Arnce, L. | bioRxiv Science

bioRxiv2025

Comparative functional and evolutionary analysis of essential germline stem cell genes across the genus Drosophila and two outgroup species

In Drosophila melanogaster, bag of marbles (bam) encodes a protein essential for germline stem cell daughter (GSC) differentiation in early gametogenesis. Despite its essential role in D. melanogaster, direct functional evaluation of bam in other closely related Drosophila species reveal this essential function is not necessarily conserved. In D. teissieri, for example, bam is not essential for GSC daughter differentiation. Here, we generated bam null alleles using CRISPR-Cas9 in a species more distantly related to D. melanogaster, D. americana, to interrogate whether bams essential GSC differentiation function is novel to the melanogaster species group or a function more basal to the Drosophila genus. To further characterize the extent of the functional flexibility of other GSC regulating genes, we generated a gene ortholog dataset for 366 GSC regulating genes essential in D. melanogaster across 15 additional Drosophila and two outgroup species. We find that bams essential GSC function is conserved between D. melanogaster and D. americana and therefore originated prior to the formation of the melanogaster species group. Additionally, we find that [~]8% of the 366 GSC genes essential in D. melanogaster are absent in at least one of the 17 species in our ortholog dataset. These results indicate that developmental systems drift (DSD), in which the specific genes regulating a function may change, but the final phenotype is retained, occurs in stem cell regulation and the production of gametes across Drosophila species. Article summaryResults from CRISPR induced bam null mutants in D. americana and comparative ortholog analysis of essential GSC regulating genes indicate that the evolutionary origin of bams essential GSC differentiation function is likely basal to the Drosophila genus, and there is functional flexibility in at least [~]8% of the 366 GSC regulating genes across the 17 included species.

genetics↗

bioRxiv2024

Comparative Analysis of Drosophila Bam and Bgcn Sequences and Predicted Protein Structural Evolution

The protein encoded by the Drosophila melanogaster gene bag of marbles (bam) plays an essential role in early gametogenesis by complexing with the gene product of benign gonial cell neoplasm (bgcn) to promote germline stem cell daughter differentiation in males and females. Here, we compared the AlphaFold2 and AlphaFold Multimer predicted structures of Bam protein and the Bam:Bgcn protein complex between D. melanogaster, D. simulans, and D. yakuba, where bam is necessary in gametogenesis to that in D. teissieri, where it is not. Despite significant sequence divergence, we find very little evidence of significant structural differences in high confidence regions of the structures across the four species. This suggests that Bam structure is unlikely to be a direct cause of its functional differences between species and that Bam may simply not be integrated in an essential manner for GSC differentiation in D. teissieri. Patterns of positive selection and significant amino acid diversification across species is consistent with the Selection, Pleiotropy, and Compensation (SPC) model, where detected selection at bam is consistent with adaptive change in one major trait followed by positively selected compensatory changes for pleiotropic effects (in this case perhaps preserving structure). In the case of bam, we suggest that the major trait could be genetic interaction with the endosymbiotic bacteria Wolbachia pipientis. Following up on detected signals of positive selection and comparative structural analysis could provide insight into the distribution of a primary adaptive change versus compensatory changes following a primary change.

molecular biology↗