BIOLOGICAL OVERVIEW

beaten path codes for a secreted immunoglobulin domain protein that is required for subsets of motor axons to correctly defasciculate from other motor axons at specific choice points. Without this protein, motor axons do not properly enter their muscle target regions. The mutant phenotype resembles a result from previously published experiments where motor axon adhesivity is increased by overexpression of the cell adhesion molecule Fasciclin II (Lin, 1994 ). This similarity suggests that Beat may normally work to oppose Fas II-mediated adhesivity and allow defasciculation of otherwise adherent sets of motor axons. Genetic analysis supports this interpretation: when motoneurons have reduced quantities of Fas II, they have less need for beat function, as evidenced by suppression of the beat bypass phenotype double mutant embryos. A similar interaction is observed with the connectin gene, which encodes another cell adhesion protein expressed by a subset of motoneurons (Fambrough, 1996).

The beat phenotype and its apparent antiadhesive function sheds light on the motoneuron outgrowth mechanism. At choice points, motor axons selectively defasciculate from the common pathway and steer into their muscle target region. Several arguments suggest that defasciculation and steering are separable events. (1) The beat mutation affects all motoneuron defasciculation events, whereas mutations in genes involved in steering events might be expected to affect subsets of steering decisions. (2) The strong genetic interactions between beat and genes coding for cell adhesion proteins, both expressed in motoneurons, are not consistent with a role in steering, which is presumably a motoneuron-target muscle interaction. (3) Motoneurons would be expected to receive steering signals, not send them, and the secreted nature of Beat is not consistent with a receptor role. The common thread that runs through the phenotypes of the beat mutant and the misexpression experiments is that Beat interferes with cellular adhesive interactions. It seems more likely that Beat works through a receptor. One candidate receptor is the Fas II protein, with Beat binding to Fas II and blocking Fas II-mediated homophilic adhesion. Several lines of evidence argue against a direct interaction between Beat and Fas II: (1) ectopic expression of Beat on Fas II-expressing axon pathways in the CNS does not disrupt axon fasciculation in those pathways. If Beat functions as an antiadhesive for motor axons by directly binding to Fas II, then one would expect that such misexpression of Beat would lead to defasciculation of these Fas II-positive pathways. (2) Beat expression by muscles interferes with motoneuron-muscle interactions, and Fas II has not been strongly implicated in such interactions. Rather, the simplest interpretation of this result is that Beat interferes with specific interactions of motor axons with their target muscles, events that do not normally involve Fas II. (3) There exists a second gene, sidestep, that mutates to an identical phenotype as beat and thus is a candidate gene to encode the Beat receptor. (4) Cell coaggregation experiments using S2 cells expressing Fas II, and S2 cells expressing Beat tethered to the membrane with a GPI anchor, do not show any evidence of a binding interaction between Beat and Fas II (Fambrough, 1996).

The axonal adhesion molecule Fasciclin II and the secreted anti-adhesion molecule Beaten path carry out critical roles in the development of at least one set of sensory organs, the larval visual organs called Bolwig's organ (BOs). Instead of playing a role in axon defasciculation, in the development of Bolwig's organ, Beat appears to be involved in cell adhesion, a related phenomenon. In normal development, secretion of Beaten path by cells of the optic lobes (a portion of the brain that processes visual information sent from the eyes) allows the Fasciclin II-expressing larval visual organ cells to detach from the optic lobes as a cohesive cell cluster. Thus mechanisms guiding neuronal development may be shared between motoneurons and sensory organs, and adhesion and anti-adhesion are likely to be critical for early steps in the development of the larval visual system (Holmes, 1999).

