Extendingjulius seizure, a bang-sensitive gene, as a model for studying epileptogenesis: Cold shock, and a new insertional mutation

Abstract

The bang-sensitive (BS) mutants of Drosophila are an important model for studying epilepsy. We recently identified a novel BS locus, julius seizure (jus), encoding a protein containing two transmembrane domains and an extracellular cysteine-rich loop. We also determined that jus, a previously identified BS mutation, is an allele of jus by recombination, deficiency mapping, complementation testing, and genetic rescue. RNAi knockdown revealed that jus expression is important in cholinergic neurons and that the critical stage of jus expression is the mid-pupa. Finally, we found that a functional, GFP-tagged genomic construct of jus is expressed mostly in axons of the neck connectives and of the thoracic abdominal ganglia. In this Extra View article, we show that a MiMiC GFP-tagged Jus is localized to the same nervous system regions as the GFP-tagged genomic construct, but its expression is mostly confined to cell bodies and it causes bang-sensitivity. The MiMiC GFP-tag lies in the extracellular loop while the genomic construct is tagged at the C-terminus. This suggests that the alternate position of the GFP tag may disrupt Jus protein function by altering its subcellular localization and/or stability. We also show that a small subset of jus-expressing neurons are responsible for the BS phenotype. Finally, extending the utility of the BS seizure model, we show that jus mutants exhibit cold-sensitive paralysis and are partially sensitive to strobe-induced seizures.

Investigating rare and ultrarare epilepsy syndromes with Drosophila models

“…Mutations in 1,200 Drosophila genes have been registered as associated with heat sensitivity. Increasing the temperature of ectothermal animals increases the temperature of the central nervous system and induces the stereotypic seizure response in sensitized genetic models 21 , 33 – 35 , 47 . Surprisingly few (~30%) of the genes that can mutate to produce mechanically induced phenotypes co-occur with those that are temperature dependent.…”

A Complex Containing the Na+/K+ATPase Regulates Epileptogenesis inDrosophila melanogaster

“…For RNAi knockdown of jus expression, the following jus enhancer GAL4 lines were used (described in Pfeiffer et al, 2008): 55F051 (BL# 39124), 55G02 (BL# 46070; no longer available from BDSC but see Horne et al, 2017 and Dean et al, 2018), 59D01 (BL# 46416) and 90B09 (this paper). Each of these jus -GAL4 lines was crossed to the UAS- jus RNAi line P{y[+t7.7] v[+t1.8]=TRiP.JF03192}attP2 (BL#28764; Ni et al 2009).…”

“…Previous work demonstrated that using the 55G02-GAL4 line to drive expression of a wildtype jus transgene could effectively rescue the bang sensitive phenotype of a mutant jus allele (Horne et al, 2018). However, this rescue was not complete, and later it was found that overlap of 55G02-GAL4 expression with endogenous Jus expression was limited (Dean et al, 2018). These findings suggested that 55G02 may not be the most effective enhancer to manipulate gene expression in the Jus-expressing neurons that affect bang-sensitivity.…”

“…In contrast, Drosophila have an even more sophisticated array of genetic and transgenic tools along with a modest-sized nervous system of about 100,000 neurons. Epilepsy models in Drosophila, known as bang sensitive (BS) mutants, undergo seizures in response to mechanical electrical or temperature shocks, or less effectively, strobe light (Benzer, 1971; Ganetzky and Wu, 1982; Lee and Wu, 2002; Pavlidis and Tanouye, 1995; Burg and Wu, 2012; Giachello and Baines, 2015; Dean et al, 2018). This behavioral output is easily scored, but there are challenges in using BS mutants to study epileptogenesis.…”

“…jus encodes a predicted two-transmembrane protein that is expressed in selected neurons in the brain, including the optic lobe and photoreceptors, and in a wide but specific pattern in the thoracic abdominal ganglion. Expression of Jus during pupal life is crucial to prevent epilepsy in the adult (Horne et al, 2017; Dean et al, 2018). In short, Jus has features that could make it valuable for studying epileptogenesis: namely, it has a critical developmental period for expression, and it is expressed in a small population of neurons.…”