Publications by authors named "Elisa Molinari"

63 Publications

A discarded synonymous variant in NPHP3 explains nephronophthisis and congenital hepatic fibrosis in several families.

Hum Mutat 2021 Jul 2. Epub 2021 Jul 2.

Renal Services, Newcastle Upon Tyne Hospitals NHS Foundation Trust, Newcastle upon Tyne, UK.

Half of patients with a ciliopathy syndrome remain unsolved after initial analysis of whole exome sequencing (WES) data, highlighting the need for improved variant filtering and annotation. By candidate gene curation of WES data, combined with homozygosity mapping, we detected a homozygous predicted synonymous allele in NPHP3 in two children with hepatorenal fibrocystic disease from a consanguineous family. Analyses on patient-derived RNA shows activation of a cryptic mid-exon splice donor leading to frameshift. Remarkably, the same rare variant was detected in four additional families with hepatorenal disease from UK, US, and Saudi patient cohorts and in addition, another synonymous NPHP3 variant was identified in an unsolved case from the Genomics England 100,000 Genomes data set. We conclude that synonymous NPHP3 variants, not reported before and discarded by pathogenicity pipelines, solved several families with a ciliopathy syndrome. These findings prompt careful reassessment of synonymous variants, especially if they are rare and located in candidate genes.
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http://dx.doi.org/10.1002/humu.24251DOI Listing
July 2021

Gene and epigenetic editing in the treatment of primary ciliopathies.

Prog Mol Biol Transl Sci 2021 6;182:353-401. Epub 2021 Mar 6.

Translational and Clinical Research Institute, Faculty of Medical Sciences, Newcastle University, International Centre for Life, Central Parkway, Newcastle upon Tyne, United Kingdom; Renal Services, The Newcastle Hospitals NHS Foundation Trust, Newcastle upon Tyne, United Kingdom; NIHR Newcastle Biomedical Research Centre, Newcastle upon Tyne, United Kingdom. Electronic address:

Primary ciliopathies are inherited human disorders that arise from mutations in ciliary genes. They represent a spectrum of severe, incurable phenotypes, differentially involving several organs, including the kidney and the eye. The development of gene-based therapies is opening up new avenues for the treatment of ciliopathies. Particularly attractive is the possibility of correcting in situ the causative genetic mutation, or pathological epigenetic changes, through the use of gene editing tools. Due to their versatility and efficacy, CRISPR/Cas-based systems represent the most promising gene editing toolkit for clinical applications. However, delivery and specificity issues have so far held back the translatability of CRISPR/Cas-based therapies into clinical practice, especially where systemic administration is required. The eye, with its characteristics of high accessibility and compartmentalization, represents an ideal target for in situ gene correction. Indeed, studies for the evaluation of a CRISPR/Cas-based therapy for in vivo gene correction to treat a retinal ciliopathy have reached the clinical stage. Further technological advances may be required for the development of in vivo CRISPR-based treatments for the kidney. We discuss here the possibilities and the challenges associated to the implementation of CRISPR/Cas-based therapies for the treatment of primary ciliopathies with renal and retinal phenotypes.
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http://dx.doi.org/10.1016/bs.pmbts.2021.01.027DOI Listing
March 2021

Genetic compensation for cilia defects in cep290 mutants by upregulation of cilia-associated small GTPases.

J Cell Sci 2021 Jul 22;134(14). Epub 2021 Jul 22.

Department of Medicine, Nephrology Division, Massachusetts General Hospital, 149 13th Street, Charlestown, MA 02129, USA.

Mutations in CEP290 (also known as NPHP6), a large multidomain coiled coil protein, are associated with multiple cilia-associated syndromes. Over 130 CEP290 mutations have been linked to a wide spectrum of human ciliopathies, raising the question of how mutations in a single gene cause different disease syndromes. In zebrafish, the expressivity of cep290 deficiencies were linked to the type of genetic ablation: acute cep290 morpholino knockdown caused severe cilia-related phenotypes, whereas deficiencies in a CRISPR/Cas9 genetic mutant were restricted to photoreceptor defects. Here, we show that milder phenotypes in genetic mutants were associated with the upregulation of genes encoding the cilia-associated small GTPases arl3, arl13b and unc119b. Upregulation of UNC119b was also observed in urine-derived renal epithelial cells from human Joubert syndrome CEP290 patients. Ectopic expression of arl3, arl13b and unc119b in cep290 morphant zebrafish embryos rescued Kupffer's vesicle cilia and partially rescued photoreceptor outer segment defects. The results suggest that genetic compensation by upregulation of genes involved in a common subcellular process, lipidated protein trafficking to cilia, may be a conserved mechanism contributing to genotype-phenotype variations observed in CEP290 deficiencies. This article has an associated First Person interview with the first author of the paper.
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http://dx.doi.org/10.1242/jcs.258568DOI Listing
July 2021

Evidence of ideal excitonic insulator in bulk MoS under pressure.

