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1. Single-cell transcriptomes of developing and adult olfactory receptor neurons in Drosophila

   https://doi.org/10.7554/eLife.63856  

   McLaughlin, Colleen N; Brbić, Maria; Xie, Qijing; Li, Tongchao; Horns, Felix; Kolluru, Sai Saroja; Kebschull, Justus M; Vacek, David; Xie, Anthony; Li, Jiefu; et al (February 2021, eLife)

   null (Ed.)

   Recognition of environmental cues is essential for the survival of all organisms. Transcriptional changes occur to enable the generation and function of the neural circuits underlying sensory perception. To gain insight into these changes, we generated single-cell transcriptomes of Drosophila olfactory- (ORNs), thermo-, and hygro-sensory neurons at an early developmental and adult stage using single-cell and single-nucleus RNA sequencing. We discovered that ORNs maintain expression of the same olfactory receptors across development. Using receptor expression and computational approaches, we matched transcriptomic clusters corresponding to anatomically and physiologically defined neuron types across multiple developmental stages. We found that cell-type-specific transcriptomes partly reflected axon trajectory choices in development and sensory modality in adults. We uncovered stage-specific genes that could regulate the wiring and sensory responses of distinct ORN types. Collectively, our data reveal transcriptomic features of sensory neuron biology and provide a resource for future studies of their development and physiology. 

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2. Inferring Network Structure from Neural Activity in a Biologically-Constrained Model of the Insect Olfactory System

   Joshi, Shruti; Navas-Zuloaga, Maria Gabriela; McLane-Svoboda, Autumn; Sanchez, Simon; Saha, Debajit; Bazhenov, Maxim (August 2024, https://2024.ccneuro.org/)

   Understanding olfactory processing in insects requires characterizing the complex dynamics and connectivity of the first olfactory relay - antennal lobe (AL). We leverage in vivo electrophysiology to train recurrent neural network (RNN) model of the locust AL, inferring the underlying connectivity and temporal dynamics. The RNN comprises 830 projection neurons (PNs) and 300 local neurons (LNs), replicating the locust AL anatomy. The trained network reveals sparse connectivity, with different connection densities between LNs and PNs and no PN-PN connections, consistent with in vivo data. The learned time constants predict slower LN dynamics and diverse PN response patterns, with low and high time constants correlating with early and late odor-evoked activity, as reported in vivo. Our approach demonstrates the utility of biologically-constrained RNNs in inferring circuit properties from empirical data, providing insights into mechanisms of odor coding in the AL. 

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3. Deciphering the combinatorial expression pattern and genetic regulatory mechanisms of Beats and Sides in the olfactory circuits of Drosophila

   https://doi.org/10.1101/2025.05.31.657193  

   Duan, Ǫichen; Okuwa, Sumie; Estrella, Rachel; Yeung, Chun; Chen, Yu-Chieh David; Rio, Laura Ǫuintana; Vien, Khanh M; Volkan, Pelin C (June 2025, bioRxiv)

   Abstract Over the past decades, many critical molecular players have been uncovered to control distinct steps in olfactory circuit assembly in Drosophila. Among these, multi-member gene families of cell surface proteins are of interest because they can act as neuron-specific identification/recognition tags in combinations and contribute to circuit assembly in complex brains through their heterophilic or homophilic interactions. Recently, a multi-protein interactome has been described between the Beat and Side families of IgSF proteins. Here, we use the publicly available single-cell RNA-seq datasets and newly generated gene trap transgenic driver lines to probe thein vivospatial expression pattern of thebeat/sidegene families in odorant receptor neurons (ORNs) and their synaptic target projection neurons (PNs).Our results revealed that each ORN and its synaptic target PN class expresses a class-specific combination ofbeat/sidegenes, hierarchically regulated by lineage-specific genetic programs. Though ORNs or PNs from closer lineages tend to possess more similarbeat/sideprofiles, we also found many examples of divergence from this pattern among closely related ORNs and closely related PNs. To explore whether the class-specific combination ofbeats/sidesdefines ORN-PN matching specificity, we perturbed presynapticbeat-IIaand postsynapticside-IVin two ORN-PN partners. However, disruption of Beat-IIa-Side-IV interaction did not produce any significant mistargeting in these two examined glomeruli. Though without affecting general glomerular targeting, knockdown ofsidein ORNs leads to the reduction of synaptic development. Interestingly, we found conserved expression patterns ofbeat/sideorthologs across ORNs in ants and mosquitoes, indicating the shared regulatory strategies specifying the expression of these duplicated paralogs in insect evolution. Overall, this comprehensive analysis of expression patterns lays a foundation for in-depth functional investigations into how Beat/Side combinatorial expression contributes to olfactory circuit assembly. 

