The regulatory roles of small nucleolar RNAs within their host locus.

Small nucleolar RNAs (snoRNAs) are a class of conserved noncoding RNAs forming complexes with proteins to catalyse site-specific modifications on ribosomal RNA. Besides this canonical role, several snoRNAs are now known to regulate diverse levels of gene expression. While these functions are carried out in by mature snoRNAs, evidence has also been emerging of regulatory roles of snoRNAs in , either within their genomic locus or as longer transcription intermediates during their maturation.

Proteomic analyses reveal new features of the box H/ACA RNP biogenesis.

The conserved H/ACA RNPs consist of one H/ACA RNA and 4 core proteins: dyskerin, NHP2, NOP10, and GAR1. Its assembly requires several assembly factors. A pre-particle containing the nascent RNAs, dyskerin, NOP10, NHP2 and NAF1 is assembled co-transcriptionally.

Emerging role of IκBζ in inflammation: Emphasis on psoriasis.

Psoriasis is a chronic inflammatory disorder affecting skin and joints that results from immunological dysfunction such as enhanced IL-23 induced Th-17 differentiation. IkappaB-Zeta (IκBζ) is an atypical transcriptional factor of the IκB protein family since, contrary to the other family members, it positively regulates NF-κB pathway by being exclusively localized into the nucleus. IκBζ deficiency reduces visible manifestations of experimental psoriasis by diminishing expression of psoriasis-associated genes.

Phylogenetic and Molecular Analyses Identify SNORD116 Targets Involved in the Prader-Willi Syndrome.

The eutherian-specific SNORD116 family of repeated box C/D snoRNA genes is suspected to play a major role in the Prader-Willi syndrome (PWS), yet its molecular function remains poorly understood. Here, we combined phylogenetic and molecular analyses to identify candidate RNA targets. Based on the analysis of several eutherian orthologs, we found evidence of extensive birth-and-death and conversion events during SNORD116 gene history.

Emerging Data on the Diversity of Molecular Mechanisms Involving C/D snoRNAs.

Box C/D small nucleolar RNAs (C/D snoRNAs) represent an ancient family of small non-coding RNAs that are classically viewed as housekeeping guides for the 2'-O-methylation of ribosomal RNA in Archaea and Eukaryotes. However, an extensive set of studies now argues that they are involved in mechanisms that go well beyond this function. Here, we present these pieces of evidence in light of the current comprehension of the molecular mechanisms that control C/D snoRNA expression and function.

Using Native RIP, UV-CLIP or fCLIP to Address Protein-RNA Interactions In Vivo.

Stable and transient interactions between molecules are determinant for cell function. Among those, numerous proteins contact coding and noncoding RNAs to modulate their fate and promote their activity. The identification of such interactions as well as the cellular and molecular conditions of these interactions represent key information for the characterization of the role of each partner.

The box C/D snoRNP assembly factor Bcd1 interacts with the histone chaperone Rtt106 and controls its transcription dependent activity.

Biogenesis of eukaryotic box C/D small nucleolar ribonucleoproteins initiates co-transcriptionally and requires the action of the assembly machinery including the Hsp90/R2TP complex, the Rsa1p:Hit1p heterodimer and the Bcd1 protein. We present genetic interactions between the Rsa1p-encoding gene and genes involved in chromatin organization including RTT106 that codes for the H3-H4 histone chaperone Rtt106p controlling H3K56ac deposition. We show that Bcd1p binds Rtt106p and controls its transcription-dependent recruitment by reducing its association with RNA polymerase II, modulating H3K56ac levels at gene body.

Bcd1p controls RNA loading of the core protein Nop58 during C/D box snoRNP biogenesis.

Biogenesis of eukaryotic box C/D small nucleolar ribonucleoproteins (C/D snoRNPs) is guided by conserved -acting factors that act collectively to assemble the core proteins SNU13/Snu13, NOP58/Nop58, NOP56/Nop56, FBL/Nop1, and box C/D small nucleolar RNAs (C/D snoRNAs), in human and in yeast, respectively. This finely elaborated process involves the sequential interplay of snoRNP-related proteins and RNA through the formation of transient pre-RNP complexes. BCD1/Bcd1 protein is essential for yeast cell growth and for the specific accumulation of box C/D snoRNAs.