Proc Natl Acad Sci U S A 2021 Mar;118(13)

Consiglio Nazionale delle Ricerche - Istituto Nanoscienze, 41125 Modena, Italy;

Spontaneous condensation of excitons is a long-sought phenomenon analogous to the condensation of Cooper pairs in a superconductor. It is expected to occur in a semiconductor at thermodynamic equilibrium if the binding energy of the excitons-electron (e) and hole (h) pairs interacting by Coulomb force-overcomes the band gap, giving rise to a new phase: the "excitonic insulator" (EI). Transition metal dichalcogenides are excellent candidates for the EI realization because of reduced Coulomb screening, and indeed a structural phase transition was observed in few-layer systems. However, previous work could not disentangle to which extent the origin of the transition was in the formation of bound excitons or in the softening of a phonon. Here we focus on bulk [Formula: see text] and demonstrate theoretically that at high pressure it is prone to the condensation of genuine excitons of finite momentum, whereas the phonon dispersion remains regular. Starting from first-principles many-body perturbation theory, we also predict that the self-consistent electronic charge density of the EI sustains an out-of-plane permanent electric dipole moment with an antiferroelectric texture in the layer plane: At the onset of the EI phase, those optical phonons that share the exciton momentum provide a unique Raman fingerprint for the EI formation. Finally, we identify such fingerprint in a Raman feature that was previously observed experimentally, thus providing direct spectroscopic confirmation of an ideal excitonic insulator phase in bulk [Formula: see text] above 30 GPa.
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http://dx.doi.org/10.1073/pnas.2010110118DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8020749PMC
March 2021

Update of genetic variants in CEP120 and CC2D2A-With an emphasis on genotype-phenotype correlations, tissue specific transcripts and exploring mutation specific exon skipping therapies.

Mol Genet Genomic Med 2021 Jan 24:e1603. Epub 2021 Jan 24.

Translational and Clinical Research Institute, Faculty of Medical Sciences, Newcastle University, Newcastle Upon Tyne, UK.

Background: Mutations in ciliary genes cause a spectrum of both overlapping and distinct clinical syndromes (ciliopathies). CEP120 and CC2D2A are paradigmatic examples for this genetic heterogeneity and pleiotropy as mutations in both cause Joubert syndrome but are also associated with skeletal ciliopathies and Meckel syndrome, respectively. The molecular basis for this phenotypical variability is not understood but basal exon skipping likely contributes to tolerance for deleterious mutations via tissue-specific preservation of the amount of expressed functional protein.

Methods: We systematically reviewed and annotated genetic variants and clinical presentations reported in CEP120- and CC2D2A-associated disease and we combined in silico and ex vivo approaches to study tissue-specific transcripts and identify molecular targets for exon skipping.

Results: We confirmed more severe clinical presentations associated with truncating CC2D2A mutations. We identified and confirmed basal exon skipping in the kidney, with possible relevance for organ-specific disease manifestations. Finally, we proposed a multimodal approach to classify exons amenable to exon skipping. By mapping reported variants, 14 truncating mutations in 7 CC2D2A exons were identified as potentially rescuable by targeted exon skipping, an approach that is already in clinical use for other inherited human diseases.

Conclusion: Genotype-phenotype correlations for CC2D2A support the deleteriousness of null alleles and CC2D2A, but not CEP120, offers potential for therapeutic exon skipping approaches.
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http://dx.doi.org/10.1002/mgg3.1603DOI Listing
January 2021

Cell preservation methods and its application to studying rare disease.

Mol Cell Probes 2021 04 9;56:101694. Epub 2021 Jan 9.

Translational and Clinical Research Institute, Faculty of Medical Sciences, Newcastle University, International Centre for Life, Central Parkway, Newcastle Upon Tyne, NE1 3BZ, UK; Renal Services, The Newcastle Upon Tyne Hospitals NHS Foundations Trust, Newcastle Upon Tyne, NE7 7DN, UK; NIHR Newcastle Biomedical Research Centre, Newcastle University, Newcastle Upon Tyne, Tyne and Wear, NE45PL, UK. Electronic address:

The ability to preserve and transport human cells in a stable medium over long distances is critical to collaborative efforts and the advancement of knowledge in the study of human disease. This is particularly important in the study of rare diseases. Recently, advancements in the understanding of renal ciliopathies has been achieved via the use of patient urine-derived cells (UDCs). However, the traditional method of cryopreservation, although considered as the gold standard, can result in decreased sample viability of many cell types, including UDCs. Delays in transportation can have devastating effects upon the viability of samples, and may even result in complete destruction of cells following evaporation of dry ice or liquid nitrogen, leaving samples in cryoprotective agents, which are cytotoxic at room temperature. The loss of any patient sample in this manner is detrimental to research, however it is even more so when samples are from patients with a rare disease. In order to overcome the associated limitations of traditional practices, new methods of preservation and shipment, including cell encapsulation within hydrogels, and transport in specialised devices are continually being investigated. Here we summarise and compare traditional methods with emerging novel alternatives for the preservation and shipment of cells, and consider the effectiveness of such methods for use with UDCs to further enable the study and understanding of kidney diseases.
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http://dx.doi.org/10.1016/j.mcp.2021.101694DOI Listing
April 2021

Intermolecular conical intersections in molecular aggregates.

Nat Nanotechnol 2021 01 16;16(1):63-68. Epub 2020 Nov 16.

Institut für Physik and Center of Interface Science, Carl von Ossietzky Universität, Oldenburg, Germany.

Conical intersections (CoIns) of multidimensional potential energy surfaces are ubiquitous in nature and control pathways and yields of many photo-initiated intramolecular processes. Such topologies can be potentially involved in the energy transport in aggregated molecules or polymers but are yet to be uncovered. Here, using ultrafast two-dimensional electronic spectroscopy (2DES), we reveal the existence of intermolecular CoIns in molecular aggregates relevant for photovoltaics. Ultrafast, sub-10-fs 2DES tracks the coherent motion of a vibrational wave packet on an optically bright state and its abrupt transition into a dark state via a CoIn after only 40 fs. Non-adiabatic dynamics simulations identify an intermolecular CoIn as the source of these unusual dynamics. Our results indicate that intermolecular CoIns may effectively steer energy pathways in functional nanostructures for optoelectronics.
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http://dx.doi.org/10.1038/s41565-020-00791-2DOI Listing
January 2021

Use of patient derived urine renal epithelial cells to confirm pathogenicity of PKHD1 alleles.

