Biology subjects

Jovanovic, K.

Publications and source records attributed to Jovanovic, K..

3 recordsLinked to original sources

bioRxiv2023

Eupatilin improves cilia defects in human CEP290 ciliopathy models

The photoreceptor outer segment is a highly specialized primary cilium essential for phototransduction and vision. Biallelic pathogenic variants in the cilia-associated gene CEP290 cause non-syndromic Leber congenital amaurosis 10 (LCA10) and syndromic diseases, where the retina is also affected. While RNA antisense oligonucleotides and gene editing are potential treatment options for the common deep intronic variant c.2991+1655A>G in CEP290, there is a need for variant-independent approaches that could be applied to a broader spectrum of ciliopathies. Here, we generated several distinct human models of CEP290-related retinal disease and investigated the effects of the flavonoid eupatilin as a potential treatment. Eupatilin improved cilium formation and length in CEP290 LCA10 patient-derived fibroblasts, in gene-edited CEP290 knockout (CEP290 KO) RPE1 cells, and in both CEP290 LCA10 and CEP290 KO iPSCs-derived retinal organoids. Furthermore, eupatilin reduced rhodopsin retention in the outer nuclear layer of CEP290 LCA10 retinal organoids. Eupatilin altered gene transcription in retinal organoids, by modulating the expression of rhodopsin, and by targeting cilia and synaptic plasticity pathways. This work sheds light into the mechanism of action of eupatilin, and supports its potential as a variant-independent approach for CEP290-associated ciliopathies. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=156 SRC="FIGDIR/small/536565v1_ufig1.gif" ALT="Figure 1"> View larger version (39K): org.highwire.dtl.DTLVardef@c2d45borg.highwire.dtl.DTLVardef@103c8b5org.highwire.dtl.DTLVardef@16e025eorg.highwire.dtl.DTLVardef@15d8be7_HPS_FORMAT_FIGEXP M_FIG C_FIG

neuroscience↗

bioRxiv2021

The role of Musashi-1 in CEP290 c.2991+1655A>G cryptic exon splicing in Leber Congenital Amaurosis

Human photoreceptors maximise alternative exon splicing to generate a unique set of gene isoforms. Conversely, the inclusion of a cryptic exon caused by the c.2991+1655A>G deep intronic change in CEP290 occurs in the human retina leading to Leber Congenital Amaurosis (LCA10). The RNA-binding protein Musashi-1 (MSI1) is a key component of alternative splicing in the developing mouse retina. Here we investigated the role of MSI1 in human photoreceptor-specific splicing and its potential role in CEP290 aberrant splicing disease. Alternative splicing was studied using human induced pluripotent stem cell derived 3D retinal organoid and RPE RNA-seq datasets and several photoreceptor gene isoforms were identified. Their temporal expression was resolved in control 3D retinal organoids in comparison to development and differentiation markers. Morpholino knockdown of MSI1 in control retinal organoids reduced the expression of several photoreceptor differentiation markers and the inclusion of photoreceptor-specific exons. Nonetheless, MSI1 knockdown in homozygous CEP290 c.2991+1655A>G LCA10 retinal organoids did not affect the inclusion of the LCA10-associated cryptic exon. These results show that while MSI1 is important for photoreceptor alternative splicing and homeostasis, it is not a major driver of the recognition of the CEP290 cryptic splice site and the manifestation of LCA10. HIGHLIGHTS{blacksquare} The human retina expresses a unique set of gene isoforms {blacksquare}Musashi-1 regulates alternative splicing in 3D human retinal organoids {blacksquare}Musashi-1 knockdown in 3D retinal organoids affects gene splicing and homeostasis in photoreceptors {blacksquare}Musashi-1 may regulate alternative splicing of cryptic exons in retina but not in LCA10

cell biology↗

bioRxiv2020

Modelling and rescue of RP2 Retinitis Pigmentosa using iPSC Derived Retinal Organoids

RP2 mutations cause a severe form of X-linked retinitis pigmentosa (XLRP). The mechanism of RP2 associated retinal degeneration in humans is unclear, and animal models of RP2 XLRP do not recapitulate this severe phenotype. Here, we developed gene edited isogenic RP2 knock-out (RP2 KO) induced pluripotent stem cells (iPSC) and RP2 patient derived iPSC to produce 3D retinal organoids as a human retinal disease model. Strikingly, the RP2 KO and RP2 patient derived organoids showed a peak in rod photoreceptor cell death at day 150 (D150) with subsequent thinning of the organoid outer nuclear layer (ONL) by D180 of culture. AAV mediated gene augmentation with human RP2 rescued the degeneration phenotype of the RP2 KO organoids, to prevent ONL thinning and restore rhodopsin expression. Notably, these data show that 3D retinal organoids can be used to model photoreceptor degeneration and test potential therapies to prevent photoreceptor cell death.

neuroscience↗