Analysis of Bub3 and Nup75 in the Drosophila male germline lineage.

Extensive communication at the stem cell-niche interface and asymmetric stem cell division is key for the homeostasis of the Drosophila male germline stem cell system. To improve our understanding of these processes, we analysed the function of the mitotic checkpoint complex (MCC) component Bub3 and the nucleoporin Nup75, a component of the nuclear pore complex realizing the transport of signalling effector molecules to the nucleus, in the Drosophila testis. By lineage-specific interference, we found that the two genes control germline development and maintenance. Bub3 is continuously required in the germline, as its loss results in the beginning in an over-proliferation of early germ cells and later on in loss of the germline. The absence of the germline lineage in such testes has dramatic cell non-autonomous consequences, as cells co-expressing markers of hub and somatic cyst cell fates accumulate and populate in extreme cases the whole testis. Our analysis of Nups showed that some of them are critical for lineage maintenance, as their depletion results in the loss of the affected lineage. In contrast, Nup75 plays a role in controlling proliferation of early germ cells but not differentiating spermatogonia and seems to be involved in keeping hub cells quiescent. In sum, our analysis shows that Bub3 and Nup75 are required for male germline development and maintenance.

Single-cell RNA sequencing of motoneurons identifies regulators of synaptic wiring in Drosophila embryos.

The correct wiring of neuronal circuits is one of the most complex processes in development, since axons form highly specific connections out of a vast number of possibilities. Circuit structure is genetically determined in vertebrates and invertebrates, but the mechanisms guiding each axon to precisely innervate a unique pre-specified target cell are poorly understood. We investigated Drosophila embryonic motoneurons using single-cell genomics, imaging, and genetics. We show that a cell-specific combination of homeodomain transcription factors and downstream immunoglobulin domain proteins is expressed in individual cells and plays an important role in determining cell-specific connections between differentiated motoneurons and target muscles. We provide genetic evidence for a functional role of five homeodomain transcription factors and four immunoglobulins in the neuromuscular wiring. Knockdown and ectopic expression of these homeodomain transcription factors induces cell-specific synaptic wiring defects that are partly phenocopied by genetic modulations of their immunoglobulin targets. Taken together, our data suggest that homeodomain transcription factor and immunoglobulin molecule expression could be directly linked and function as a crucial determinant of neuronal circuit structure.

The Hox transcription factor Ultrabithorax binds RNA and regulates co-transcriptional splicing through an interplay with RNA polymerase II.

Transcription factors (TFs) play a pivotal role in cell fate decision by coordinating gene expression programs. Although most TFs act at the DNA layer, few TFs bind RNA and modulate splicing. Yet, the mechanistic cues underlying TFs activity in splicing remain elusive. Focusing on the Drosophila Hox TF Ultrabithorax (Ubx), our work shed light on a novel layer of Ubx function at the RNA level. Transcriptome and genome-wide binding profiles in embryonic mesoderm and Drosophila cells indicate that Ubx regulates mRNA expression and splicing to promote distinct outcomes in defined cellular contexts. Our results demonstrate a new RNA-binding ability of Ubx. We find that the N51 amino acid of the DNA-binding Homeodomain is non-essential for RNA interaction in vitro, but is required for RNA interaction in vivo and Ubx splicing activity. Moreover, mutation of the N51 amino acid weakens the interaction between Ubx and active RNA Polymerase II (Pol II). Our results reveal that Ubx regulates elongation-coupled splicing, which could be coordinated by a dynamic interplay with active Pol II on chromatin. Overall, our work uncovered a novel role of the Hox TFs at the mRNA regulatory layer. This could be an essential function for other classes of TFs to control cell diversity.