BMC Nephrol 2020 10 15;21(1):435. Epub 2020 Oct 15.

Translational and Clinical Research Institute, Faculty of Medical Sciences, Newcastle University, International Centre for Life, Central Parkway, Newcastle upon Tyne, NE1 3BZ, UK.

Background: PKHD1 is the main genetic cause of autosomal recessive polycystic kidney disease (ARPKD), a hereditary hepato-renal fibrocystic disorder which is the most important cause of end-stage renal disease during early childhood. ARPKD can also present in adulthood with milder phenotypes. In this study, we describe a 24-year-old woman with atypical polycystic kidney, no family history of renal disease and no obvious extra-renal manifestations who was referred for genetic investigation.

Methods: We used a combination of next generation sequencing, Sanger sequencing and RNA and microscopy studies performed on urine-derived renal epithelial cells (URECs) to provide a genetic diagnosis of ARPKD.

Results: A next generation sequencing panel of cystic ciliopathy genes allowed the identification of two heterozygous sequence changes in PKHD1 (c.6900C > T; p.(Asn2300=) and c.7964A > C; p.(His2655Pro)). The pathogenicity of the synonymous PKHD1 variant is not clear and requires RNA studies, which cannot be carried out efficiently on RNA extracted from proband blood, due to the low expression levels of PKHD1 in lymphocytes. Using URECs as a source of kidney-specific RNA, we show that PKHD1 is alternatively spliced around exon 43, both in control and proband URECs. The variant p.(Asn2300=) shifts the expression ratio in favour of a shorter, out-of-frame transcript. To further study the phenotypic consequence of these variants, we investigated the ciliary phenotype of patient URECs, which were abnormally elongated and presented multiple blebs along the axoneme.

Conclusions: We confirm the power of URECs as a tool for functional studies on candidate variants in inherited renal disease, especially when the expression of the gene of interest is restricted to the kidney and we describe, for the first time, ciliary abnormalities in ARPKD patient cells.
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http://dx.doi.org/10.1186/s12882-020-02094-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7559414PMC
October 2020

Vibrational signature of the graphene nanoribbon edge structure from high-resolution electron energy-loss spectroscopy.

Nanoscale 2020 Oct;12(38):19681-19688

Dipartimento di Scienze Fisiche, Informatiche e Matematiche (FIM), Università degli Studi di Modena e Reggio Emilia, 41125, Modena, Italy. and Centro S3, Istituto Nanoscienze CNR-NANO, 41125, Modena, Italy.

Bottom-up approaches exploiting on-surface synthesis reactions allow atomic-scale precision in the fabrication of graphene nanoribbons (GNRs); this is essential for their technological applications since their unique electronic and optical properties are largely controlled by the specific edge structure. By means of a combined experimental-theoretical investigation of some prototype GNRs, we show here that high-resolution electron energy-loss spectroscopy (HREELS) can be successfully employed to fingerprint the details of the GNR edge structure. In particular, we demonstrate how the features of HREEL vibrational spectra - mainly dictated by edge CH out-of-plane modes - are unambiguously related to the GNR edge structure. Moreover, we single out those modes which are localized at the GNR termini and show how their relative intensity can be related to the average GNR length.
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http://dx.doi.org/10.1039/d0nr05763kDOI Listing
October 2020

Clinical and genetic characteristics of autosomal recessive polycystic kidney disease in Oman.

BMC Nephrol 2020 08 14;21(1):347. Epub 2020 Aug 14.

Translational and Clinical Research Institute, Faculty of Medical Sciences, Newcastle University, Central Parkway, Newcastle upon Tyne, NE1 3BZ, UK.

Background: There is a high prevalence of rare genetic disorders in the Middle East, and their study provides unique clinical and genetic insights. Autosomal recessive polycystic kidney disease (ARPKD) is one of the leading causes of kidney and liver-associated morbidity and mortality in Oman. We describe the clinical and genetic profile of cohort of ARPKD patients.

Methods: We studied patients with a clinical diagnosis of ARPKD (n = 40) and their relatives (parents (n = 24) and unaffected siblings (n = 10)) from 32 apparently unrelated families, who were referred to the National Genetic Centre in Oman between January 2015 and December 2018. Genetic analysis of PKHD1 if not previously known was performed using targeted exon PCR of known disease alleles and Sanger sequencing.

Results: A clinical diagnosis of ARPKD was made prenatally in 8 patients, 21 were diagnosed during infancy (0-1 year), 9 during early childhood (2-8 years) and 2 at later ages (9-13 years). Clinical phenotypes included polycystic kidneys, hypertension, hepatic fibrosis and splenomegaly. Twenty-four patients had documented chronic kidney disease (median age 3 years). Twenty-four out of the 32 families had a family history suggesting an autosomal recessive pattern of inherited kidney disease, and there was known consanguinity in 21 families (66%). A molecular genetic diagnosis with biallelic PKHD1 mutations was known in 18 patients and newly identified in 20 other patients, totalling 38 patients from 30 different families. Two unrelated patients remained genetically unsolved. The different PKHD1 missense pathogenic variants were: c.107C > T, p.(Thr36Met); c.406A > G, p.(Thr136Ala); c.4870C > T, p.(Arg1624Trp) and c.9370C > T, p.(His3124Tyr) located in exons 3, 6, 32 and 58, respectively. The c.406A > G, p.(Thr136Ala) missense mutation was detected homozygously in one family and heterozygously with a c.107C > T, p.(Thr36Met) allele in 5 other families. Overall, the most commonly detected pathogenic allele was c.107C > T; (Thr36Met), which was seen in 24 families.

