Cbln3, a Novel Member of the Precerebellin Family that Binds Specifically to Cbln1

Abstract

Precerebellin (Cbln1) is the precursor of the brain-specific hexadecapeptide cerebellin. Although cerebellin has properties of a conventional neuropeptide, its function is controversial because Cbln1 has structural features characteristic of circulating atypical collagens. Cbln1 is related to the three subunits of the complement C1q complex. Therefore, we hypothesized that Cbln1 participated in analogous heteromeric complexes with precerebellin-related proteins. Using LexA-Cbln1 as bait in a yeast two-hybrid screen, we isolated a cDNA encoding a novel Cbln1-related protein, designated Cbln3. The gene encoding cbln3 had the same intron-exon structure as cbln1 but mapped to a different mouse chromosome (14). The deduced amino acid sequence of Cbln3 was 55% identical to Cbln1 and also contained a C1q signature domain and signal sequence for secretion. In addition to binding avidly to Cbln3, Cbln1 also formed homomeric complexes. In contrast, Cbln3 homomeric association was weak. These interactions exhibited specificity because C1qB bound to neither Cbln1 nor Cbln3. Like cbln1, cbln3 was expressed in the cerebellum and dorsal cochlear nucleus in which it was detected in granule neurons. Because Cbln1 and Cbln3 are coexpressed in the brain and interact avidly, they may function as a secreted heteromeric complex in vivo.

Cbln1 Is Essential for Interaction-Dependent Secretion of Cbln3

“…Cbln1 is the prototype of a family of proteins (Cbln1 to Cbln4) that share a conserved C1q globular domain at their C termini (1,27,40,41). This raises the possibility that other family members fulfill functions analogous to that of Cbln1.…”

“…In the case of Cbln1, two homotrimers assemble into hexamers via intermolecular disulfide bonds between conserved cysteine-containing motifs in the N termini of all family members (1). Although Cbln1 forms homomeric complexes, it also binds to Cbln3 in yeast two-hybrid assays (1,27). As Cbln3 is coexpressed with Cbln1 in mature granule neurons (9,27), this raises the question of whether Cbln3 can contribute to the maintenance of synaptic structure and function in cerebellum either by acting alone or by participating in heteromeric complexes with Cbln1.…”

“…The method and probes for Northern blotting were described previously (27). Briefly, total RNA from mouse cerebellum was extracted using RNAzol B (Tel-Test, Friendswood, TX) and hybridized to 32 P-labeled probes from either cbln3 or cbln1.…”

Precerebellin-related genes and precerebellin 1 peptide in the adrenal gland of the rat: Expression pattern, localization, developmental regulation and effects on corticosteroidogenesis

“…However, not only Cbln-derived polypeptides are biologically active compounds. Recently it was demonstrated that cells are able to secrete entire Cbln molecules which in extracellular space form homomeric and heteromeric complexes involved in paracrine or autocrine regulation of nervous cells, among others in synapse development and synaptic plasticity, especially in the cerebellum (5,6,8,42). We cannot exclude that a similar mechanism of Cbln actions may operate in the adrenal gland.…”

Cerebellin and des-cerebellin exert ACTH-like effects on corticosterone secretion and the intracellular signaling pathway gene expression in cultured rat adrenocortical cells - DNA microarray and QPCR studies

“…Cbln1 is homologous to originally described human precerebellin (3) while Cbln2 is homologous to rat and murine cerebellin-like proteins identified by Wada and Ohtani (4) and Kavety et al (5). Cbln3 binds specifically to Cbln1 (6) and recently Cbln4 was identified (7) (NCBI accession number NM_175631). All members of the Cbln family form homomeric and also heteromeric complexes with each other in vitro and it was suggested that such complexes play a crucial role in normal development of the cerebellum (2,8).…”