Clustered Protocadherins Are Required for Building Functional Neural Circuits

Abstract

Neuronal identity is generated by the cell-surface expression of clustered protocadherin (Pcdh) isoforms. In mice, 58 isoforms from three gene clusters, Pcdhα, Pcdhβ, and Pcdhγ, are differentially expressed in neurons. Since cis-heteromeric Pcdh oligomers on the cell surface interact homophilically with that in other neurons in trans, it has been thought that the Pcdh isoform repertoire determines the binding specificity of synapses. We previously described the cooperative functions of isoforms from all three Pcdh gene clusters in neuronal survival and synapse formation in the spinal cord. However, the neuronal loss and the following neonatal lethality prevented an analysis of the postnatal development and characteristics of the clustered-Pcdh-null (Δαβγ) neural circuits. Here, we used two methods, one to generate the chimeric mice that have transplanted Δαβγ neurons into mouse embryos, and the other to generate double mutant mice harboring null alleles of both the Pcdh gene and the proapoptotic gene Bax to prevent neuronal loss. First, our results showed that the surviving chimeric mice that had a high contribution of Δαβγ cells exhibited paralysis and died in the postnatal period. An analysis of neuronal survival in postnatally developing brain regions of chimeric mice clarified that many Δαβγ neurons in the forebrain were spared from apoptosis, unlike those in the reticular formation of the brainstem. Second, in Δαβγ/Bax null double mutants, the central pattern generator (CPG) for locomotion failed to create a left-right alternating pattern even in the absence of neurodegeneraton. Third, calcium imaging of cultured hippocampal neurons showed that the network activity of Δαβγ neurons tended to be more synchronized and lost the variability in the number of simultaneously active neurons observed in the control network. Lastly, a comparative analysis for trans-homophilic interactions of the exogenously introduced single Pcdh-γA3 isoforms between the control and the Δαβγ neurons suggested that the isoform-specific trans-homophilic interactions require a complete match of the expressed isoform repertoire at the contacting sites between interactive neurons. These results suggested that combinations of clustered Pcdh isoforms are required for building appropriate neural circuits.

The gamma-Protocadherins regulate the survival of GABAergic interneurons during developmentally-regulated cell death

“…week (Duan et al, 2019;Minlebaev et al, 2011;Pangratz-Fuehrer and Hestrin, 2011). cPcdh deletions result in abnormal network activities in spinal cord and synchronized activities in vitro (Hasegawa et al, 2017)…”

Regulation of Wnt signaling by protocadherins

“…These observations are consistent with functions revealed by analyses of Pcdha or Pcdhg mutant mice, which indicated that α- and γ-Pcdhs play critical roles in neuronal survival, axon and dendrite arborization, self-avoidance, and tiling, and synaptogenesis, depending on the neuronal subtype examined [109, 137, 140–152]. Comparison of mice lacking all three Pcdh gene clusters with those lacking only the Pcdha, Pcdhb , or Pcdhg clusters indicates overlapping and synergistic functions for the clustered Pcdh families, though the most severe phenotypes are generally attributable to loss of the γ-Pcdhs [153, 154]. Individual non-clustered δ-Pcdhs are expressed by discrete neuronal subsets throughout the brain, and functional studies collectively have revealed roles in axon outgrowth and pathfinding, synaptic plasticity, and synapse elimination [84, 86, 110, 155–163].…”

CRISPR/Cas9 interrogation of the mouse Pcdhg gene cluster reveals a crucial isoform-specific role for Pcdhgc4

“…In mouse, the Pcdha cluster encodes 14 α-Pcdhs, the Pcdhb cluster encodes 22 β-Pcdhs, and the Pcdhg cluster encodes 22 γ-Pcdhs ( Fig 1A). While all three Pcdh gene clusters contribute to neural development to some extent [4][5][6][7][8][9][10], the Pcdhg locus is the only one required for postnatal viability, and disruption of this locus results in the strongest phenotypes (reviewed in [11]).…”