Conclusions: Molecular genetic screening of PKHD1 in clinically suspected ARPKD cases produced a high diagnostic rate. The limited number of PKHD1 missense variants identified in ARPKD cases suggests these may be common founder alleles in the Omani population. Cost effective targeted PCR analysis of these specific alleles can be a useful diagnostic tool for future cases of suspected ARPKD in Oman.
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http://dx.doi.org/10.1186/s12882-020-02013-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7429752PMC
August 2020

Disease Modeling To Understand the Pathomechanisms of Human Genetic Kidney Disorders.

Clin J Am Soc Nephrol 2020 06 5;15(6):855-872. Epub 2020 Mar 5.

Faculty of Medical Sciences, Translational and Clinical Research Institute, International Centre for Life, Newcastle University, Newcastle upon Tyne, United Kingdom

The class of human genetic kidney diseases is extremely broad and heterogeneous. Accordingly, the range of associated disease phenotypes is highly variable. Many children and adults affected by inherited kidney disease will progress to ESKD at some point in life. Extensive research has been performed on various different disease models to investigate the underlying causes of genetic kidney disease and to identify disease mechanisms that are amenable to therapy. We review some of the research highlights that, by modeling inherited kidney disease, contributed to a better understanding of the underlying pathomechanisms, leading to the identification of novel genetic causes, new therapeutic targets, and to the development of new treatments. We also discuss how the implementation of more efficient genome-editing techniques and tissue-culture methods for kidney research is providing us with personalized models for a precision-medicine approach that takes into account the specificities of the patient and the underlying disease. We focus on the most common model systems used in kidney research and discuss how, according to their specific features, they can differentially contribute to biomedical research. Unfortunately, no definitive treatment exists for most inherited kidney disorders, warranting further exploitation of the existing disease models, as well as the implementation of novel, complex, human patient-specific models to deliver research breakthroughs.
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http://dx.doi.org/10.2215/CJN.08890719DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7274277PMC
June 2020

A monolayer transition-metal dichalcogenide as a topological excitonic insulator.

Nat Nanotechnol 2020 May 2;15(5):367-372. Epub 2020 Mar 2.

CNR-NANO, Modena, Italy.

Monolayer transition-metal dichalcogenides in the T' phase could enable the realization of the quantum spin Hall effect at room temperature, because they exhibit a prominent spin-orbit gap between inverted bands in the bulk. Here we show that the binding energy of electron-hole pairs excited through this gap is larger than the gap itself in the paradigmatic case of monolayer T' MoS, which we investigate from first principles using many-body perturbation theory. This paradoxical result hints at the instability of the T' phase in the presence of spontaneous generation of excitons, and we predict that it will give rise to a reconstructed 'excitonic insulator' ground state. Importantly, we show that in this monolayer system, topological and excitonic order cooperatively enhance the bulk gap by breaking the crystal inversion symmetry, in contrast to the case of bilayers where the frustration between the two orders is relieved by breaking time reversal symmetry. The excitonic topological insulator is distinct from the bare topological phase because it lifts the band spin degeneracy, which results in circular dichroism. A moderate biaxial strain applied to the system leads to two additional excitonic phases, different in their topological character but both ferroelectric as an effect of electron-electron interaction.
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http://dx.doi.org/10.1038/s41565-020-0650-4DOI Listing
May 2020

Mouse genetics reveals Barttin as a genetic modifier of Joubert syndrome.

Proc Natl Acad Sci U S A 2020 01 26;117(2):1113-1118. Epub 2019 Dec 26.

Translational and Clinical Research Institute, Faculty of Medical Sciences, Newcastle upon Tyne NE1 3BZ, United Kingdom;

Genetic and phenotypic heterogeneity and the lack of sufficiently large patient cohorts pose a significant challenge to understanding genetic associations in rare disease. Here we identify (alias ) as a genetic modifier of cystic kidney disease in Joubert syndrome, using a -deficient mouse model to recapitulate the phenotypic variability observed in patients by mixing genetic backgrounds in a controlled manner and performing genome-wide analysis of these mice. Experimental down-regulation of in the parental mouse strain phenocopied the severe cystic kidney phenotype. A common polymorphism within human significantly associates with kidney disease severity in a patient cohort with mutations. The striking phenotypic modifications we describe are a timely reminder of the value of mouse models and highlight the significant contribution of genetic background. Furthermore, if appropriately managed, this can be exploited as a powerful tool to elucidate mechanisms underlying human disease heterogeneity.
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http://dx.doi.org/10.1073/pnas.1912602117DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6969532PMC
January 2020

A CEP104-CSPP1 Complex Is Required for Formation of Primary Cilia Competent in Hedgehog Signaling.

Cell Rep 2019 08;28(7):1907-1922.e6

Department of Radiation Biology, Institute for Cancer Research, OUH-Norwegian Radium Hospital, Oslo, Norway. Electronic address:

CEP104 is an evolutionarily conserved centrosomal and ciliary tip protein. CEP104 loss-of-function mutations are reported in patients with Joubert syndrome, but their function in the etiology of ciliopathies is poorly understood. Here, we show that cep104 silencing in zebrafish causes cilia-related manifestations: shortened cilia in Kupffer's vesicle, heart laterality, and cranial nerve development defects. We show that another Joubert syndrome-associated cilia tip protein, CSPP1, interacts with CEP104 at microtubules for the regulation of axoneme length. We demonstrate in human telomerase reverse transcriptase-immortalized retinal pigmented epithelium (hTERT-RPE1) cells that ciliary translocation of Smoothened in response to Hedgehog pathway stimulation is both CEP104 and CSPP1 dependent. However, CEP104 is not required for the ciliary recruitment of CSPP1, indicating that an intra-ciliary CEP104-CSPP1 complex controls axoneme length and Hedgehog signaling competence. Our in vivo and in vitro analyses of CEP104 define its interaction with CSPP1 as a requirement for the formation of Hedgehog signaling-competent cilia, defects that underlie Joubert syndrome.
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http://dx.doi.org/10.1016/j.celrep.2019.07.025DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6702141PMC
August 2019

Targeted exon skipping rescues ciliary protein composition defects in Joubert syndrome patient fibroblasts.