Deletion of the miR-379/miR-410 gene cluster at the imprinted Dlk1-Dio3 locus enhances anxiety-related behaviour.

The brain-specific miR-379/miR-410 gene cluster at the imprinted Dlk1-Dio3 domain is implicated in several aspects of brain development and function, particularly in fine-tuning the dendritic outgrowth and spine remodelling of hippocampal neurons. Whether it might influence behaviour and memory-related processes has not yet been explored at the whole organism level. We previously reported that constitutive deletion of the miR-379/miR-410 gene cluster affects metabolic adaptation in neonatal mice.

The miR-379/miR-410 cluster at the imprinted Dlk1-Dio3 domain controls neonatal metabolic adaptation.

In mammals, birth entails complex metabolic adjustments essential for neonatal survival. Using a mouse knockout model, we identify crucial biological roles for the miR-379/miR-410 cluster within the imprinted Dlk1-Dio3 region during this metabolic transition. The miR-379/miR-410 locus, also named C14MC in humans, is the largest known placental mammal-specific miRNA cluster, whose 39 miRNA genes are expressed only from the maternal allele.

The Yin and Yang of pain: variability in formalin test nociception and morphine analgesia produced by the Yin Yang 1 transcription factor gene.

We recently observed a reliable phenotypic difference in the inflammatory pain sensitivity of a congenic mouse strain compared to its background strain. By constructing and testing subcongenic strains combined with gene-expression assays, we provide evidence for the candidacy of the Yy1 gene - encoding the ubiquitously expressed and multifunctional Yin Yang 1 transcription factor - as responsible. To confirm this hypothesis, we used a Cre/lox strategy to produce mutant mice in which Yy1 expression was ablated in Nav 1.

Do repeated arrays of regulatory small-RNA genes elicit genomic imprinting?: Concurrent emergence of large clusters of small non-coding RNAs and genomic imprinting at four evolutionarily distinct eutherian chromosomal loci.

The basic premise of the host-defense theory is that genomic imprinting, the parent-of-origin expression of a subset of mammalian genes, derives from mechanisms originally dedicated to silencing repeated and retroviral-like sequences that deeply colonized mammalian genomes. We propose that large clusters of tandemly-repeated C/D-box small nucleolar RNAs (snoRNAs) or microRNAs represent a novel category of sequences recognized as "genomic parasites", contributing to the emergence of genomic imprinting in a subset of chromosomal regions that contain them. Such a view is supported by evidence derived from studies of the imprinted snoRNA- and/or miRNA-encoding Dlk1-Dio3, Snurf-Snrpn, Sfbmt2, and C19MC domains.

Novel imprinted transcripts from the Dlk1-Gtl2 intergenic region, Mico1 and Mico1os, show circadian oscillations.

Most of the known imprinted genes are assembled into clusters that share common imprinting control regions (ICRs). Non-coding transcripts are often associated with ICRs and implicated in imprinting regulation. We undertook a systematic search for transcripts originating from the Dlk1-Gtl2 intergenic region that contains the ICR for the chromosome 12 imprinted cluster and identified two overlapping transcripts expressed from opposite strands exclusively from the maternal chromosome.

Parental effect of DNA (Cytosine-5) methyltransferase 1 on grandparental-origin-dependent transmission ratio distortion in mouse crosses and human families.

Transmission ratio distortion (TRD) is a deviation from the expected Mendelian 1:1 ratio of alleles transmitted from parents to offspring and may arise by different mechanisms. Earlier we described a grandparental-origin-dependent sex-of-offspring-specific TRD of maternal chromosome 12 alleles closely linked to an imprinted region and hypothesized that it resulted from imprint resetting errors in the maternal germline. Here, we report that the genotype of the parents for loss-of-function mutations in the Dnmt1 gene influences the transmission of grandparental chromosome 12 alleles.

Coordinated diurnal regulation of genes from the Dlk1-Dio3 imprinted domain: implications for regulation of clusters of non-paralogous genes.

The functioning of the genome is tightly related to its architecture. Therefore, understanding the relationship between different regulatory mechanisms and the organization of chromosomal domains is essential for understanding genome regulation. The majority of imprinted genes are assembled into clusters, share common regulatory elements, and, hence, represent an attractive model for studies of regulation of clusters of non-paralogous genes.

Gene therapy of the typical multidrug resistance phenotype of cancers: a new hope?