The larval visual system (LVS) is a relatively simple sensory system composed of BOs: two clusters, each composed of 12 photoreceptor cells from which axons extend in a single fascicle to the brain. Development of the LVS differs in several significant ways from that of the motoneurons. Notably, the axon tracks of the Drosophila CNS and PNS develop through axonal outgrowth and migration, characterized by growth cone extension and selective fasciculation. In contrast, the initial connections of the pioneer axons of the LVS are established when the BOs are close to their target cells, the developing larval brain. Thus, it is neuronal cell bodies, the BOs, that move through a complex environment during LVS development, rather than the axons as in development of the CNS and PNS. It is currently unclear whether the anterior relocation of the BOs occurs as a passive consequence of head involution, or whether an active migratory process is involved. Preliminary analysis of two identified mutations, not enough anterior extension and out of place, which result in the failure of the BOs to attain their proper location, reveals that, despite failures in BO development, head involution appears to occur normally (Holmes, 1998a). The apparent genetic separation of BO movement and head involution suggests the processes may also be mechanistically separable. However, the developmental mechanism driving BO migration remains to be determined (Holmes, 1999).

The BOs arise from cells of the optic lobe placodes (OLs), ectodermal tissues that will go on to form the optic lobes of the adult fly. The BOs detach from the OLs and remain at the periphery of the embryo when the OLs invaginate (Green, 1993). As the BOs detach from the OLs, axons extend from the BOs and establish initial connections with the brain, which is adjacent to the OLs. Therefore, the axons need only navigate a relatively short distance to reach their targets in the brain at this early stage of development. The BO cells remain clustered for the remainder of development, during which they migrate anteriorly and the Bolwig's nerves (BNs) increase significantly in length. Therefore, proper development of the LVS requires that the BO precursors detach from the OLs, that the pioneer connections between the BOs and the brain be established and maintained, that axons properly fasciculate with the pioneer axons, and that the BOs migrate correctly to their final positions (Holmes, 1999).

By genetic analysis of LVS development, a mutant allele of beat has been identified that disrupts early stages of BO development (Holmes, 1998). Beat is expressed in a cluster of cells in the OLs before the BOs become distinct. In LVS development, as in motor neuron development, beat interacts genetically with fas II. A mutation in fas II also disrupts LVS development, resulting in a phenotype that is, in some ways, the opposite of that resulting from mutations in beat. Conversely, overexpression of fas II in the BO causes defects in BO development resembling those that result from mutations in beat. By directing expression of beat to either the BOs or the OLs, the LVS phenotype of beat mutations is reversed. Together, these results demonstrate that detachment of the BO precursors from the OL may involve interaction between Beat and Fas II. Thus, at least some of the mechanisms important for regulating intercellular interactions during motor axon development also function to guide development of sensory organs, suggesting that some of the general principles of neuronal development may overlap in these two different neuronal systems (Holmes, 1999).

PROTEIN STRUCTURE

Amino Acids - 427

Structural Domains

Sequence analysis suggests that the first 26 amino acids are a cleaved signal peptide. The predicted protein does not contain any other significant hydrophobic stretches, and thus the putative Beat protein should be secreted. There are four possible N-linked glycosylation sites. The mature 401 amino-acid chain of Beat can be reliably parsed into three discrete globular domains: a tandem repeat of Ig domain modules with distinctive Cys and Try landmarks (D1, residues 1-114; D2, 115-220) separated by an unstructured linker (221-293) from a Cys-rich C-terminal segment. The Amino-terminal Ig D1 module resembles vertebrate T-cell receptor sequences, whereas the more economical Ig D2 resembles more divergent Ig superfamily molecules like Drosophila Fasciclin III. Structural analysis suggests that Beat Ig domain consist of nine-beta-stranded V-type structures; the D2 module displays shorter loops. The carboxy-terminal D3 module contains a pattern of six Cys residues loosely similar to a 'cysteine-knot' motif conserved in the beta-sheet folds of growth factors like transforming growth factor-beta, neurotrophins, platelet-derived growth factors and glycoprotein hormones (Mushegian, 1997 and Bazan, 1997)

date revised: 6 January 99

Home page: The Interactive Fly © 1995, 1996 Thomas B. Brody, Ph.D.

The Interactive Fly resides on the
Society for Developmental Biology's Web server.