Sci Rep 2019 07 25;9(1):10828. Epub 2019 Jul 25.

Institute of Genetic Medicine, Newcastle University, International Centre for Life, Central Parkway, Newcastle upon Tyne, NE1 3BZ, United Kingdom.

Joubert syndrome (JBTS) is an incurable multisystem ciliopathy syndrome. The most commonly mutated gene in JBTS patients with a cerebello-retinal-renal phenotype is CEP290 (alias JBTS5). The encoded CEP290 protein localises to the proximal end of the primary cilium, in the transition zone, where it controls ciliary protein composition and signalling. We examined primary cilium structure and composition in fibroblast cells derived from homozygous and compound heterozygous JBTS5 patients with nonsense mutations in CEP290 and show that elongation of cilia, impaired ciliogenesis and ciliary composition defects are typical features in JBTS5 cells. Targeted skipping of the mutated exon c.5668 G > T using antisense oligonucleotide (ASO) therapy leads to restoration of CEP290 protein expression and functions at the transition zone in homozygous and compound heterozygous JBTS5 cells, allowing a rescue of both cilia morphology and ciliary composition. This study, by demonstrating that targeted exon skipping is able to rescue ciliary protein composition defects, provides functional evidence for the efficacy of this approach in the treatment of JBTS.
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http://dx.doi.org/10.1038/s41598-019-47243-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6658666PMC
July 2019

Tailoring optical properties and stimulated emission in nanostructured polythiophene.

Sci Rep 2019 May 14;9(1):7370. Epub 2019 May 14.

IFN-CNR, c\o Dipartimento di Fisica, di Milano, I-20132, Milano, Italy.

Polythiophenes are the most widely utilized semiconducting polymers in organic electronics, but they are scarcely exploited in photonics due to their high photo-induced absorption caused by interchain polaron pairs, which prevents the establishment of a window of net optical gain. Here we study the photophysics of poly(3-hexylthiophene) configured with different degrees of supramolecular ordering, spin-coated thin films and templated nanowires, and find marked differences in their optical properties. Transient absorption measurements evidence a partially-polarized stimulated emission band in the nanowire samples, in contrast with the photo-induced absorption band observed in spin-coated thin films. In combination with theoretical modeling, our experimental results reveal the origin of the primary photoexcitations dominating the dynamics for different supramolecular ordering, with singlet excitons in the nanostructured samples superseding the presence of polaron pairs, which are present in the disordered films. Our approach demonstrates a viable strategy to direct optical properties through structural control, and the observation of optical gain opens the possibility to the use of polythiophene nanostructures as building blocks of organic optical amplifiers and active photonic devices.
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http://dx.doi.org/10.1038/s41598-019-43719-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6517583PMC
May 2019

A case of ocular cystinosis associated with two potentially severe CTNS mutations.

Ophthalmic Genet 2019 04 6;40(2):157-160. Epub 2019 Apr 6.

b Institute of Genetic Medicine, International Centre for Life, Newcastle University , Newcastle upon Tyne , UK.

Background: Ocular cystinosis is a rare autosomal recessive disorder caused by one severe and one mild mutation in the CTNS gene. It is characterised by cystine deposition within the cornea and conjunctiva however, the kidneys are not affected. We report a case of ocular cystinosis caused by two potentially severe CTNS mutations and discuss the possible mechanism of renal sparing.

Methods: This is an observational case report of the proband and her unaffected relatives. All subjects underwent ophthalmic examination, whilst in the proband, In vivo laser scanning confocal microscopy was used to demonstrate cystine crystals within her corneas and conjunctiva. Genetic diagnosis was confirmed by DNA sequencing of the proband and the segregation of the mutations was established in her relatives. RT-PCR of leukocyte RNA was undertaken to determine if aberrant splicing of the CTNS gene was taking place Results: The proband was found to have cystine crystals limited to the anterior corneal stroma and the conjunctiva. Sequencing of the proband's CTNS gene found her to be a compound heterozygote for a 27bp deletion in exon8/intron 8 (c.559_561 + 24del) and a novel c.635C>T variant in exon 9 that is predicted be pathogenic and to result in the substitution of alanine with valine at amino acid position 212 (p.Ala212Val), which is within the 3 transmembrane spanning domain of the CTNS protein. Examination of the proband's leukocyte RNA failed to demonstrate any aberrant CTNS gene splicing.

Conclusion: We present a case of ocular cystinosis caused by two potentially severe CTNS gene mutations. The lack of renal involvement may be due to localised (ocular) aberrant CTNS RNA splicing.
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http://dx.doi.org/10.1080/13816810.2019.1592198DOI Listing
April 2019

Using zebrafish to study the function of nephronophthisis and related ciliopathy genes.

F1000Res 2018 25;7:1133. Epub 2018 Jul 25.

Institute of Genetic Medicine, Newcastle University, Newcastle upon Tyne, NE1 3BZ, UK.

