PersistentengrailedExpression Is Required to Determine Sensory Axon Trajectory, Branching, and Target Choice

Abstract

The transcription factor Engrailed (En) directs, in the cockroach cercal system, the shape of the axonal arborization and the choice of postsynaptic partners of an identified sensory neuron (6m). Knock-out of En using double-stranded RNA interference transforms 6m so that it resembles a neighboring neuron that normally does not express the en gene, has a different arbor anatomy, and makes different connections. We characterized the development of 6m and perturbed en expression at different stages. Our results show that En is not required before birth for 6m to become a neuron, but that it is required in the postmitotic neuron to control axonal arborization and synaptic specificity. Knock-out of En after 6m has entered the CNS does not change the axonal trajectory and has minor effects on axonal branches but causes the formation of synaptic connections typical of an En-negative cell. This suggests that En controls target recognition molecules independently from those guiding the axon. In contrast, double-stranded RNA injection 1 d later does not have any effects on the phenotype of 6m, suggesting that the period of synapse formation is over by the time En levels have fallen or, if synapse turnover occurs, that En is not required to maintain the specificity of synaptic connections. We conclude that persistent en expression is required to determine successive stages in the differentiation of the neuron, suggesting that it is not far upstream from those genes encoding axon guidance and synaptic recognition molecules.

Engrailed Alters the Specificity of Synaptic Connections ofDrosophilaAuditory Neurons with the Giant Fiber

“…When we did use a driver (ato-GAL4 ) that is actually expressed in the precursors but not the early neurons, we found no effect of En knockdown. This apparent postmitotic effect is reminiscent of our previous studies on the cockroach cercal system, which also indicate that En is required postmitotically in sensory neurons to determine first their branching pattern and then their pattern of synaptic connections (Marie et al, 2002). In that system, it seems that, once formed, synapses cannot be altered by En knockdown.…”

“…En regulates the development of some spinal cord interneurons (Wenner et al, 2000;Simon et al, 2001) and affects the survival of dopaminergic midbrain neurons (Simon et al, 2001;Sgadò et al, 2006). In Drosophila and grasshopper CNS, En controls neuron/ glia fate decisions, neuronal identity, and axon pathfinding (Condron et al, 1994;Lundell et al, 1996;Joly et al, 2007), whereas in cockroach mechanosensory neurons we showed that it also controls axon guidance, synaptic target recognition, and, as a result, escape behavior (Marie et al, 2000(Marie et al, , 2002Marie and Blagburn, 2003;Blagburn and Bacon, 2004;Booth et al, 2009).…”

“…In view of the controlling effect of En on synaptic connectivity in the cockroach cercal system (Marie et al, 2000(Marie et al, , 2002Marie and Blagburn, 2003), we hypothesized that the expression of En could influence the formation of JON synapses with the GF. Therefore, we used the JO15-GAL4 line (Sharma et al, 2002), which is expressed in all of the JO-A and the majority of the JO-B neurons (Kamikouchi et al, 2006;Fig.…”

Pax6andEngrailed 1Regulate Two Distinct Aspects of Renshaw Cell Development

“…In this respect it is interesting to note that En1 activity regulates ephrin gene expression in the chick tectum, thereby altering the topographic targeting of retinal axons (Friedman and O'Leary, 1996;Logan et al, 1996). Furthermore, in Drosophila, the expression of synaptic recognition molecules is modulated by engrailed, the fly homolog of En1 (Marie et al, 2002). Interestingly, the recurrent synapses that do form on motor neurons in the En1 mutant mice appear to be functional, because the calbindin ϩ terminals are presynaptic to gephyrin-containing clusters, which are known to be formed and stabilized by activitydependent mechanisms (Kirsch and Betz, 1998;Levi et al, 1999).…”

Engrailed genes are cell-autonomously required to prevent apoptosis in mesencephalic dopaminergic neurons

“…Involvement of En in axonal outgrowth has been demonstrated in mammals and insects (Marie et al, 2002;Saueressig et al, 1999). To address this possibility, we placed dissociated E12 ventral midbrain into a three-dimensional collagen matrix with or without an explant of basal telencephalon of the same age.…”