids fail to elongate normally and to individualize. Whole-testis poly(A) tail sequencing revealed that, unexpectedly for the loss of a poly(A) polymerase, median tail length increased in mutants, with lengthened transcripts enriched for roles in axoneme assembly. Both wild-type and a predicted catalytically inactive Isep rescued sterility. We propose that Isep controls localized translation of axonemal transcripts at the spermatid tip, largely independently of its poly(A) polymerase activity.

developmental biology↗

bioRxiv2026

A genetic screen to identify novel interactions of the phosphatidylinositol phosphate signaling pathway in Drosophila spermatogenesis

Phosphatidylinositol (PI) phosphates (PIPs) are membrane lipids with crucial roles in intracellular signaling, membrane trafficking and cytoskeletal organization. To perturb PIP function in Drosophila melanogaster sperm development, we previously expressed the Salmonella PIP phosphatase SigD (also known as SopB) under control of a spermatocyte- specific promoter. SigD expression results in reduced levels of plasma membrane PI 4,5-bisphosphate [PI(4,5)P2; henceforth PIP2] and male sterility. Transgenic lines expressing high levels of SigD (SigD-high) exhibit defects in spermatocyte cytokinesis, ciliary transition zone maturation and spermatid elongation, whereas lines expressing low levels of SigD (SigD-low) have defects in nuclear shaping and spermatid cell polarization. To further investigate the role of PIPs during Drosophila sperm development, we conducted a sensitized genetic screen to identify suppressors and enhancers of SigD-low phenotypes. From a screen of 760 chromosomal deficiencies spanning 83.6% of chromosomes 2 and 3, we identified 80 interacting regions that were further narrowed down to 40 individual suppressors. Among the strongest interactors were genes involved in microtubule dynamics, nucleocytoplasmic transport and membrane trafficking. The novel genetic interactions identified by our screen pave the way for further investigation into molecular mechanisms underlying the affected aspects of sperm development.

genetics↗

bioRxiv2025

The C-terminal ZZ domain of the Drosophila ORB2 RNA-binding protein is required for spermatid individualization

ORB2 is the Drosophila ortholog of the human CPEB2-4 family of RNA-binding proteins, which include a conserved C-terminal Zinc-binding ( ZZ) domain. We have recently shown that this domain interacts with several translation co-repressors in the early embryo, and that deletion of this domain from the endogenous orb2 gene results in derepression of its target mRNAs (LOW et al. 2025). Here we assess the effect of deletion of the ZZ domain on spermatogenesis. We find that ORB2{Delta}ZZ protein is no longer localized to the distal tip of spermatids; that in orb2{Delta}ZZ testes, additional proteins known to play a role in spermatid individualization - ORB, IMP, SOTI - are mislocalized; that the SOTI-dependent Cleaved Caspase 3 gradient no longer forms; that individualization complexes are defective; and that orb2{Delta}ZZ flies are sterile and lack mature sperm.

genetics↗

bioRxiv2023

Mutual dependence of Osbp and PI4KII in the maturation of regulated secretory granules

Secretory granules (SGs) are crucial for normal animal physiology due to their role in regulated exocytosis of biologically active molecules. SG membranes are enriched in phosphatidylinositol 4-phosphate (PI4P) and cholesterol, and previous studies suggest lipid composition is important for SG biogenesis and function. Nonetheless, the molecular details of how lipids are regulated during SG biogenesis remain poorly understood. Here, we identify Oxysterol binding protein (Osbp) as a novel regulator of SG biogenesis in a Drosophila model. We show Osbp expression level positively correlates with SG size and that Osbp requires type II phosphatidylinositol 4-kinase (PI4KII) to increase SG size. Moreover, Osbp is needed for proper PI4KII and PI4P distribution, autophagic resolution and formation of cholesterol-rich endosomal tubules that are positive for PI4KII. Feeding larvae food supplemented with sterol leads to partial suppression of SG size and PI4P distribution defects in Osbp mutants. Our results indicate that reciprocal regulation of Osbp and PI4KII drives formation of membrane tubules that mediate SG maturation through elevating PI4P levels on SG membranes. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=200 SRC="FIGDIR/small/551178v1_ufig1.gif" ALT="Figure 1"> View larger version (67K): org.highwire.dtl.DTLVardef@19566fborg.highwire.dtl.DTLVardef@19beeadorg.highwire.dtl.DTLVardef@52e048org.highwire.dtl.DTLVardef@1029b3c_HPS_FORMAT_FIGEXP M_FIG C_FIG HighlightsO_LIOsbp is needed for formation of PI4KII-positive endosomal tubules that enrich secretory granule membranes with PI4P and facilitate granule maturation. C_LIO_LIReciprocally, PI4KII is needed for Osbp-mediated secretory granule growth in salivary gland cells. C_LIO_LIChanges in Osbp expression levels alter autophagy initiation and autophagosome resolution in salivary gland cells. C_LIO_LIPI4KII-positive endosomal tubules are enriched in sterols, and sterol feeding suppresses defects caused by loss of Osbp. C_LI

cell biology↗

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

Eukaryotic translation initiation factor eIF4E-5 is required for spermiogenesis in Drosophila melanogaster

Drosophila sperm development is characterized by extensive post-transcriptional regulation whereby thousands of transcripts are preserved for translation during later stages. A key step in translation initiation is the binding of eukaryotic initiation factor 4E (eIF4E) to the 5 mRNA cap. Drosophila has multiple paralogs of eIF4E, including four (eIF4E-3, -4, -5, and -7) that are highly expressed in the testis. Other than eIF4E-3, none of these has been characterized genetically. Here, using CRISPR/Cas9 mutagenesis, we determined that eIF4E-5 is essential for male fertility. eIF4E-5 mutants exhibit defects during post-meiotic stages, including a fully penetrant defect in individualization, resulting in failure to produce mature sperm. eIF4E-5 protein localizes to the distal ends of elongated spermatid cysts, where it regulates non-apoptotic caspase activity during individualization by promoting local accumulation of the E3 ubiquitin ligase inhibitor Soti. eIF4E-5 mutants also have mild defects in spermatid cyst polarization, similar to mutants affecting the cytoplasmic polyadenylation-element binding protein Orb2 and atypical protein kinase C (aPKC). Our results further extend the diversity of non-canonical eIF4Es that carry out distinct spatiotemporal roles during spermatogenesis. Summary StatementThe testis-enriched translation initiation factor eIF4E-5 is needed for spermatid cyst polarization, individualization of mature sperm and male fertility in Drosophila.

developmental biology↗