The multidrug resistance (MDR) phenotype of cancers has generated a large amount of research, owing to its constant fatal clinical outcome. Many studies have focused on the discovery of chemomodulators; however, in spite of this huge effort, the side effects that these products induce, and their additive toxicity when used in the presence of anticancer drugs, have led to the disaffection of the pharmaceutical industry and possibly slowed down research in pharmacological modulation. New tools developed using molecular biology techniques have opened the way for gene therapy and given birth to new therapeutic hopes.

[Multidrug resistance in uveal melanoma].

In spite of important progress in the local treatment of uveal melanoma, the most frequent primitive intraocular tumor, 15%-30% of patients still die because of tumor metastasis. This tumor is characterized by constitutive chemoresistance, thwarting any attempt to control it using the usual chemotherapy protocols. The chemoresistance of uveal melanoma is mainly due to the typical multidrug resistance phenotype (MDR), which is linked to overexpression of membrane proteins that actively extrude anticancer drugs from the cell.

Characterization of the typical multidrug resistance profile in human uveal melanoma cell lines and in mouse liver metastasis derivatives.

Uveal melanoma is the most common intraocular malignancy. To study its biology, stable cell lines provide a useful tool, but these are very difficult to obtain. A stable and rapidly growing human choroidal melanoma cell line composed of pure epithelioid cells was established and maintained for at least 4 years.

Major cytogenetic aberrations and typical multidrug resistance phenotype of uveal melanoma: current views and new therapeutic prospects.

Uveal melanoma is the most frequent intra-ocular cancer. The recent development of new chromosome-related technologies have permitted the elucidation of both the cytogenetics and the natural history of this disease. Fifty to 60% of uveal melanomas are linked to a monosomy 3, which appears as an early and determinant event in tumor progression.

Control of P-glycoprotein activity by membrane cholesterol amounts and their relation to multidrug resistance in human CEM leukemia cells.

P-glycoprotein (P-gp) is the most well-known ATP-binding cassette (ABC) transporter involved in unidirectional substrate translocation across the membrane lipid bilayer, thereby causing the typical multidrug resistance (MDR) phenotype expressed in many cancers. We observed that in human CEM acute lymphoblastic leukemia cells expressing various degrees of chemoresistance and where P-gp was the sole MDR-related ABC transporter detected, the amount of esterified cholesterol increased linearly with the level of resistance to vinblastine while the amounts of total and free cholesterol increased in a nonlinear way. Membrane cholesterol controlled the ATPase activity of P-gp in a linear manner, whereas the P-gp-induced daunomycin efflux decreased nonlinearly with the depletion of membrane cholesterol.

Multidrug-resistant cancer cells contain two populations of P-glycoprotein with differently stimulated P-gp ATPase activities: evidence from atomic force microscopy and biochemical analysis.

Considerable interest exists about the localization of P-gp (P-glycoprotein) in DRMs (detergent-resistant membranes) of multidrug resistant cancer cells, in particular concerning the potential modulating role of the closely related lipids and proteins on P-gp activity. Our observation of the opposite effect of verapamil on P-gp ATPase activity from DRM and solubilized-membrane fractions of CEM-resistant leukaemia cells, and results from Langmuir experiments on membrane monolayers from resistant CEM cells, strongly suggest that two functional populations of P-gp exist. The first is located in DRM regions: it displays its optimal P-gp ATPase activity, which is almost completely inhibited by orthovanadate and activated by verapamil.

[Toward monosomy 3 as the main prognosis factor of uveal melanoma: current cytogenetic data].

Uveal melanoma is the most frequent intraocular cancer. The recent development of new technologies such as microsatellite analysis and comparative genomic hybridization have elucidated both the cytogenetics and the natural history of this disease. Fifty to 60% of uveal melanomas are linked to monosomy 3, which appears as an early and determinant event in tumor progression.

New invMED1 element cis-activates human multidrug-related MDR1 and MVP genes, involving the LRP130 protein.

The MDR1 gene is a key component of the cytotoxic defense network and its overexpression results in the multidrug resistance (MDR) phenotype. However, the molecular mechanisms that regulate the MDR1 gene and coordinate multiple MDR-related genes expression are poorly understood. In a previous study, we identified a new 12 bp cis-activating region in the 5'-flanking region of the human MDR1 gene, which we called inverted MED1.