Zebrafish are a valuable vertebrate model in which to study development and characterize genes involved in cystic kidney disease. Zebrafish embryos and larvae are transparent, allowing non-invasive imaging during their rapid development, which takes place over the first 72 hours post fertilisation. Gene-specific knockdown of nephronophthisis-associated genes leads to ciliary phenotypes which can be assessed in various developmental structures. Here we describe in detail the methods used for imaging cilia within Kupffer's vesicle to assess nephronophthisis and related ciliopathy phenotypes.
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http://dx.doi.org/10.12688/f1000research.15511.2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6127739PMC
September 2019

Targeted exon skipping of a mutation rescues Joubert syndrome phenotypes in vitro and in a murine model.

Proc Natl Acad Sci U S A 2018 12 16;115(49):12489-12494. Epub 2018 Nov 16.

Institute of Genetic Medicine, Newcastle University, International Centre for Life, Central Parkway, Newcastle upon Tyne NE1 3BZ, United Kingdom;

Genetic treatments of renal ciliopathies leading to cystic kidney disease would provide a real advance in current therapies. Mutations in underlie a ciliopathy called Joubert syndrome (JBTS). Human disease phenotypes include cerebral, retinal, and renal disease, which typically progresses to end stage renal failure (ESRF) within the first two decades of life. While currently incurable, there is often a period of years between diagnosis and ESRF that provides a potential window for therapeutic intervention. By studying patient biopsies, patient-derived kidney cells, and a mouse model, we identify abnormal elongation of primary cilia as a key pathophysiological feature of -associated JBTS and show that antisense oligonucleotide (ASO)-induced splicing of the mutated exon (41, G1890*) restores protein expression in patient cells. We demonstrate that ASO-induced splicing leading to exon skipping is tolerated, resulting in correct localization of CEP290 protein to the ciliary transition zone, and restoration of normal cilia length in patient kidney cells. Using a gene trap mouse model of JBTS, we show that systemic ASO treatment can reduce the cystic burden of diseased kidneys in vivo. These findings indicate that ASO treatment may represent a promising therapeutic approach for kidney disease in -associated ciliopathy syndromes.
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http://dx.doi.org/10.1073/pnas.1809432115DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6298104PMC
December 2018

Outpatient management of the anemic patient: is it a possible challenge?

Minerva Med 2019 Apr 11;110(2):171-173. Epub 2018 Oct 11.

Department of Internal Medicine, IRCCS AOU San Martino IST, Genoa, Italy.

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http://dx.doi.org/10.23736/S0026-4806.18.05761-0DOI Listing
April 2019

Interaction-Driven Giant Orbital Magnetic Moments in Carbon Nanotubes.

Phys Rev Lett 2018 Sep;121(12):127704

Kavli Institute of Nanoscience, Delft University of Technology, Lorentzweg 1, 2628 CJ Delft, Netherlands.

Carbon nanotubes continue to be model systems for studies of confinement and interactions. This is particularly true in the case of so-called "ultraclean" carbon nanotube devices offering the study of quantum dots with extremely low disorder. The quality of such systems, however, has increasingly revealed glaring discrepancies between experiment and theory. Here, we address the outstanding anomaly of exceptionally large orbital magnetic moments in carbon nanotube quantum dots. We perform low temperature magnetotransport measurements of the orbital magnetic moment and find it is up to 7 times larger than expected from the conventional semiclassical model. Moreover, the magnitude of the magnetic moment monotonically drops with the addition of each electron to the quantum dot directly contradicting the widely accepted shell filling picture of single-particle levels. We carry out quasiparticle calculations, both from first principles and within the effective-mass approximation, and find the giant magnetic moments can only be captured by considering a self-energy correction to the electronic band structure due to electron-electron interactions.
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http://dx.doi.org/10.1103/PhysRevLett.121.127704DOI Listing
September 2018

ARL3 Mutations Cause Joubert Syndrome by Disrupting Ciliary Protein Composition.

Am J Hum Genet 2018 10 27;103(4):612-620. Epub 2018 Sep 27.

Institute of Genetic Medicine, Newcastle University, International Centre for Life, Central Parkway, Newcastle upon Tyne NE1 3BZ, UK; Renal Services, Newcastle Hospitals NHS Foundation Trust, Newcastle upon Tyne NE7 7DN, UK. Electronic address:

Joubert syndrome (JBTS) is a genetically heterogeneous autosomal-recessive neurodevelopmental ciliopathy. We investigated further the underlying genetic etiology of Joubert syndrome by studying two unrelated families in whom JBTS was not associated with pathogenic variants in known JBTS-associated genes. Combined autozygosity mapping of both families highlighted a candidate locus on chromosome 10 (chr10: 101569997-109106128, UCSC Genome Browser hg 19), and exome sequencing revealed two missense variants in ARL3 within the candidate locus. The encoded protein, ADP ribosylation factor-like GTPase 3 (ARL3), is a small GTP-binding protein that is involved in directing lipid-modified proteins into the cilium in a GTP-dependent manner. Both missense variants replace the highly conserved Arg149 residue, which we show to be necessary for the interaction with its guanine nucleotide exchange factor ARL13B, such that the mutant protein is associated with reduced INPP5E and NPHP3 localization in cilia. We propose that ARL3 provides a potential hub in the network of proteins implicated in ciliopathies, whereby perturbation of ARL3 leads to the mislocalization of multiple ciliary proteins as a result of abnormal displacement of lipidated protein cargo.
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http://dx.doi.org/10.1016/j.ajhg.2018.08.015DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6174286PMC
October 2018

Human urine-derived renal epithelial cells provide insights into kidney-specific alternate splicing variants.

