Piccolo is essential for the maintenance of mouse retina but not cochlear hair cell function

Abstract

Piccolo is a presynaptic protein with high conservation among different species, and the expression of Piccolo is extensive in vertebrates. Recently, a small fragment of Piccolo (Piccolino), arising due to the incomplete splicing of intron 5/6, was found to be present in the synapses of retinas and cochleae. However, the comprehensive function of Piccolo in the retina and cochlea remains unclear. In this study, we generated Piccolo knockout mice using CRISPR-Cas9 technology to explore the function of Piccolo. Unexpectedly, whereas no abnormalities were found in the cochlear hair cells of the mutant mice, significant differences were found in the retinas, in which two layers (the outer nuclear layer and the outer plexiform layer) were absent. Additionally, the amplitudes of electroretinograms were significantly reduced and pigmentation was observed in the fundoscopy of the mutant mouse retinas. The expression levels of Bassoon, a homolog of Piccolo, as well as synapse-associated proteins CtBP1, CtBP2, Kif3A, and Rim1 were down-regulated. The numbers of ribbon synapses in the retinas of the mutant mice were also reduced. Altogether, the phenotype of Piccolo -/- mice resembled the symptoms of retinitis pigmentosa (RP) in humans, suggesting Piccolo might be a candidate gene of RP and indicates Piccolo knockout mice are a good model for elucidating the molecular mechanisms of RP.

Piccolino regulates the architecture of the ribbon at cochlear inner hair cell synapses

“…Piccolino lacks the interactions site to bassoon (Regus-Leidig et al, 2013), and different from conventional synapses, piccolino and bassoon segregate at ribbon-type AZs (Dick et al, 2001; Khimich et al, 2005; Limbach et al, 2011; Michanski et al, 2019; tom Dieck et al, 1998; Wong et al, 2014). In contrast to the mild sensory coding phenotypes of piccolino mutants (the present study, (Li et al, 2021; Müller et al, 2019)), disruption of bassoon strongly alters transmission at afferent synapses of retina and cochlea (Fig 9). The deletion of exon 4 and 5 ( Bsn ΔEx4/5 ) resulted in detached synaptic ribbons in photoreceptors and IHCs and impaired sensory coding assayed at the systems and synaptic levels (Dick et al, 2003; Khimich et al, 2005).…”

“…In these mutants, the piccolino splice variant was not affected (Butola et al, 2017), thus normal sound encoding on the level of ribbon synapses was expected, given that piccolino is the only isoform present at ribbon synapses (Regus-Leidig et al, 2013). Recently, Li and co-workers reported findings in mice with complete loss of piccolo and piccolino demonstrating functional impairments of the retina while hearing seemed unaffected with normal ARBs up to 8 months of age (Li et al, 2021). Our findings on piccolino gene-trap rats contrast the normal hearing in KO mice for which we currently do not have an explanation.…”

“…Piccolino’s absence or reduction had a striking structural impact on photoreceptor ribbon synapses: the altered ribbon shape went along with a reduced number of SVs around the ribbons (Müller et al, 2019; Regus-Leidig et al, 2014). A previous study of the auditory system reported that piccolino KO mice lack functional deficits on the level of auditory brain-stem response (ABR) thresholds (Li et al, 2021).…”

Non-viral gene therapy using RNA interference with PDGFR-α mediated epithelial-mesenchymal transformation for proliferative vitreoretinopathy

“…The electroretinogram (ERG) is a complex electrical response that indicates retinal function [ 91 ]. As shown in Fig.…”

Metabotropic Glutamate Receptors at Ribbon Synapses in the Retina and Cochlea

“…A recent study used CRISPR-Cas9 to generate a complete knock-out of piccolo and piccolino. These mice showed severe anatomical and functional deficits in layers of the outer retina where the photoreceptors are located [ 25 ]. Interestingly, no changes were found in hair cells of the cochlea.…”