Lola regulates midline crossing

In each hemisegment of the developing Drosophila ventral nerve cord, the axons of ~20% of the 350 or so embryonic interneurons grow longitudinally on the same side of the midline where their cell bodies lie; they never cross the midline of the CNS (Schmid et al., 1999). In part, this pattern of axon growth reflects the existence of molecular forces that repel axons away from the midline (Klambt et al., 1991; Seeger et al., 1993; Kidd et al., 1998b), and other forces that attract axons to the developing longitudinal axon tracts (Giniger et al., 1993; Giniger, 1998). Midline repulsion alone cannot explain CNS axon pattern, however, as ~70% of interneurons do cross the midline – once – to project in the contralateral longitudinal tract (Schmid et al., 1999). Evidently, the trajectory of a CNS axon reflects a delicate balance of repulsion from and attraction toward the midline, as well as attraction to the longitudinal tracts (Seeger et al., 1993; Hummel et al., 1999a). In recent years, a great deal has been learned about the extracellular molecules that provide some of these competing signals to CNS axons, and about the growth cone receptors that read and interpret those signals. Thus, the secreted protein Slit is made by cells of the midline glia (Rothberg et al., 1990) and repels susceptible axons away from the midline (Kidd et al., 1999). Growth cones recognize and respond to Slit because they have on their surface a family of receptors related to the protein Roundabout (Robo), a repulsive guidance receptor that binds and is activated by Slit (Kidd et al., 1999; Rajagopalan et al., 2000; Simpson et al., 2000). All CNS neurons express Robo, but its activity is modulated post-translationally by the transmembrane protein Commissureless (Comm) (Tear et al., 1996; Kidd et al., 1998b). Comm apparently removes Robo from the plasma membrane, perhaps by activating its endocytosis (Kidd et al., 1998b; Wolf et al., 1998). Comm thus allows particular axons to cross the midline by rendering them insensitive to Slit. The complement of proteins on the surface of a growth cone, and thus the trajectory of that axon, depends upon the genetic program which specifies the identity of that neuron (Ghysen et al., 1985; Miller et al., 1992; Nottebohm et al., 1992; Jurata et al., 2000). For example, a combinatorial ‘code’ of Lim family homeodomain proteins determines particular motoneuron trajectories both in flies (Thor et al., 1999) and in vertebrates (Tsuchida et al., 1994). In this example, the nuclear control of axonal trajectory is intimately intertwined with the very definition of the identities of these neurons. It is not enough, however, for a particular spectrum of proteins to be present on the growth cone of a given neuron. Rather, the precise level (Winberg et al., 1998) and timing (Rose et al., 1997) of the expression of these proteins must also be coordinated exactly (Daston and Koester, 1996; Madden et al., 1999). At the Drosophila midline, the net effect of Robo, Slit and Comm depends on their relative levels of expression (Kidd et al., 1998b; Rajagopalan et al., 2000), and of their level of activity relative to that of the midline attractant(s) (Bashaw and Goodman, 1999). Thus, modest overexpression of Comm in a normally ipsilateral neuron can reduce Robo level – and consequently the sensitivity to Slit – to the point where the axon crosses the midline inappropriately (Kidd et al., 1998b). Similarly, simultaneous reduction of both Robo and Slit levels by just 50% suffices to cause widespread midline crossing (Battye et al., 1999; Kidd et al., 1999). Mechanisms must therefore exist that coordinate the programs of guidance gene expression to ensure that all of the many guidance proteins 1317 Development 129, 1317-1325 (2002) Printed in Great Britain © The Company of Biologists Limited 2002 DEV9836