570 research outputs found
Reduced splicing efficiency induced by synonymous substitutions may generate a substrate for natural selection of new splicing isoforms: the case of CFTR exon 12
Alternative splicing has been associated with increased evolutionary changes and with recent exon creation or loss. The addition of a new exon can be explained by its inclusion in only a fraction of the transcripts leaving the original form intact and giving to the new form the possibility to evolve independently but the exon loss phenomenon is less clear. To explore the mechanism that could be involved in CFTR exon 12 lower splicing efficiency in primates, we have analyzed the effect of multiple synonymous variations. Random patterns of synonymous variations were created in CFTR exon12 and the majority of them induced exon inclusion, suggesting a suboptimal splicing efficiency of the human gene. In addition, the effect of each single synonymous substitution on splicing is strongly dependent on the exonic context and does not correlate with available in silico exon splicing prediction programs. We propose that casual synonymous substitutions may lead to a reduced splicing efficiency that can result in a variable proportion of exon loss. If this phenomenon happens in in-frame exons and to an extent tolerated by the cells it can have an important evolutionary effect since it may generate a substrate for natural selection of new splicing isoforms
Direct evidence of dust growth in L183 from MIR light scattering
Theoretical arguments suggest that dust grains should grow in the dense cold
parts of molecular clouds. Evidence of larger grains has so far been gathered
in near/mid infrared extinction and millimeter observations. Interpreting the
data is, however, aggravated by the complex interplay of density and dust
properties (as well as temperature for thermal emission). We present new
Spitzer data of L183 in bands that are sensitive and insensitive to PAHs. The
visual extinction AV map derived in a former paper was fitted by a series of 3D
Gaussian distributions. For different dust models, we calculate the scattered
MIR radiation images of structures that agree agree with the AV map and compare
them to the Spitzer data. The Spitzer data of L183 show emission in the 3.6 and
4.5 micron bands, while the 5.8 micron band shows slight absorption. The
emission layer of stochastically heated particles should coincide with the
layer of strongest scattering of optical interstellar radiation, which is seen
as an outer surface on I band images different from the emission region seen in
the Spitzer images. Moreover, PAH emission is expected to strongly increase
from 4.5 to 5.8 micron, which is not seen. Hence, we interpret this emission to
be MIR cloudshine. Scattered light modeling when assuming interstellar medium
dust grains without growth does not reproduce flux measurable by Spitzer. In
contrast, models with grains growing with density yield images with a flux and
pattern comparable to the Spitzer images in the bands 3.6, 4.5, and 8.0 micron.Comment: 13 pages, 11 figures, accepted for publication in Astronomy and
Astrophysic
Inguinocrural dermolipectomy A retrospective study
Inguinocrural dermolipectomy is a constantly increasing surgical procedure, especially for postbariatric patients with significant weight loss. The "crescent medial" and "vertical medial" thigh lifting techniques (CMTL/VMTL) are the most effective type of surgery to treat laxity and excess of skin and soft tissues in the medial region of the thighs Aim of this article is to suggest which patients may be eligible for a surgical thighplasty with an acceptable risk of postoperative complications
Missense, Nonsense, and Neutral Mutations Define Juxtaposed Regulatory Elements of Splicing in Cystic Fibrosis Transmembrane Regulator Exon 9
