Cbln1 and Cbln4 are structurally similar but differ in GluD2 binding interactions

The Cerebellin (Cbln) family proteins contain four members (Cbln1-4) that share high sequence identity and similar structural organization with a C1q domain at the C-terminus. Intriguingly, Cbln1 has been found to function in synaptogenesis through bridging neurexin (Nrxn) proteins and  type glutamate receptors at the synaptic cleft, whereas Cbln4 has been shown to have no or very weak binding to Nrxn or δ type glutamate receptors but is a ligand for DCC, despite its high sequence identity (~74%) with Cbln1.

A research team with leading scientists of Dr. DING Jianping and Dr. ZHONG Chen at Shanghai Institute of Biochemistry and Cell Biology, Chinese Academy of Sciences (CAS) reported a most recent progress on the similarities and divergence of Cbln proteins. The study was published in Cell Reports.

Researchers determined the crystal structures of the homotrimers of the C1q domain of Cbln1 and Cbln4 (Cbln1C1q and Cbln4C1q) at 2.2 Å and 2.3 Å resolution, respectively. The structures show that Cbln1C1q and Cbln4C1q take a very similar overall conformation but exhibit significant differences at the region equivalent to the Cbln1C1q-GluD2ATD interface.

For the hexamer of full-length Cbln1, negative-stain single-particle electron microscopy (EM) reconstruction was calculated at 13 Å resolution. The EM density map is well fitted with the crystal structure of the Cbln1C1q homotrimer and a predicted model of the N-terminal region of Cbln1. The reconstruction displays that the overall shape of Cbln1 likes two cherries connected with stems: two copies of the crystal structure of the Cbln1C1q trimer are well fitted into the two “cherries” in the density; and the N-terminal region of Cbln1 interlocks with each other to form a relatively flat surface, through which the hexamer is formed.

In addition, to resolve the discrepancy of some of the data in the literature, researchers developed a semi-quantitative cell surface binding method to determine the binding affinity of Cbln1 and Cbln4 to the full-length Nrxn1β. Surprisingly, the binding of Cbln4 to the full-length Nrxn1β is not weak. Indeed, Cbln4 and the LNS domain of Nrxn1β can form a stable complex in vitro. Furthermore, the EM density map of the Cbln4/Nrxn1β complex at 19 Å resolution suggests that Nrxn1β binds to the N-terminal region of Cbln4, likely through strand β10 of the S4 splicing insert. Taken together, these findings provide new insights into the similarities and divergence of the functional roles of the Cbln proteins in synaptogenesis at the molecular level.

This study was supported by the grants from National Natural Science Foundation of China, Ministry of Science and Technology, and CAS.


AUTHOR CONTACT:
DING Jianping
Shanghai Institute of Biochemistry and Cell Biology, Chinese Academy of Sciences, Shanghai, China
E-mail: jpding@sibcb.ac.cn