Biomedical subjects

Uberto Pozzoli

Publications and source records attributed to Uberto Pozzoli.

17 recordsLinked to original sources

PubMed2006

Intron size in mammals: complexity comes to terms with economy.

Different and contrasting models have been proposed to explain intron size evolution in mammals. Here, we demonstrate that intron and intergenic size per se has no adaptive role in gene expression regulation but reflects the need to preserve conserved intronic elements. Although the amount of non-coding functional elements explains the within-genome size variation of intergenic spacers, we show that an additional, additive pressure has been acting on highly expressed introns to reduce the cost of their transcription.

Animals↗

PubMed2006

A region in the dystrophin gene major hot spot harbors a cluster of deletion breakpoints and generates double-strand breaks in yeast.

Deletions within the dystrophin gene (DMD) account for >70% of mutations leading to Duchenne and Becker muscular dystrophies (DMD and BMD). Deletion breakpoints were reported to be scattered within regions that also represent meiotic recombination hot spots. Recent studies indicates that deletion junctions arise from nonhomologous end joining (NHEJ), a major pathway for repairing DNA double-strand breaks (DSBs) in mammals. Here we show that a region in intron 47 (i.e., a major deletion hot spot in the DMD gene) generates DSBs during meiosis in yeast and harbors a cluster of previously sequenced deletion breaks. Mapping of breakpoints in 26 BMD/DMD patients indicated that the frequency of breakpoint occurrence around this region is 3-fold higher than expected by chance. These findings suggest that DSBs mediate deletion formation in intron 47 and possibly account for the high frequency of meiotic recombination in the region. Statistical analysis indicated the presence of at least one other breakpoint cluster in intron 47. Taken together, these results suggest that the primary events in deletion formation occur within discrete regions and that the scattered breakpoint distribution reflects both a variable degree of DSB end processing and the availability of a small (compared to the huge regions involved) deletion junction sample.

Base Sequence↗

PubMed2006

Gene function and expression level influence the insertion/fixation dynamics of distinct transposon families in mammalian introns.

BACKGROUND: Transposable elements (TEs) represent more than 45% of the human and mouse genomes. Both parasitic and mutualistic features have been shown to apply to the host-TE relationship but a comprehensive scenario of the forces driving TE fixation within mammalian genes is still missing. RESULTS: We show that intronic multispecies conserved sequences (MCSs) have been affecting TE integration frequency over time. We verify that a selective economizing pressure has been acting on TEs to decrease their frequency in highly expressed genes. After correcting for GC content, MCS density and intron size, we identified TE-enriched and TE-depleted gene categories. In addition to developmental regulators and transcription factors, TE-depleted regions encompass loci that might require subtle regulation of transcript levels or precise activation timing, such as growth factors, cytokines, hormones, and genes involved in the immune response. The latter, despite having reduced frequencies of most TE types, are significantly enriched in mammalian-wide interspersed repeats (MIRs). Analysis of orthologous genes indicated that MIR over-representation also occurs in dog and opossum immune response genes, suggesting, given the partially independent origin of MIR sequences in eutheria and metatheria, the evolutionary conservation of a specific function for MIRs located in these loci. Consistently, the core MIR sequence is over-represented in defense response genes compared to the background intronic frequency. CONCLUSION: Our data indicate that gene function, expression level, and sequence conservation influence TE insertion/fixation in mammalian introns. Moreover, we provide the first report showing that a specific TE family is evolutionarily associated with a gene function category.

Animals↗

PubMed2005

Analysis of intronic conserved elements indicates that functional complexity might represent a major source of negative selection on non-coding sequences.

The non-coding portion of human genome is punctuated by a large number of multispecies conserved sequence (MCS) elements with largely unknown function. We demonstrate that MCSs are unevenly distributed in human introns with the majority of relatively short introns (< 9 kb long) displaying no or a few MCSs and that MCS density reaching up to 10% of total size in longer introns. After correction for intron length, MCSs were found to be enriched within genes involved in development and transcription, whereas depleted in immune response loci. Moreover, many central nervous system tissues show a preferential expression of MCS-rich genes and MCS enrichment significantly correlates with gene functional complexity in terms of distinct protein domains. Analysis of human-mouse orthologous pairs indicated a significant association between intronic MCS density and conservation of protein sequence, promoter regions and untranslated sequences. Moreover, MCS density correlates with the predicted occurrence of human-mouse conserved alternative splicing events. These observations suggest that evolution acts on human genes as integrated units of coding and regulatory capacity and that functional complexity might represent a major source of negative selection on non-coding sequences. To substantiate our result, we also searched previously experimentally identified intronic regulatory elements and indicate that about half of these sequences map to an MCS; in particular, support to the notion whereby mutations in MCSs can result in human genetic diseases is provided, because three previously identified intronic pathological variations were found to occur within MCSs, and human disease and cancer genes were found significantly enriched in MCSs.

Animals↗

PubMed2005

Skeletal muscle gene expression profiling in mitochondrial disorders.

