The LHX2-OTX2 transcriptional regulatory module controls retinal pigmented epithelium differentiation and underlies genetic risk for age-related macular degeneration.

Tissue-specific transcription factors (TFs) control the transcriptome through an association with noncoding regulatory regions (cistromes). Identifying the combination of TFs that dictate specific cell fate, their specific cistromes and examining their involvement in complex human traits remain a major challenge. Here, we focus on the retinal pigmented epithelium (RPE), an essential lineage for retinal development and function and the primary tissue affected in age-related macular degeneration (AMD), a leading cause of blindness. By combining mechanistic findings in stem-cell-derived human RPE, in vivo functional studies in mice and global transcriptomic and proteomic analyses, we revealed that the key developmental TFs LHX2 and OTX2 function together in transcriptional module containing LDB1 and SWI/SNF (BAF) to regulate the RPE transcriptome. Importantly, the intersection between the identified LHX2-OTX2 cistrome with published expression quantitative trait loci, ATAC-seq data from human RPE, and AMD genome-wide association study (GWAS) data, followed by functional validation using a reporter assay, revealed a causal genetic variant that affects AMD risk by altering TRPM1 expression in the RPE through modulation of LHX2 transcriptional activity on its promoter. Taken together, the reported cistrome of LHX2 and OTX2, the identified downstream genes and interacting co-factors reveal the RPE transcription module and uncover a causal regulatory risk single-nucleotide polymorphism (SNP) in the multifactorial common blinding disease AMD.

Shared and organ-specific gene-expression programs during the development of the cochlea and the superior olivary complex.

The peripheral and central auditory subsystems together form a complex sensory network that allows an organism to hear. The genetic programs of the two subsystems must therefore be tightly coordinated during development. Yet, their interactions and common expression pathways have never been systematically explored. MicroRNAs (miRNAs) are short non-coding RNAs that regulate gene expression and are essential for normal development of the auditory system. We performed mRNA and small-RNA sequencing of organs from both auditory subsystems at three critical developmental timepoints (E16, P0, P16) to obtain a comprehensive and unbiased insight of their expression profiles. Our analysis reveals common and organ-specific expression patterns for differentially regulated mRNAs and miRNAs, which could be clustered with a particular selection of functions such as inner ear development, Wnt signalling, K+ transport, and axon guidance, based on gene ontology. Bioinformatics detected enrichment of predicted targets of specific miRNAs in the clusters and predicted regulatory interactions by monitoring opposite trends of expression of miRNAs and their targets. This approach identified six miRNAs as strong regulatory candidates for both subsystems. Among them was miR-96, an established critical factor for proper development in both subsystems, demonstrating the strength of our approach. We suggest that other miRNAs identified by this analysis are also common effectors of proper hearing acquirement. This first combined comprehensive analysis of the developmental program of the peripheral and central auditory systems provides important data and bioinformatics insights into the shared genetic program of the two sensory subsystems and their regulation by miRNAs.

[TUMOR-LIKE EFFECT AS INITIAL PRESENTATION OF PEDIATRIC MULTIPLE SCLEROSIS].

INTRODUCTION: In both children and adults, magnetic resonance imaging of the brain in cases of multiple sclerosis (MS) has typical indications, where one of the key points for differentiating between demyelinating processes and place-taking processes is the fact that most of the lesions that appear in multiple sclerosis do not cause a mass effect or much edema around them. There are several uncommon subtypes of multiple sclerosis that can appear specifically in adolescents, presenting with a stormy clinical course and accompanied by brain lesions that resemble space-occupying lesions. These include Marburg disease, Balò's concentric sclerosis, and tumefactive MS. These unusual presentations raise the question regarding the ability to distinguish between neoplastic and demyelinating processes. In this article we present two case studies that illustrate this diagnostic dilemma and an accompanying literature review.

Expression pattern of cochlear microRNAs in the mammalian auditory hindbrain.

