Biology subjects

Krauchunas, A. R.

Publications and source records attributed to Krauchunas, A. R..

7 recordsLinked to original sources

bioRxiv2026

Characterization of spe-40/Fam187 identifies a deeply conserved sperm protein at the C. elegans fertilization synapse

Fertilization is the process in which two specialized cells, the sperm and egg, interact, adhere, and fuse their membranes. This occurs in all sexually reproducing organisms. Several transmembrane and secreted proteins have been shown to be required for fertilization. Genetic mutations can alter these proteins and disrupt fertilization, leading to reduced or no offspring. When fertilization-specific sperm proteins are mutated, sperm production, motility, and activation are unaffected, but the sperm lose the ability to successfully fertilize an egg. In this study, we report on the sperm-specific protein SPE-40/FAM187, which is a single-pass transmembrane protein with an immunoglobulin-like domain. When spe-40 is mutated in C. elegans the animals are severely sub-fertile due to a sperm-specific defect. All the characteristics of the sperm that we have evaluated in the mutant are normal, yet sperm lacking SPE-40 do not fertilize. SPE-40 has orthologs in other species, including humans. Thus, we have established a role for SPE-40/FAM187 in fertilization that suggests it represents a conserved component of the fertilization synapse.

developmental biology↗

bioRxiv2025

The Caenorhabditis elegans spe-21 gene that encodes a palmitoyltransferase is necessary for spermiogenesis.

In most animals, spermatids that are produced must further differentiate into fertilization-competent spermatozoa after completing meiosis. In Caenorhabditis elegans, this process is known as spermiogenesis or spermatid activation and it results in the transformation of round, non-motile spermatids into amoeboid, motile spermatozoa. Spermatid activation in C. elegans is also associated with the fusion of Golgi-derived vesicles called the membranous organelles (MOs) with the plasma membrane. This fusion process is required for producing a fertilization-competent surface on the sperm by placing MO-carried, resident proteins onto the spermatozoon membrane. We have identified, cloned, and characterized the role of spe-21, also designated as dhhc-5, during both hermaphrodite and male spermatid activation. spe-21 null mutant worms are severely sub-fertile at all studied growth temperatures. spe-21 mutant spermatids fail to activate either in vivo or in vitro after treatment with known chemical activators. We have found that spe-21 is necessary for MO fusion and pseudopod formation in spermatids. The spe-21 gene encodes a predicted four pass transmembrane protein with a conserved Asp-His-His-Cys (DHHC) tetrapeptide zinc finger motif embedded in a cysteine rich region (DHHC-CRD type zinc finger motif). Generally, proteins with DHHC-CRD motifs catalyze the post-translational addition of palmitate to their protein substrates and are called palmitoyltransferases or palmitoylacyltransferases (PATs). We also found that mNeonGreen-tagged SPE-21 localizes to the MOs in spermatids. Together, our findings show that SPE-21 is an MO localized palmitoyltransferase required for proper spermatid activation and creation of fertilization-competent spermatozoa.

developmental biology↗

bioRxiv2021

Tracking extracellular vesicle (EV) cargo as a platform for studying EVomics, signaling, and targeting in vivo

Extracellular vesicle (EV)-based signaling is a challenge to study, due to EV small size, heterogeneity, and limited information on cargo content in vivo. We present Caenorhabditis elegans as a discovery platform that allows single EV tracking from source to target tissue in living animals. We enriched ciliary EVs using GFP-tagged PKD-2 cargo followed by mass spectrometry analysis to identify 2,888 cargo candidates. By integrating our dataset with single-cell transcriptomic data, we identified EV cargo produced by individual neurons and other cell and tissue types. A single cilium produces multiple EVs with distinct protein content. Ciliary EVs carry nucleic acid binding proteins. We observed transfer of EV cargo from the male reproductive tract to the hermaphrodite uterus during mating, a direct demonstration of animal-to-animal EV targeting. One-Sentence SummaryHere we present a discovery platform for studying animal extracellular vesicle composition and biogenesis.

cell biology↗

bioRxiv2021

SPE-51, a sperm secreted protein with an Immunoglobulin-like domain, is required for sperm-egg fusion in C. elegans

