hkb is required for DIP-α expression and target recognition in the Drosophila neuromuscular circuit.

Our nervous system contains billions of neurons that form precise connections with each other through interactions between cell surface proteins. In Drosophila, the Dpr and DIP immunoglobulin protein subfamilies form homophilic or heterophilic interactions to instruct synaptic connectivity, synaptic growth, and cell survival. However, the upstream regulatory mechanisms of Dprs and DIPs are not clear. On the other hand, while transcription factors have been implicated in target recognition, their downstream cell surface proteins remain mostly unknown. We conduct an F1 dominant modifier genetic screen to identify regulators of Dprs and DIPs. We identify huckebein (hkb), a transcription factor previously implicated in target recognition of the dorsal Is motor neuron. We show that hkb genetically interacts with DIP-α and loss of hkb leads to complete removal of DIP-α expression specifically in dorsal Is motor neurons. We then confirm that this specificity is through the dorsal Is motor neuron specific transcription factor, even-skipped (eve), which acts downstream of hkb. Analysis of the genetic interaction between hkb and eve reveals that they act in the same pathway to regulate dorsal Is motor neuron connectivity. Our study provides insight into the transcriptional regulation of DIP-α and suggests that distinct regulatory mechanisms exist for the same CSP in different neurons.

hkb is required for DIP-α expression and target recognition in the Drosophila neuromuscular circuit.

Our nervous system contains billions of neurons that form precise connections with each other through interactions between cell surface proteins (CSPs). In Drosophila, the Dpr and DIP immunoglobulin protein subfamilies form homophilic or heterophilic interactions to instruct synaptic connectivity, synaptic growth and cell survival. However, the upstream regulation and downstream signaling mechanisms of Dprs and DIPs are not clear. In the Drosophila larval neuromuscular system, DIP-α is expressed in the dorsal and ventral type-Is motor neurons (MNs). We conducted an F1 dominant modifier genetic screen to identify regulators of Dprs and DIPs. We found that the transcription factor, huckebein (hkb), genetically interacts with DIP-α and is important for target recognition specifically in the dorsal Is MN, but not the ventral Is MN. Loss of hkb led to complete removal of DIP-α expression. We then confirmed that this specificity is through the dorsal Is MN specific transcription factor, even-skipped (eve), which acts downstream of hkb. Genetic interaction between hkb and eve revealed that they act in the same pathway to regulate dorsal Is MN connectivity. Our study provides insight into the transcriptional regulation of DIP-α and suggests that distinct regulatory mechanisms exist for the same CSP in different neurons.

Molecular heterogeneity of C. elegans glia across sexes.

A comprehensive description of nervous system function, and sex dimorphism within, is incomplete without clear assessment of the diversity of its component cell types, neurons and glia. C. elegans has an invariant nervous system with the first mapped connectome of a multicellular organism and single-cell atlas of component neurons. Here we present single nuclear RNA-seq evaluation of glia across the entire adult C. elegans nervous system, including both sexes. Machine learning models enabled us to identify both sex-shared and sex-specific glia and glial subclasses. We have identified and validated molecular markers in silico and in vivo for these molecular subcategories. Comparative analytics also reveals previously unappreciated molecular heterogeneity in anatomically identical glia between and within sexes, indicating consequent functional heterogeneity. Furthermore, our datasets reveal that while adult C. elegans glia express neuropeptide genes, they lack the canonical unc-31/CAPS-dependent dense core vesicle release machinery. Thus, glia employ alternate neuromodulator processing mechanisms. Overall, this molecular atlas, available at www.wormglia.org, reveals rich insights into heterogeneity and sex dimorphism in glia across the entire nervous system of an adult animal.

Transsynaptic interactions between IgSF proteins DIP-α and Dpr10 are required for motor neuron targeting specificity.

The Drosophila larval neuromuscular system provides an ideal context in which to study synaptic partner choice, because it contains a small number of pre- and postsynaptic cells connected in an invariant pattern. The discovery of interactions between two subfamilies of IgSF cell surface proteins, the Dprs and the DIPs, provided new candidates for cellular labels controlling synaptic specificity. Here we show that DIP-α is expressed by two identified motor neurons, while its binding partner Dpr10 is expressed by postsynaptic muscle targets. Removal of either DIP-α or Dpr10 results in loss of specific axonal branches and NMJs formed by one motor neuron, MNISN-1s, while other branches of the MNISN-1s axon develop normally. The temporal and spatial expression pattern of dpr10 correlates with muscle innervation by MNISN-1s during embryonic development. We propose a model whereby DIP-α and Dpr10 on opposing synaptic partners interact with each other to generate proper motor neuron connectivity.