Biology subjects

Pangeni, G.

Publications and source records attributed to Pangeni, G..

3 recordsLinked to original sources

bioRxiv2025

RHO1-2 meganuclease gene editing targets human P23H rhodopsin-induced retinitis pigmentosa to rejuvenate rods and maintain cones

Autosomal dominant retinitis pigmentosa (adRP) is an inherited retinal dystrophy characterized by progressive vision loss and eventual blindness. The P23H mutation (proline to histidine substitution at codon 23) in the rhodopsin (RHO) gene represents the most common form of adRP in North Americans. Currently, there is no cure for P23H adRP. Genome editing targeting the mutant RHO allele, leaving a functional wildtype (WT) allele, is an attractive approach for P23H adRP, as only one copy of RHO is needed for normal retinal function. We re-engineered an I-Cre meganuclease, called RHO1-2, to target a 22bp recognition sequence encompassing the mutation responsible for the p.P23H RHO mutation. In vitro, RHO1-2, cuts human P23H RHO but not WT RHO. In vivo, we delivered scAAV5:GRK1:RHO1-2 via subretinal injection in early-stage degeneration using the only large animal model of human p.P23H RHO adRP (TgP23H pigs). We tested RHO1-2 efficacy and durability, on retinal function using full-field electroretinograms and on retinal structure using spectral domain optical coherence tomography and immunohistochemistry. We observe that RHO1-2 treatment: arrests rod photoreceptor degeneration, resurrects rod-driven retinal function that does not exist in untreated TgP23H pigs, restores mislocalized rhodopsin expression and rebuilds rod inner and outer segments (IS/OS). Rod rescue maintains cones. A year after RHO1-2 treatment, we show that TgP23H pigs use rod-driven vision to navigate a maze. Our results demonstrate that genome editing via RHO1-2 meganuclease is a viable treatment to cure human p.P23H RHO adRP. They also suggest that meganuclease-based editors can be effective for other IRDs. One Sentence SummaryEngineered meganuclease, RHO1-2 is a safe and promising therapeutic genome editing approach to cure human p.P23H RHO adRP.

neuroscience↗

bioRxiv2019

Nystagmus in patients with congenital stationary night blindness (CSNB) originates from synchronously firing direction-selective retinal ganglion cells

Congenital nystagmus, involuntary oscillating small eye movements, is commonly thought to originate from aberrant interactions between brainstem nuclei and foveal cortical pathways. Here we investigated whether nystagmus associated with congenital stationary nightblindness (CSNB) can result from primary deficits in the retina. We found that CSNB patients as well as an animal model (nob mice), both of which lack functional nyctalopin protein (NYX, nyx) in ON bipolar cells (ON-BC) at their synapse with photoreceptors, showed oscillating eye movements at a frequency of 4-7Hz. nob ON direction selective ganglion cells (ON-DSGC), which detect global motion and project to the accessory optic system (AOS), oscillated with the same frequency as their eyes. In the dark, individual ganglion cells (GC) oscillated asynchronously, but their oscillations became synchronized by light stimulation. Likewise, both patient and nob mice oscillating eye movements were only present in the light. Retinal pharmacological manipulations that blocked nob ON-DSGC oscillations also eliminated their oscillating eye movements, and retinal pharmacological manipulations that reduced oscillation frequency of nob ON-DSGCs also reduced oscillation frequency of their eye movements. We conclude that, in nob mice, oscillations of retinal ON-DSGCs cause nystagmus with properties similar to those associated with CSNB in humans. These results show that the nob mouse is the first animal model for a form of congenital nystagmus paving the way for development of therapeutic strategies.

neuroscience↗

bioRxiv2018

LRIT3 is required for nyctalopin expression and normal ON and OFF pathway signaling in the retina

At its first synapse, the retina establishes two parallel channels that encode light increments (ON) or decrements (OFF). At the same synapse, changes in photoreceptor glutamate release are sensed by ON bipolar cells (BCs) via the metabotropic glutamate receptor 6 (mGluR6), and OFF BCs via ionotropic BCs, which differ in their synaptic configuration with the photoreceptor terminal. ON BCs form invaginating synapses that bring them in close proximity to presynaptic ribbons and the presumed sole source of glutamate release. OFF bipolar cells form flat contacts distal to the ribbon synapse. We investigated the role of LRIT3 in normal assembly and function of the mGlur6 signaling cascade present in ON BCs. We demonstrate that LRIT3 is required for nyctalopin expression and thus TRPM1 expression and function. Using glutamate imaging, whole-cell electrophysiology, and multi-electrode array extracellular recordings we demonstrate that the loss of LRIT3 impacts both the ON and OFF pathways at the level of the BCs. The effect on ON pathway signaling, a lack of ON BC response, is shared by mutants lacking mGluR6, TRPM1 GPR179 or nyctalopin. The effects on the OFF pathway are unique to LRIT3, and include a decrease in response amplitude of both OFF BC and GCs. Based on these results, we propose a working model where LRIT3 is required for either efficient glutamate release or reuptake from the first retinal synapse.\n\nSIGNIFICANCE STATEMENTAt the first visual synapse, photoreceptor cells signal to two distinct bipolar cell (BC) populations, one characterized by a depolarizing response to light onset (ON or DBCs), the other by a hyperpolarizing response (OFF or HBCs). The DBC light response depends on a G protein-coupled receptor and associated protein complex, known as the signalplex. Mutations in signalplex proteins lead to DBC pathway-specific loss of visual function. Here we show how loss of LRIT3, a previously identified signalplex protein, prevents functional assembly of the DBC signalplex and alters visual function in both ON and OFF signaling pathways. Thus, our results indicate that the function of LRIT3 at this first synapse extends beyond assembly of the DBC signalplex.

neuroscience↗