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4. Coordinated control of neuronal differentiation and wiring by sustained transcription factors

   https://doi.org/10.1126/science.add1884  

   Özel, Mehmet Neset; Gibbs, Claudia Skok; Holguera, Isabel; Soliman, Mennah; Bonneau, Richard; Desplan, Claude (December 2022, Science)

   INTRODUCTION Neurons are by far the most diverse of all cell types in animals, to the extent that “cell types” in mammalian brains are still mostly heterogeneous groups, and there is no consensus definition of the term. The Drosophila optic lobes, with approximately 200 well-defined cell types, provides a tractable system with which to address the genetic basis of neuronal type diversity. We previously characterized the distinct developmental gene expression program of each of these types using single-cell RNA sequencing (scRNA-seq), with one-to-one correspondence to the known morphological types. RATIONALE The identity of fly neurons is determined by temporal and spatial patterning mechanisms in stem cell progenitors, but it remained unclear how these cell fate decisions are implemented and maintained in postmitotic neurons. It was proposed in Caenorhabditis elegans that unique combinations of terminal selector transcription factors (TFs) that are continuously expressed in each neuron control nearly all of its type-specific gene expression. This model implies that it should be possible to engineer predictable and complete switches of identity between different neurons just by modifying these sustained TFs. We aimed to test this prediction in the Drosophila visual system. RESULTS Here, we used our developmental scRNA-seq atlases to identify the potential terminal selector genes in all optic lobe neurons. We found unique combinations of, on average, 10 differentially expressed and stably maintained (across all stages of development) TFs in each neuron. Through genetic gain- and loss-of-function experiments in postmitotic neurons, we showed that modifications of these selector codes are sufficient to induce predictable switches of identity between various cell types. Combinations of terminal selectors jointly control both developmental (e.g., morphology) and functional (e.g., neurotransmitters and their receptors) features of neurons. The closely related Transmedullary 1 (Tm1), Tm2, Tm4, and Tm6 neurons (see the figure) share a similar code of terminal selectors, but can be distinguished from each other by three TFs that are continuously and specifically expressed in one of these cell types: Drgx in Tm1, Pdm3 in Tm2, and SoxN in Tm6. We showed that the removal of each of these selectors in these cell types reprograms them to the default Tm4 fate. We validated these conversions using both morphological features and molecular markers. In addition, we performed scRNA-seq to show that ectopic expression of pdm3 in Tm4 and Tm6 neurons converts them to neurons with transcriptomes that are nearly indistinguishable from that of wild-type Tm2 neurons. We also show that Drgx expression in Tm1 neurons is regulated by Klumpfuss, a TF expressed in stem cells that instructs this fate in progenitors, establishing a link between the regulatory programs that specify neuronal fates and those that implement them. We identified an intronic enhancer in the Drgx locus whose chromatin is specifically accessible in Tm1 neurons and in which Klu motifs are enriched. Genomic deletion of this region knocked down Drgx expression specifically in Tm1 neurons, leaving it intact in the other cell types that normally express it. We further validated this concept by demonstrating that ectopic expression of Vsx (visual system homeobox) genes in Mi15 neurons not only converts them morphologically to Dm2 neurons, but also leads to the loss of their aminergic identity. Our results suggest that selector combinations can be further sculpted by receptor tyrosine kinase signaling after neurogenesis, providing a potential mechanism for postmitotic plasticity of neuronal fates. Finally, we combined our transcriptomic datasets with previously generated chromatin accessibility datasets to understand the mechanisms that control brain wiring downstream of terminal selectors. We built predictive computational models of gene regulatory networks using the Inferelator framework. Experimental validations of these networks revealed how selectors interact with ecdysone-responsive TFs to activate a large and specific repertoire of cell surface proteins and other effectors in each neuron at the onset of synapse formation. We showed that these network models can be used to identify downstream effectors that mediate specific cellular decisions during circuit formation. For instance, reduced levels of cut expression in Tm2 neurons, because of its negative regulation by pdm3 , controls the synaptic layer targeting of their axons. Knockdown of cut in Tm1 neurons is sufficient to redirect their axons to the Tm2 layer in the lobula neuropil without affecting other morphological features. CONCLUSION Our results support a model in which neuronal type identity is primarily determined by a relatively simple code of continuously expressed terminal selector TFs in each cell type throughout development. Our results provide a unified framework of how specific fates are initiated and maintained in postmitotic neurons and open new avenues to understanding synaptic specificity through gene regulatory networks. The conservation of this regulatory logic in both C. elegans and Drosophila makes it likely that the terminal selector concept will also be useful in understanding and manipulating the neuronal diversity of mammalian brains. Terminal selectors enable predictive cell fate reprogramming. Tm1, Tm2, Tm4, and Tm6 neurons of the Drosophila visual system share a core set of TFs continuously expressed by each cell type (simplified). The default Tm4 fate is overridden by the expression of a single additional terminal selector to generate Tm1 ( Drgx ), Tm2 ( pdm3 ), or Tm6 ( SoxN ) fates. 