Extremely variable clinic and genetic features characterize mitochondrial encephalomyopathy (MEM). Pathogenic mitochondrial DNA (mtDNA) defects can be divided into large-scale rearrangements and single point mutations. Clinical manifestations become evident when a threshold percentage of the total mtDNA is mutated. In some MEM, the "mutant load" in an affected tissue is directly related to the severity of the phenotype. However, the clinical phenotype is not simply a direct consequence of the relative abundance of mutated mtDNA. Other factors, such as nuclear background, can contribute to the disease process, resulting in a wide range of phenotypes caused by the same mutation. Using Affymetrix oligonucleotide cDNA microarrays (HG-U133A), we studied the gene expression profile of muscle tissue biopsies obtained from 12 MEM patients [4 common 4977 bp deleted mtDNA and 8 A3243G: 4 progressive external ophthalmoplegia (PEO) and 4 mitochondrial myopathy, encephalopathy, lactic cidosis, and stroke-like episodes syndrome (MELAS) phenotypes] compared with age-matched healthy individuals. We found several differentially expressed genes: 35 were markedly up-regulated in the mtDNA macro-deletion group (vs. the control group) and 4 decreased; 56 genes were dysregulated in A3243G-related disorders (53 down-regulated in PEO and 3 up-regulated in MELAS). Finally, 12 genes were similarly regulated in the majority of the MEM patients under study. Amongst these, we identified an increased expression of genes related to the metabolism of the amino groups, as well as of several genes involved in genetic information processing. Moreover, few genes were similarly decreased in MEM patients vs. the control group. Real-time PCR demonstrated excellent reproducibility of the microarray-based findings. The observed expression changes are likely to represent a molecular signature for mitochondrial disorders. Furthermore, the differential expression profile of MELAS(A3243G) vs. PEO(A3243G) may support a role of nuclear background in contributing to these different clinical phenotypes. MEM microarray data are available from GEO database (http://www.ncbi.nlm.nih.gov/geo/) with the accession number: GSE1462.

Adolescent↗

PubMed2005

Influence of the serotonin transporter promoter gene and shyness on children's cerebral responses to facial expressions.

BACKGROUND: Childhood shyness can predate social anxiety disorder and may be associated with biased discrimination of facial expressions of emotions. OBJECTIVE: To determine whether childhood shyness, or the serotonin transporter promoter polymorphism genotype, can predict participants' visual event-related potentials in response to expressions of children of similar ages. DESIGN: Study group drawn from an inception cohort of 149 subjects characterized 1 year before the present study by their degree of shyness. SETTING: Third- and fourth-grade schoolchildren. PARTICIPANTS: Forty-nine of the inception cohort children, randomly selected. MAIN OUTCOME MEASURES: Latencies and amplitudes of the N400 waveform in response to happy, neutral, and angry expressions. RESULTS: Shyness predicted significantly smaller N400 amplitudes in response to anger (at Pz: P < or = .04) and to a neutral expression (at Pz: P < or = .047). Shyness was significantly different across the 3 genotypes, the SS genotype being associated with higher shyness levels (analysis of variance: F(2,42) = 4.47, P < or = .02; Tukey honestly significant difference, SS vs LL, P < or = .01). An analysis of covariance showed that neither the type of expression nor the genotype per se influenced the N400 amplitudes, but a significant expression X genotype interaction was found (F(4,72) = 3.57, P < or = .01), sustained by the difference in amplitude of the SS and S carrier subjects compared with the LL subjects when exposed to the anger expression (Tukey honestly significant difference, P < or = .02). CONCLUSION: Children who manifest higher levels of shyness or have 1 or 2 copies of the short allele of the serotonin transporter promoter gene appear to have a different pattern of processing affective stimuli of interpersonal hostility.

Affect↗

PubMed2005

Comprehending semantic and grammatical violations in Italian. N400 and P600 comparison with visual and auditory stimuli.

Arguments about the existence of language-specific neural systems and specifically about the independence of syntactic and semantic processing have focused on the event-related brain measures (ERPs) as tool to monitoring moment-by-moment the cognitive processes underlaid. In the present experiments, the available evidence indicates that the ERP response to semantic anomalies is at least partially distinct from the ERP response to syntactic anomalies and that two distinct processes are activated in sentences comprehension. ERPs were recorded from 10 electrodes while subjects read (Experiment 1) or listened (Experiment 2) to sentences containing semantic or syntactic violations. Final-words that were inconsistent with the sentence context elicited a negative-going wave at about 400 ms poststimulus, whereas penultimate-word incongruous with the grammatical structure (subject-verb non-agreement) elicited a positive-going wave about 600ms poststimulus. No differences based on the perceptual modality of the stimulus (visual or auditory) nor different ERP correlates as a function of task-relevance (explicit/implicit task induction) were found The implications of our results for Italian language are explained.

Adult↗

PubMed2005

Fixation of conserved sequences shapes human intron size and influences transposon-insertion dynamics.

