Vuong-Brender, Thanh Thi Kim et al. (2021) International Worm Meeting "CAMTA tunes neural excitability and behavior my modulating Calmodulin expression"

Flynn, Sean, Vuong-Brender, Thanh Thi Kim, de Bono, Mario

[International Worm Meeting, 2021]

Calcium signaling (Ca2+) regulates a wide range of essential biological processes in multiple tissues. In particular, in the nervous system, it mediates signal transduction, synaptic release and activity-dependent transcription important for learning and memory. The Ca2+ sensor Calmodulin (CaM) plays a key role in Ca2+ signaling, however, what mechanism regulates neuronal CaM levels is unclear. The CaM-binding transcription activators (CAMTAs) are expressed broadly and selectively in the nervous system, and CAMTA mutations confer multiple behavioral defects in different species including mouse and human. Using single-neuronal-type RNA-seq and ChIP-seq, we show that the sole C. elegans CAMTA, camt-1, is a master regulator of neuronal CaM expression. The pleiotropic behavioral and neuronal Ca2+ signaling defects in camt-1 mutants can be rescued by supplementing neuronal CaM. CAMT-1 binds multiple sites in the calmodulin promoter and deleting these sites phenocopies camt-1. CAMT-1 not only stimulates CaM expression, but can also inhibit it when CaM levels are high, by a feedback mechanism requiring CaM binding sites in CAMT-1. We also show that Drosophila CAMTA regulates CaM levels suggesting that CAMTAs mediate an evolutionally conserved mechanism that controls neuronal CaM levels, thereby regulating Ca2+ signaling, physiology and behavior.

Artan, Murat et al. (2021) International Worm Meeting "Interactome analysis of C. elegans synapses by TurboID-based proximity labeling"

Skehel, Mark, Nicolas, Armel, Barratt, Stephen, de Bono, Mario, Artan, Murat, Begum, Farida, Flynn, Sean M.

[International Worm Meeting, 2021]

Characterizing the proteome of subcellular structures and the interactome of specific proteins provides entry points to probe molecular function. Several methods for screening protein-protein interactions (PPIs) have proven useful, including affinity purification and two hybrid screens. However, these methods have limitations, including poor detection of transient or weak PPIs, high false positives, and difficulty detecting PPIs in specific cell types or subcellular compartments. Many of these limitations can be overcome using proximity-dependent protein labeling (PL). We optimize a PL protocol for C. elegans using TurboID, a recently improved E. coli promiscuous biotin ligase. We analyse the proteome of individual tissues including the nervous system, intestine, hypodermis and muscle, and, to demonstrate single cell resolution, of the pair of AFD neurons. We observe enrichment of tissue-specific proteins, and identify both known and previously unknown AFD-specific proteins. To characterize the interactome of a specific protein, we knocked TurboID into the endogenous elks-1 gene, which encodes a presynaptic active zone protein. ELKS-1-TurboID highlights both characterized and previously uncharacterized synaptic proteins. We use our data to define non-specific contaminants that should help interpretation of C. elegans PL data. TurboID proximity labeling provides a powerful tool to investigate C. elegans biology. Research support: This work was supported by an Advanced ERC Grant (269058 ACMO) and a Wellcome Investigator Award (209504/Z/17/Z) to MdB and an ISTplus Fellowship to MA (Marie Sklodowska-Curie agreement No 754411).

Prasad BC et al. (1997) International C. elegans Meeting "THE C. ELEGANS unc-3 GENE ENCODES A HOMOLOGUE OF THE O/E FAMILY OF MAMMALIAN TRANSCRIPTION FACTORS"

Prasad BC, Reed RR, Seydoux GC

[International C. elegans Meeting, 1997]

The expression of specialized signal transduction components in mammalian olfactory neurons is thought to be regulated by the O/E (Olf-1/EBF) family of transcription factors. The O/E proteins are expressed in cells of the olfactory neuronal lineage throughout development, and are also expressed transiently in neurons in the developing nervous system during embryogenesis. We have identified a C. elegans homologue of the mammalian O/E proteins, which displays greater than 80% similarity over 350 amino acids. The CeO/E cDNA maps to cosmid F42D1, previously shown to encode unc-3(1). We demonstrate that CeO/E is the product of the unc-3 gene, mutations in which cause defects in the axonal outgrowth of motor neurons as well as defects in dauer formation, a process requiring chemosensory input. Like its mammalian homologues, CeO/E is expressed in certain chemosensory neurons (ASI amphid neurons) throughout development, and is also expressed transiently in developing motor neurons when these cells undergo axonal outgrowth. Currently, we are defining the DNA sequence binding specificity of the CeO/E protein. These observations suggest that the O/E family of transcription factors play a central and evolutionarily conserved role in the expression of proteins essential for axonal pathfinding and neuronal differentiation. (1) Sean Eddy, WBG 12(5):86 (February 1, 1993)