Exonic sequence variations may induce exon inclusion or exclusion from the mature mRNA by disrupting exonic regulatory elements and/or by affecting a nuclear reading frame scanning mechanism. We have carried out a systematic study of the effect on cystic fibrosis transmembrane regulator exon 9 splicing of natural and site-directed sequence mutations. We have observed that changes in the splicing pattern were not related to the creation of premature termination codons, a fact that indicates the lack of a significant nuclear check of the reading frame in this system. In addition, the splice pattern could not be predicted by available Ser/Arg protein matrices score analysis. An extensive site-directed mutagenesis of the 3' portion of the exon has identified two juxtaposed splicing enhancer and silencer elements. The study of double mutants at these regulatory elements showed a complex regulatory activity. For example, one natural mutation (146C) enhances exon inclusion and overrides all of the downstream silencing mutations except for a C to G transversion (155G). This unusual effect is explained by the creation of a specific binding site for the inhibitory splicing factor hnRNPH. In fact, on the double mutant 146C-155G, the silencing effect is dominant. These results indicate a strict dependence between the two juxtaposed enhancer and silencer sequences and show that many point mutations in these elements cause changes in splicing efficiency by different mechanisms
An Intronic Polypyrimidine-rich Element Downstream of the Donor Site Modulates Cystic Fibrosis Transmembrane Conductance Regulator Exon 9 Alternative Splicing *
Two intronic elements, a polymorphic TGmTn locus at the end of intron 8 and an intronic splicing silencer in intron 9, regulate aberrant splicing of human cystic fibrosis transmembrane conductance regulator (CFTR) exon 9. Previous studies (Pagani, F., Buratti, E., Stuani, C., Romano, M., Zuccato, E., Niksic, M., Giglio, L., Faraguna, D., and Baralle, F. E. (2000) J. Biol. Chem. 275, 21041â21047 and Buratti, E., Dork, T., Zuccato, E., Pagani, F., Romano, M., and Baralle, F. E. (2001) Embo J. 20, 1774â1784) have demonstrated that trans-acting factors that bind to these sequences, TDP43 and Ser/Arg-rich proteins, respectively, mediate splicing inhibition. Here, we report the identification of two polypyrimidine-binding proteins, TIA-1 and polypyrimidine tract-binding protein (PTB), as novel players in the regulation of CFTR exon 9 splicing. In hybrid minigene experiments, TIA-1 induced exon inclusion, whereas PTB induced exon skipping. TIA-1 bound specifically to a polypyrimidine-rich controlling element (PCE) located between the weak 5âČ-splice site (ss) and the intronic splicing silencer. Mutants of the PCE polypyrimidine motifs did not bind TIA-1 and, in a splicing assay, did not respond to TIA-1 splicing enhancement. PTB antagonized in vitro TIA-1 binding to the PCE, but its splicing inhibition was independent of its binding to the PCE. Recruitment of U1 small nuclear RNA to the weak 5âČ-ss by complementarity also induced exon 9 inclusion, consistent with the facilitating role of TIA-1 in weak 5âČ-ss recognition by U1 small nuclear ribonucleoprotein. Interestingly, in the presence of a high number of TG repeats and a low number of T repeats in the TGmTn locus, TIA-1 activated a cryptic exonic 3âČ-ss. This effect was independent of both TIA-1 binding to the PCE and U1 small nuclear RNA recruitment to the 5âČ-ss. Moreover, it was abolished by deletion of either the TG or T sequence. These data indicate that, in CFTR exon 9, TIA-1 binding to the PCE recruits U1 small nuclear ribonucleoprotein to the weak 5âČ-ss and induces exon inclusion. The TIA-1-mediated alternative usage of the 3âČ-splice sites, which depends on the composition of the unusual TGmTn element, represents a new mechanism of splicing regulation by TIA-1
Biharmonic Conformal Field Theories
The main result of this paper is the construction of a conformally covariant
operator in two dimensions acting on scalar fields and containing fourth order
derivatives. In this way it is possible to derive a class of Lagrangians
invariant under conformal transformations. They define conformal field theories
satisfying equations of the biharmonic type. Two kinds of these biharmonic
field theories are distinguished, characterized by the possibility or not of
the scalar fields to transform non-trivially under Weyl transformations. Both
cases are relevant for string theory and two dimensional gravity. The
biharmonic conformal field theories provide higher order corrections to the
equations of motion of the metric and give a possibility of adding new terms to
the Polyakov action.Comment: 11 pages, LaTeX, no figure
The Role of Portable Incisional Negative Pressure Wound Therapy (piNPWT) in Reducing Local Complications of Post-bariatric Brachioplasty: A Case-Control Study
New lysosomal acid lipase gene mutants explain the phenotype of Wolman disease and cholesteryl ester storage disease.
