Behavioral characterization of Capn15 conditional knockout mice.

Calpain 15 (CAPN15) is an intracellular cysteine protease belonging to the non-classical small optic lobe (SOL) family of calpains, which has an important role in development. Loss of Capn15 in mice leads to developmental eye anomalies and volumetric changes in the brain. Human individuals with biallelic variants in CAPN15 have developmental delay, neurodevelopmental disorders, as well as congenital malformations. In Aplysia, a reductionist model to study learning and memory, SOL calpain is important for non-associative long-term facilitation, the cellular analog of sensitization behavior. However, how CAPN15 is involved in adult behavior or learning and memory in vertebrates is unknown. Here, using Capn15 conditional knockout mice, we show that loss of the CAPN15 protein in excitatory forebrain neurons reduces self-grooming and marble burying, decreases performance in the accelerated roto-rod and reduces pre-tone freezing after strong fear conditioning. Thus, CAPN15 plays a role in regulating behavior in the adult mouse.

The molecular diversity of plasticity mechanisms underlying memory: An evolutionary perspective.

Experience triggers molecular cascades in organisms (learning) that lead to alterations (memory) to allow the organism to change its behavior based on experience. Understanding the molecular mechanisms underlying memory, particularly in the nervous system of animals, has been an exciting scientific challenge for neuroscience. We review what is known about forms of neuronal plasticity that underlie memory highlighting important issues in the field: (1) the importance of being able to measure how neurons are activated during learning to identify the form of plasticity that underlies memory, (2) the many distinct forms of plasticity important for memories that naturally decay both within and between organisms, and (3) unifying principles underlying the formation and maintenance of long-term memories. Overall, the diversity of molecular mechanisms underlying memories that naturally decay contrasts with more unified molecular mechanisms implicated in long-lasting changes. Despite many advances, important questions remain as to which mechanisms of neuronal plasticity underlie memory.

MRI of Capn15 Knockout Mice and Analysis of Capn 15 Distribution Reveal Possible Roles in Brain Development and Plasticity.

The Small Optic Lobe (SOL) family of calpains are intracellular cysteine proteases that are expressed in the nervous system and play an important role in neuronal development in both Drosophila, where loss of this calpain leads to the eponymous small optic lobes, and in mouse and human, where loss of this calpain leads to eye anomalies. Some human individuals with biallelic variants in CAPN15 also have developmental delay and autism. However, neither the specific effect of the loss of the Capn15 protein on brain development nor the brain regions where this calpain is expressed in the adult is known. Here we show using small animal MRI that mice with the complete loss of Capn15 have smaller brains overall with larger decreases in the thalamus and subregions of the hippocampus. These losses are not seen in Capn15 conditional knockout (KO) mice where Capn15 is knocked out only in excitatory neurons in the adult. Based on ß-galactosidase expression in an insert strain where lacZ is expressed under the control of the Capn15 promoter, we show that Capn15 is expressed in adult mice, particularly in neurons involved in plasticity such as the hippocampus, lateral amygdala and Purkinje neurons, and partially in other non-characterized cell types. The regions of the brain in the adult where Capn15 is expressed do not correspond well to the regions of the brain most affected by the complete knockout suggesting distinct roles of Capn15 in brain development and adult brain function.

Biallelic variants in the small optic lobe calpain CAPN15 are associated with congenital eye anomalies, deafness and other neurodevelopmental deficits.

Microphthalmia, coloboma and cataract are part of a spectrum of developmental eye disorders in humans affecting ~12 per 100 000 live births. Currently, variants in over 100 genes are known to underlie these conditions. However, at least 40% of affected individuals remain without a clinical genetic diagnosis, suggesting variants in additional genes may be responsible. Calpain 15 (CAPN15) is an intracellular cysteine protease belonging to the non-classical small optic lobe (SOL) family of calpains, an important class of developmental proteins, as yet uncharacterized in vertebrates. We identified five individuals with microphthalmia and/or coloboma from four independent families carrying homozygous or compound heterozygous predicted damaging variants in CAPN15. Several individuals had additional phenotypes including growth deficits, developmental delay and hearing loss. We generated Capn15 knockout mice that exhibited similar severe developmental eye defects, including anophthalmia, microphthalmia and cataract, and diminished growth. We demonstrate widespread Capn15 expression throughout the brain and central nervous system, strongest during early development, and decreasing postnatally. Together, these findings demonstrate a critical role of CAPN15 in vertebrate developmental eye disorders, and may signify a new developmental pathway.

The zinc fingers of the small optic lobes calpain bind polyubiquitin.

The small optic lobes (SOL) calpain is a highly conserved member of the calpain family expressed in the nervous system. A dominant negative form of the SOL calpain inhibited consolidation of one form of synaptic plasticity, non-associative facilitation, in sensory-motor neuronal cultures in Aplysia, presumably by inhibiting cleavage of protein kinase Cs (PKCs) into constitutively active protein kinase Ms (PKMs) (Hu et al. 2017a). SOL calpains have a conserved set of 5-6 N-terminal zinc fingers. Bioinformatic analysis suggests that these zinc fingers could bind to ubiquitin. In this study, we show that both the Aplysia and mouse SOL calpain (also known as Calpain 15) zinc fingers bind ubiquitinated proteins, and we confirm that Aplysia SOL binds poly- but not mono- or diubiquitin. No specific zinc finger is required for polyubiquitin binding. Neither polyubiquitin nor calcium was sufficient to induce purified Aplysia SOL calpain to autolyse or to cleave the atypical PKC to PKM in vitro. In Aplysia, over-expression of the atypical PKC in sensory neurons leads to an activity-dependent cleavage event and an increase in nuclear ubiquitin staining. Activity-dependent cleavage is partially blocked by a dominant negative SOL calpain, but not by a dominant negative classical calpain. The cleaved PKM was stabilized by the dominant negative classical calpain and destabilized by a dominant negative form of the PKM stabilizing protein KIdney/BRAin protein. These studies provide new insight into SOL calpain's function and regulation. Open Data: Materials are available on https://cos.io/our-services/open-science-badges/ https://osf.io/93n6m/.