Germ granule analysis reveals conserved and diverse features among Drosophila species
Authors: Dominique Doyle; Bianca Ulrich; Bianca Ortega; Matthew Niepielko
Affiliation: Kean University
Keywords: l. evo-devo; k. RNA transport and localization
The co-packaging of different mRNA types into macromolecular structures called ribonucleoproteins (RNPs) is a conserved strategy for the regulation of mRNA metabolism. In many animals, the formation of complex RNPs called germ granules is essential for the post-transcriptional regulation of mRNAs that are required for germline development, maintenance, and function. In Drosophila, germ granules are assembled at the posterior of the egg and are inherited by the primordial germ cells during embryogenesis. In D. melanogaster, mRNAs accumulate in germ granules by forming homotypic clusters, which are distinct aggregates that contain multiple copies of a specific mRNA type. Homotypic clusters in D. melanogaster are generated through a stochastic seeding and self-recruitment process, relying on cis-regulatory sequences found in the 3′UTR of germ granule mRNAs called "clustering elements." We hypothesize that clustering elements may be susceptible to evolutionary changes, creating diversity in the abundance of mRNAs found in germ granules from different Drosophila species. To test our hypothesis, we first investigated the homotypic clustering of two germ granule mRNAs, nanos (nos) and polar granule component (pgc) in three Drosophila species. By combining single molecule in situ hybridization (smFISH), super-resolution microscopy, and quantitative image analysis, we found that seeding and self-recruitment is a conserved process that generates homotypic clusters to enrich germ granules with mRNAs. Interestingly, we found that the mRNA content of nos and pgc homotypic clusters, as measured by absolute transcript number, were strikingly diverse among Drosophila species. Specifically, a 50% difference in the number of transcripts in nos and/or pgc homotypic clusters was discovered between D. melanogaster, D. virilis, and D. pseudoobscura. By employing computational modeling, we recreated the diversity in germ granule mRNA content from all three species. Our simulations suggest that a combination of factors, including differences in mRNA clustering efficacy, generates germ granule diversity. Currently, we are investigating if variability found in clustering elements underlies Drosophila germ granule diversity.