Deficiency of lysosomal acid lipase (LAL) leads to either Wolman disease(WD) or the more benign cholesteryl ester storage disease (CESD). To identifythe molecular basis of the different phenotypes we have characterised the LALgene mutations in three new patients with LAL deficiency. A patient with WD washomozygote for a null allele Y303X. The other two patients, with CESD, presentedeither homozygosity for T267I or compound heterozygosity consisting of Q64R andan exon 8 donor splice site substitution (GâA in positionâ1). The mutants T267I and Q64R and the previously reported L273S, G66V,and H274Y CESD substitutions, overexpressed in stable clones, were found to befully glycosylated and show an enzymatic activity of 3â8% of that ofnormal LAL. On the other hand, the Î254â277 mutant proteinderived from exon 8 skipping and the Y303X protein were totally inactive. Bytransient transfection of hybrid minigene constructs, the CESD GâA(â1) substitution resulted in partial exon inclusion, thus allowing theproduction of a small amount of normal LAL mRNA and hence of a functionalenzyme. In contrast, a GâAsubstitution observed in WD at position +1 of the same exon 8 donor siteresulted in complete exon skipping and the sole production of an inactiveÎ254â277 protein.In conclusion,LAL genotypes determine the level of residual enzymatic activity, thusexplaining the severity of the phenotype.âPagani, F., R. Pariyarath, R.Garcia, C. Stuani, A. B. Burlina, G. Ruotolo, M. Rabusin, and F. E. Baralle. Newlysosomal acid lipase gene mutants explain the phenotype of Wolman disease andcholesteryl ester storage disease. J. Lipid Res. 1998. 39:1382â1388
Interaction of hnRNPA1/A2 and DAZAP1 with an Alu-Derived Intronic Splicing Enhancer Regulates ATM Aberrant Splicing
We have previously identified an Alu-derived Intronic Splicing enhancer (ISE) in the Ataxia Teleangectasia Mutated gene (ATM) that facilitates intron pre-mRNA processing and leads to the inclusion of a cryptic exon in the final mRNA transcript. By using an RNA pull-down assay, we show here that hnRNPA1/A2, HuR and DAZAP1 splicing factors and DHX36 RNA helicase bind to the ISE. By functional studies (overexpression and siRNA experiments), we demonstrate that hnRNPA1 and DAZAP1 are indeed involved in ISE-dependent ATM cryptic exon activation, with hnRNPA1 acting negatively and DAZAP1 positively on splicing selection. On the contrary, HuR and DHX36 have no effect on ATM splicing pattern. These data suggest that splicing factors with both negative and positive effect can assemble on the intronic Alu repeats and regulate pre-mRNA splicing
Splicing Factors Induce Cystic Fibrosis Transmembrane Regulator Exon 9 Skipping through a Nonevolutionary Conserved Intronic Element
In monosymptomatic forms of cystic fibrosis such as congenital bilateral absence of vas deferens, variations in the TG(m) and T(n) polymorphic repeats at the 3' end of intron 8 of the cystic fibrosis transmembrane regulator (CFTR) gene are associated with the alternative splicing of exon 9, which results in a nonfunctional CFTR protein. Using a minigene model system, we have previously shown a direct relationship between the TG(m)T(n) polymorphism and exon 9 splicing. We have now evaluated the role of splicing factors in the regulation of the alternative splicing of this exon. Serine-arginine-rich proteins and the heterogeneous nuclear ribonucleoprotein A1 induced exon skipping in the human gene but not in its mouse counterpart. The effect of these proteins on exon 9 exclusion was strictly dependent on the composition of the TG(m) and T(n) polymorphic repeats. The comparative and functional analysis of the human and mouse CFTR genes showed that a region of about 150 nucleotides, present only in the human intron 9, mediates the exon 9 splicing inhibition in association with exonic regulatory elements. This region, defined as the CFTR exon 9 intronic splicing silencer, is a target for serine-arginine-rich protein interactions. Thus, the nonevolutionary conserved CFTR exon 9 alternative splicing is modulated by the TG(m) and T(n) polymorphism at the 3' splice region, enhancer and silencer exonic elements, and the intronic splicing silencer in the proximal 5' intronic region. Tissue levels and individual variability of splicing factors would determine the penetrance of the TG(m)T(n) locus in monosymptomatic forms of cystic fibrosis
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