Axon–glial interactions are critical for the induction of myelination and the domain organization of myelinated fibers. Although molecular complexes that mediate these interactions in the nodal region are known, their counterparts along the internode are poorly defined. We report that neurons and Schwann cells express distinct sets of nectin-like (Necl) proteins: axons highly express Necl-1 and -2, whereas Schwann cells express Necl-4 and lower amounts of Necl-2. These proteins are strikingly localized to the internode, where Necl-1 and -2 on the axon are directly apposed by Necl-4 on the Schwann cell; all three proteins are also enriched at Schmidt-Lanterman incisures. Binding experiments demonstrate that the Necl proteins preferentially mediate heterophilic rather than homophilic interactions. In particular, Necl-1 on axons binds specifically to Necl-4 on Schwann cells. Knockdown of Necl-4 by short hairpin RNA inhibits Schwann cell differentiation and subsequent myelination in cocultures. These results demonstrate a key role for Necl-4 in initiating peripheral nervous system myelination and implicate the Necl proteins as mediators of axo–glial interactions along the internode.

Axon initial segments (AISs) and nodes of Ranvier are sites of action potential generation and propagation, respectively. Both domains are enriched in sodium channels complexed with adhesion molecules (neurofascin [NF] 186 and NrCAM) and cytoskeletal proteins (ankyrin G and βIV spectrin). We show that the AIS and peripheral nervous system (PNS) nodes both require ankyrin G but assemble by distinct mechanisms. The AIS is intrinsically specified; it forms independent of NF186, which is targeted to this site via intracellular interactions that require ankyrin G. In contrast, NF186 is targeted to the node, and independently cleared from the internode, by interactions of its ectodomain with myelinating Schwann cells. NF186 is critical for and initiates PNS node assembly by recruiting ankyrin G, which is required for the localization of sodium channels and the entire nodal complex. Thus, initial segments assemble from the inside out driven by the intrinsic accumulation of ankyrin G, whereas PNS nodes assemble from the outside in, specified by Schwann cells, which direct the NF186-dependent recruitment of ankyrin G.

In Drosophila, glial cell differentiation requires the expression of glial cells missing (gcm) in multiple neural cell lineages, where gcm acts as a binary switch for glial vs. neuronal fate. Thus, the primary event controlling gliogenesis in neural progenitors is the transcription of gcm. In addition, gcm is also required for the differentiation of macrophages, and is expressed in the hemocyte lineage. This dual role of gcm in glial cell and blood cell development underscores the need for the precise temporal and spatial regulation of gcm transcription. To understand how gcm transcription is regulated, we have undertaken an analysis of the cis-regulatory DNA elements of gcm using lacZ reporter activity in transgenic embryos, testing the activity of approximately 35 kilobases of DNA from the gcm locus. We have identified several distinct DNA regions that promote most of the elements of gcm expression. These include elements for general neural expression, gcm-independent and gcm-dependent glial-specific expression, as well as early and late hemocyte expression. We show that expression of an abdominal glial-specific element is dependent on the homeotic gene abdominal-A. Our results indicate that gcm transcription is controlled by a combination of general and lineage-specific elements, positive autoregulation, and neuronal repression.