Eur J Hum Genet 2018 12 12;26(12):1791-1796. Epub 2018 Jul 12.

Institute of Genetic Medicine, Newcastle University, Central Parkway, Newcastle, NE1 3BZ, UK.

The majority of multi-exon genes undergo alternative splicing to produce different mRNA transcripts and this may occur in a tissue-specific manner. Assessment of mRNA transcripts isolated from blood samples may sometimes be unhelpful in determining the affect on function of putative splice-site variants affecting kidney-specific mRNA transcripts. Here we present data demonstrating the power of using human urine-derived renal epithelial cells (hUREC) as a source of kidney RNA. We report clinical and molecular genetic data from three affected cases from two families all with end-stage renal disease by 15 years of age. In both families, heterozygous variants which are predicted to effect function in NPHP3 were found on one allele, in combination with a synonymous SNV (c.2154C>T; p.Phe718=), 18 base pairs from the exon-intron boundary within exon 15 of NPHP3. The only mRNA transcript amplified from wild-type whole blood showed complete splicing out of exon 15. Urine samples obtained from control subjects and the father of family 2, who carried the synonymous SNV variant, were therefore used to culture hUREC and allowed us to obtain kidney-specific mRNA. Control kidney mRNA showed retention of exon 15, while the mRNA from the patient's father confirmed evidence of a heterozygous alternate splicing of exon 15 of NPHP3. Analysis of RNA derived from hUREC allows for a comparison of kidney-specific and whole-blood RNA transcripts and for assessment of the effect on function of putative splice variants leading to end-stage kidney disease.
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http://dx.doi.org/10.1038/s41431-018-0212-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6244279PMC
December 2018

Bandgap Engineering of Graphene Nanoribbons by Control over Structural Distortion.

J Am Chem Soc 2018 06 15;140(25):7803-7809. Epub 2018 Jun 15.

Max Planck Institute for Polymer Research , Ackermannweg 10 , D-55128 Mainz , Germany.

Among organic electronic materials, graphene nanoribbons (GNRs) offer extraordinary versatility as next-generation semiconducting materials for nanoelectronics and optoelectronics due to their tunable properties, including charge-carrier mobility, optical absorption, and electronic bandgap, which are uniquely defined by their chemical structures. Although planar GNRs have been predominantly considered until now, nonplanarity can be an additional parameter to modulate their properties without changing the aromatic core. Herein, we report theoretical and experimental studies on two GNR structures with "cove"-type edges, having an identical aromatic core but with alkyl side chains at different peripheral positions. The theoretical results indicate that installment of alkyl chains at the innermost positions of the "cove"-type edges can "bend" the peripheral rings of the GNR through steric repulsion between aromatic protons and the introduced alkyl chains. This structural distortion is theoretically predicted to reduce the bandgap by up to 0.27 eV, which is corroborated by experimental comparison of thus synthesized planar and nonplanar GNRs through UV-vis-near-infrared absorption and photoluminescence excitation spectroscopy. Our results extend the possibility of engineering GNR properties, adding subtle structural distortion as a distinct and potentially highly versatile parameter.
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http://dx.doi.org/10.1021/jacs.8b02209DOI Listing
June 2018

Many Genes-One Disease? Genetics of Nephronophthisis (NPHP) and NPHP-Associated Disorders.

Front Pediatr 2017 5;5:287. Epub 2018 Jan 5.

Institute of Genetic Medicine, Newcastle University, Newcastle upon Tyne, United Kingdom.

Nephronophthisis (NPHP) is a renal ciliopathy and an autosomal recessive cause of cystic kidney disease, renal fibrosis, and end-stage renal failure, affecting children and young adults. Molecular genetic studies have identified more than 20 genes underlying this disorder, whose protein products are all related to cilia, centrosome, or mitotic spindle function. In around 15% of cases, there are additional features of a ciliopathy syndrome, including retinal defects, liver fibrosis, skeletal abnormalities, and brain developmental disorders. Alongside, gene identification has arisen molecular mechanistic insights into the disease pathogenesis. The genetic causes of NPHP are discussed in terms of how they help us to define treatable disease pathways including the cyclic adenosine monophosphate pathway, the mTOR pathway, Hedgehog signaling pathways, and DNA damage response pathways. While the underlying pathology of the many types of NPHP remains similar, the defined disease mechanisms are diverse, and a personalized medicine approach for therapy in NPHP patients is likely to be required.
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http://dx.doi.org/10.3389/fped.2017.00287DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5770800PMC
January 2018

Carbon nanotubes as excitonic insulators.

Nat Commun 2017 11 13;8(1):1461. Epub 2017 Nov 13.

CNR-NANO, Via Campi 213a, 41125, Modena, Italy.

Fifty years ago Walter Kohn speculated that a zero-gap semiconductor might be unstable against the spontaneous generation of excitons-electron-hole pairs bound together by Coulomb attraction. The reconstructed ground state would then open a gap breaking the symmetry of the underlying lattice, a genuine consequence of electronic correlations. Here we show that this excitonic insulator is realized in zero-gap carbon nanotubes by performing first-principles calculations through many-body perturbation theory as well as quantum Monte Carlo. The excitonic order modulates the charge between the two carbon sublattices opening an experimentally observable gap, which scales as the inverse of the tube radius and weakly depends on the axial magnetic field. Our findings call into question the Luttinger liquid paradigm for nanotubes and provide tests to experimentally discriminate between excitonic and Mott insulators.
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http://dx.doi.org/10.1038/s41467-017-01660-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5684388PMC
November 2017

A human patient-derived cellular model of Joubert syndrome reveals ciliary defects which can be rescued with targeted therapies.