The auditory system comprises the auditory periphery, engaged in sound transduction and the central auditory system, implicated in auditory information processing and perception. Recently, evidence mounted that the mammalian peripheral and central auditory systems share a number of genes critical for proper development and function. This bears implication for auditory rehabilitation and evolution of the auditory system. To analyze to which extent microRNAs (miRNAs) belong to genes shared between both systems, we characterize the expression pattern of 12 cochlea-abundant miRNAs in the central auditory system. Quantitative real-time PCR (qRT-PCR) demonstrated expression of all 12 genes in the cochlea, the auditory hindbrain and the non-auditory prefrontal cortex (PFC) at embryonic stage (E)16 and postnatal stages (P)0 and P30. Eleven of them showed differences in expression between tissues and nine between the developmental time points. Hierarchical cluster analysis revealed that the temporal expression pattern in the auditory hindbrain was more similar to the PFC than to the cochlea. Spatiotemporal expression analysis by RNA in situ hybridization demonstrated widespread expression throughout the cochlear nucleus complex (CNC) and the superior olivary complex (SOC) during postnatal development. Altogether, our data indicate that miRNAs represent a relevant class of genetic factors functioning across the auditory system. Given the importance of gene regulatory network (GRN) components for development, physiology and evolution, the 12 miRNAs provide promising entry points to gain insights into their molecular underpinnings in the auditory system.

Identification and characterization of key long non-coding RNAs in the mouse cochlea.

The auditory system is a complex sensory network with an orchestrated multilayer regulatory programme governing its development and maintenance. Accumulating evidence has implicated long non-coding RNAs (lncRNAs) as important regulators in numerous systems, as well as in pathological pathways. However, their function in the auditory system has yet to be explored. Using a set of specific criteria, we selected four lncRNAs expressed in the mouse cochlea, which are conserved in the human transcriptome and are relevant for inner ear function. Bioinformatic characterization demonstrated a lack of coding potential and an absence of evolutionary conservation that represent properties commonly shared by their class members. RNAscope®  analysis of the spatial and temporal expression profiles revealed specific localization to inner ear cells. Sub-cellular localization analysis presented a distinct pattern for each lncRNA and mouse tissue expression evaluation displayed a large variability in terms of level and location. Our findings establish the expression of specific lncRNAs in different cell types of the auditory system and present a potential pathway by which the lncRNA Gas5 acts in the inner ear. Studying lncRNAs and deciphering their functions may deepen our knowledge of inner ear physiology and morphology and may reveal the basis of as yet unresolved genetic hearing loss-related pathologies. Moreover, our experimental design may be employed as a reference for studying other inner ear-related lncRNAs, as well as lncRNAs expressed in other sensory systems.

LncRNA-OIS1 regulates DPP4 activation to modulate senescence induced by RAS.

Oncogene-induced senescence (OIS), provoked in response to oncogenic activation, is considered an important tumor suppressor mechanism. Long non-coding RNAs (lncRNAs) are transcripts longer than 200 nt without a protein-coding capacity. Functional studies showed that deregulated lncRNA expression promote tumorigenesis and metastasis and that lncRNAs may exhibit tumor-suppressive and oncogenic function. Here, we first identified lncRNAs that were differentially expressed between senescent and non-senescent human fibroblast cells. Using RNA interference, we performed a loss-function screen targeting the differentially expressed lncRNAs, and identified lncRNA-OIS1 (lncRNA#32, AC008063.3 or ENSG00000233397) as a lncRNA required for OIS. Knockdown of lncRNA-OIS1 triggered bypass of senescence, higher proliferation rate, lower abundance of the cell-cycle inhibitor CDKN1A and high expression of cell-cycle-associated genes. Subcellular inspection of lncRNA-OIS1 indicated nuclear and cytosolic localization in both normal culture conditions as well as following oncogene induction. Interestingly, silencing lncRNA-OIS1 diminished the senescent-associated induction of a nearby gene (Dipeptidyl Peptidase 4, DPP4) with established role in tumor suppression. Intriguingly, similar to lncRNA-OIS1, silencing DPP4 caused senescence bypass, and ectopic expression of DPP4 in lncRNA-OIS1 knockdown cells restored the senescent phenotype. Thus, our data indicate that lncRNA-OIS1 links oncogenic induction and senescence with the activation of the tumor suppressor DPP4.