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5. Genipin Crosslinks the Extracellular Matrix to Rescue Developmental and Degenerative Defects, and Accelerates Regeneration of Peripheral Neurons

   Kenyi Saito-Diaz, Paula Dietrich (March 2023, Science translational medicine)

   The peripheral nervous system (PNS) is essential for proper body function. A high percentage of the population suffer nerve degeneration or peripheral damage. For example, over 40% of patients with diabetes or undergoing chemotherapy develop peripheral neuropathies. Despite this, there are major gaps in the knowledge of human PNS development and therefore, there are no available treatments. Familial Dysautonomia (FD) is a devastating disorder that specifically affects the PNS making it an ideal model to study PNS dysfunction. FD is caused by a homozygous point mutation in ELP1 leading to developmental and degenerative defects in the sensory and autonomic lineages. We previously employed human pluripotent stem cells (hPSCs) to show that peripheral sensory neurons (SNs) are not generated efficiently and degenerate over time in FD. Here, we conducted a chemical screen to identify compounds able to rescue this SN differentiation inefficiency. We identified that genipin, a compound prescribed in Traditional Chinese Medicine for neurodegenerative disorders, restores neural crest and SN development in FD, both in the hPSC model and in a FD mouse model. Additionally, genipin prevented FD neuronal degeneration, suggesting that it could be offered to patients suffering from PNS neurodegenerative disorders. We found that genipin crosslinks the extracellular matrix, increases the stiffness of the ECM, reorganizes the actin cytoskeleton, and promotes transcription of YAP-dependent genes. Finally, we show that genipin enhances axon regeneration in an in vitro axotomy model in healthy sensory and sympathetic neurons (part of the PNS) and in prefrontal cortical neurons (part of the central nervous system, CNS). Our results suggest genipin can be used as a promising drug candidate for treatment of neurodevelopmental and neurodegenerative diseases, and as a enhancer of neuronal regeneration. 

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