Hum Mol Genet 2017 12;26(23):4657-4667

Newcastle University, Institute of Genetic Medicine, Central Parkway, Newcastle upon Tyne, NE1 3BZ, UK.

Joubert syndrome (JBTS) is the archetypal ciliopathy caused by mutation of genes encoding ciliary proteins leading to multi-system phenotypes, including a cerebello-retinal-renal syndrome. JBTS is genetically heterogeneous, however mutations in CEP290 are a common underlying cause. The renal manifestation of JBTS is a juvenile-onset cystic kidney disease, known as nephronophthisis, typically progressing to end-stage renal failure within the first two decades of life, thus providing a potential window for therapeutic intervention. In order to increase understanding of JBTS and its associated kidney disease and to explore potential treatments, we conducted a comprehensive analysis of primary renal epithelial cells directly isolated from patient urine (human urine-derived renal epithelial cells, hURECs). We demonstrate that hURECs from a JBTS patient with renal disease have elongated and disorganized primary cilia and that this ciliary phenotype is specifically associated with an absence of CEP290 protein. Treatment with the Sonic hedgehog (Shh) pathway agonist purmorphamine or cyclin-dependent kinase inhibition (using roscovitine and siRNA directed towards cyclin-dependent kinase 5) ameliorated the cilia phenotype. In addition, purmorphamine treatment was shown to reduce cyclin-dependent kinase 5 in patient cells, suggesting a convergence of these signalling pathways. To our knowledge, this is the most extensive analysis of primary renal epithelial cells from JBTS patients to date. It demonstrates the feasibility and power of this approach to directly assess the consequences of patient-specific mutations in a physiologically relevant context and a previously unrecognized convergence of Shh agonism and cyclin-dependent kinase inhibition as potential therapeutic targets.
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http://dx.doi.org/10.1093/hmg/ddx347DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5886250PMC
December 2017

From disease modelling to personalised therapy in patients with mutations.

F1000Res 2017 12;6:669. Epub 2017 May 12.

Institute of Genetic Medicine, Newcastle University, Newcastle, NE1 3BZ, UK.

Mutations that give rise to premature termination codons are a common cause of inherited genetic diseases. When transcripts containing these changes are generated, they are usually rapidly removed by the cell through the process of nonsense-mediated decay. Here we discuss observed changes in transcripts of the centrosomal protein CEP290 resulting not from degradation, but from changes in exon usage. We also comment on a landmark paper (Drivas . Sci Transl Med. 2015) where modelling this process of exon usage may be used to predict disease severity in ciliopathies, and how understanding this process may potentially be used for therapeutic benefit in the future.
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http://dx.doi.org/10.12688/f1000research.11553.1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5482330PMC
May 2017

How To Identify Plasmons from the Optical Response of Nanostructures.

ACS Nano 2017 07 5;11(7):7321-7335. Epub 2017 Jul 5.

ICFO-Institut de Ciencies Fotoniques, The Barcelona Institute of Science and Technology , 08860 Castelldefels, Barcelona, Spain.

A promising trend in plasmonics involves shrinking the size of plasmon-supporting structures down to a few nanometers, thus enabling control over light-matter interaction at extreme-subwavelength scales. In this limit, quantum mechanical effects, such as nonlocal screening and size quantization, strongly affect the plasmonic response, rendering it substantially different from classical predictions. For very small clusters and molecules, collective plasmonic modes are hard to distinguish from other excitations such as single-electron transitions. Using rigorous quantum mechanical computational techniques for a wide variety of physical systems, we describe how an optical resonance of a nanostructure can be classified as either plasmonic or nonplasmonic. More precisely, we define a universal metric for such classification, the generalized plasmonicity index (GPI), which can be straightforwardly implemented in any computational electronic-structure method or classical electromagnetic approach to discriminate plasmons from single-particle excitations and photonic modes. Using the GPI, we investigate the plasmonicity of optical resonances in a wide range of systems including: the emergence of plasmonic behavior in small jellium spheres as the size and the number of electrons increase; atomic-scale metallic clusters as a function of the number of atoms; and nanostructured graphene as a function of size and doping down to the molecular plasmons in polycyclic aromatic hydrocarbons. Our study provides a rigorous foundation for the further development of ultrasmall nanostructures based on molecular plasmonics.
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http://dx.doi.org/10.1021/acsnano.7b03421DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5607458PMC
July 2017

Tracking the coherent generation of polaron pairs in conjugated polymers.

Nat Commun 2016 12 8;7:13742. Epub 2016 Dec 8.

Institut für Physik, Carl von Ossietzky Universität, Oldenburg 26129, Germany.

The optical excitation of organic semiconductors not only generates charge-neutral electron-hole pairs (excitons), but also charge-separated polaron pairs with high yield. The microscopic mechanisms underlying this charge separation have been debated for many years. Here we use ultrafast two-dimensional electronic spectroscopy to study the dynamics of polaron pair formation in a prototypical polymer thin film on a sub-20-fs time scale. We observe multi-period peak oscillations persisting for up to about 1 ps as distinct signatures of vibronic quantum coherence at room temperature. The measured two-dimensional spectra show pronounced peak splittings revealing that the elementary optical excitations of this polymer are hybridized exciton-polaron-pairs, strongly coupled to a dominant underdamped vibrational mode. Coherent vibronic coupling induces ultrafast polaron pair formation, accelerates the charge separation dynamics and makes it insensitive to disorder. These findings open up new perspectives for tailoring light-to-current conversion in organic materials.
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http://dx.doi.org/10.1038/ncomms13742DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5155154PMC
December 2016
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