Despite the importance of fertilization, the molecular basis of sperm-egg interaction is not well understood. In a forward genetics screen for fertility mutants in Caenorhabditis elegans we identified spe-51. Mutant worms make sperm that are unable to fertilize the oocyte but otherwise normal by all available measurements. The spe-51 gene encodes a secreted protein that includes an immunoglobulin (Ig)-like domain and a hydrophobic sequence of amino acids. The SPE-51 protein acts cell-autonomously and localizes to the surface of the spermatozoa. This is the first example of a secreted protein required for the interactions between the sperm and egg with genetic validation for a specific function in fertilization. Our analyses of these genes begin to build a paradigm for sperm-secreted or reproductive tract-secreted proteins that coat the sperm surface and influence their survival, motility, and/or the ability to fertilize the egg.

developmental biology↗

bioRxiv2021

The EGF-motif containing protein SPE-36 is a secreted protein required for sperm function at fertilization in C. elegans

The growing number of genes specifically required for fertilization suggests that there is a significant amount of molecular complexity at the sperm-egg interface. Thus, we have adopted a model of a "fertilization synapse" where specialized zones of interaction and multi-protein complexes mediate gamete interaction and fusion. The fertilization synapse is likely to be composed of both trans and cis protein-protein interactions at the surface of each gamete. Mutations in the Caenorhabditis elegans spe-36 gene result in a sperm-specific fertility defect. Surprisingly, spe-36 encodes a secreted EGF-motif containing protein that functions cell autonomously. Despite the fact that morphology and migratory behavior of spe-36 sperm are indistinguishable from wild-type sperm, spe-36 sperm make close contact with oocytes but fail to fertilize them. The genetic requirement for a secreted sperm-derived protein for fertilization is novel and represents a paradigm-shifting discovery in the molecular understanding of fertilization.

developmental biology↗

bioRxiv2017

Maternal proteins that are phospho-regulated upon egg activation include crucial factors for oogenesis, egg activation and embryogenesis in Drosophila melanogaster

Egg activation is essential for the successful transition from a mature oocyte to a developmentally competent egg. It consists of a series of events including the resumption and completion of meiosis, initiation of translation of some maternal mRNAs and destruction of others, and changes to the vitelline envelope. This drastic change of cell state is accompanied by large scale alteration of the phospho-proteome of the cell. Despite the importance of this transition in cell and developmental state, it has been difficult to find many of its regulators. We hypothesize that phosphorylation state changes between oocyte and early embryo regulate the activities of proteins that are necessary during or after this transition, and thus that the set of phospho-regulated proteins would be an enriched source for finding critical players in the egg-to-embryo transition. To test this, we used germline-specific RNAi to examine the function of 189 maternal proteins that are phospho-regulated during egg activation in Drosophila melanogaster. We identified 53 genes whose knockdown reduced or abolished egg production, as well as 50 genes for which maternal knockdown led to significant impairment or abolishment of the eggs ability to hatch (hatchability). We observed different stages of developmental arrest in the embryos with impaired hatchability and several distinct categories of abnormalities in the maternal knockdown embryos that arrest early in development, suggesting potential roles of the candidates in egg activation and early embryogenesis. Our results, validated by our detection of multiple genes with previously-documented maternal-effect phenotypes among the proteins we tested, revealed 15 genes with new roles in egg activation and early embryogenesis. Given that protein phospho-regulation also occurs during this transition in other organisms, we suggest that the phospho-regulated proteins may provide an enriched dataset for identifying important players in the egg-to-embryo transition.

developmental biology↗

bioRxiv2017

spe-43 is required for sperm activation in C. elegans

Successful fertilization requires that sperm are activated prior to contacting an oocyte. In C. elegans, this activation process, called spermiogenesis, transforms round immobile spermatids into motile, fertilization-competent spermatozoa. We describe the phenotypic and genetic characterization of spe-43, a new component of the spe-8 pathway, which is required for spermiogenesis in hermaphrodites; spe-43 hermaphrodites are self-sterile, while spe-43 males show wild-type fertility. When exposed to Pronase to activate sperm in vitro, spe-43 spermatids form long rigid spikes radiating outward from the cell periphery instead of forming a motile pseudopod, indicating that spermiogenesis initiates but is not completed. Using a combination of recombinant and deletion mapping and whole genome sequencing, we identified F09E8.1 as spe-43. SPE-43 is predicted to exist in two isoforms; one isoform appears to be a single-pass transmembrane protein while the other is predicted to be a secreted protein. SPE-43 can bind to other known sperm proteins, including SPE-4 and SPE-29, which are known to impact spermiogenesis. In summary, we have identified a membrane protein that is present in C. elegans sperm and is required for sperm activation via the hermaphrodite activation signal.

developmental biology↗