The basis for intron expansion in humans is largely unexplored. In this article, we demonstrate that intron expansion has primarily been determined by fixation of multispecies conserved sequences (MCSs) over time. The presence of MCSs has shaped intron features: the insertion of transposable elements (TEs) has been constrained as more MCSs were fixed. Analysis of TE and MCS distribution suggested an unprecedented estimate of information requirements for proper splicing of long introns with indication of sequence constraints extending up to >3 kb downstream 5' splice sites.

Alternative Splicing↗

PubMed2005

Morphed facial expressions elicited a N400 ERP effect: a domain-specific semantic module?

Previous studies have revealed that decoding of facial expressions is a specific component of face comprehension and that semantic information might be processed separately from the basic stage of face perception. In order to explore event-related potentials (ERPs) related to recognition of facial expressions and the effect of the semantic content of the stimulus, we analyzed 20 normal subjects. Faces with three prototypical emotional expressions (fear, happiness, and sadness) and with three morphed expressions were presented in random order. The neutral stimuli represented the control condition. Whereas ERP profiles were similar with respect to an early negative ERP (N170), differences in peak amplitude were observed later between incongruous (morphed) expressions and congruous (prototypical) ones. In fact, the results demonstrated that the emotional morphed faces elicited a negative peak at about 360 ms, mainly distributed over the posterior site. The electrophysiological activity observed may represent a specific cognitive process underlying decoding of facial expressions in case of semantic anomaly detection. The evidence is in favor of the similarity of this negative deflection with the N400 ERP effect elicited in linguistic tasks. A domain-specific semantic module is proposed to explain these results.

Adult↗

PubMed2004

Over-representation of exonic splicing enhancers in human intronless genes suggests multiple functions in mRNA processing.

The human transcriptome is constituted of a great majority of intron-containing and a minority of intron-lacking mRNAs; given the different processing these transcripts undergo, they are expected to carry, intermingled with coding properties, very different editing information. Here we applied a computational approach to compare intronless and intron-containing coding sequences. Hexamer composition comparison allowed the definition of over- and under-represented motifs in intronless genes; surprisingly, experimental testing revealed that intron-lacking coding sequences are enriched rather than depleted in elements with splicing enhancement ability. Similarly, we show evidence that intronless transcripts display a significantly higher frequency of both shuttling and non-shuttling SR protein binding sites compared to intron-containing sequences. These observations suggest that SR proteins (and possibly other splicing factors) play a role in cellular processes distinct from splicing.

Base Sequence↗

PubMed2004

Silencer elements as possible inhibitors of pseudoexon splicing.

Human pre-mRNAs contain a definite number of exons and several pseudoexons which are located within intronic regions. We applied a computational approach to address the question of how pseudoexons are neglected in favor of exons and to possibly identify sequence elements preventing pseudoexon splicing. A search for possible splicing silencers was carried out on a pseudoexon selection that resembled exons in terms of splice site strength and exon splicing enhancer (ESE) representation; three motifs were retrieved through hexamer composition comparisons. One of these functions as a powerful silencer in transfection-based splicing assays and matches a previously identified silencer sequence with hnRNP H binding ability. The other two motifs are novel and failed to induce skipping of a constitutive exon, indicating that they might act as weak repressors or in synergy with other unidentified elements. All three motifs are enriched in pseudoexons compared with intronic regions and display higher frequencies in intronless gene-coding sequences compared with exons. We consider that a subpopulation of pseudoexons might rely on negative regulators for splicing repression; this hypothesis, if experimentally verified, might improve our understanding of exonic splicing regulatory sequences and provide the identification of a novel mutation target for human genetic diseases.

Animals↗

PubMed2004

N400 and P600 or the role of the ERP correlates in sentence comprehension: some applications to the Italian language.

In the present study, event-related brain potentials (ERPs) were applied to the study of language comprehension in the Italian language. The ERPs were recorded from 10 electrodes while the participants read (Experiment 1) or listened (Experiment 2) to sentences containing semantic or syntactic anomalies. Final words that were inconsistent with the sentence context elicited a negative wave at about 400 ms poststimulus that was more concentrated in the posterior sites of the scalp, whereas final words that were incongruous with the grammatical structure (subject-verb nonagreement) elicited a positive wave at about 600 ms poststimulus that was homogeneously distributed on the scalp. The authors found no differences based on the perceptual modality of the stimulus (visual or auditory), nor did they find different ERP correlates as a function of task relevance (explicit-implicit task induction). The available evidence indicated that the ERP response to semantic anomalies was at least partially distinct from the ERP response to syntactic anomalies, and that a syntactic parser is a plausible process included in sentence comprehension. The two semantic and syntactic effects appear as automatic processes of the decoding of the anomalies and also modality-independent processes. Cross-linguistic applications are considered in the general discussion.

Acoustic Stimulation↗

PubMed2003

The 129 codon polymorphism of the prion protein gene influences earlier cognitive performance in Down syndrome subjects.

Recently, a frequent prion protein gene (PRNP) polymorphism consisting of a methionine (M) for valine (V) substitution at codon 129 has been associated with cognitive impairment in elderly individuals. Down syndrome (DS) is associated with mental retardation and development of Alzheimer-like brain abnormalities. In the present study, we investigated the role of the PRNP polymorphism in 122 relatively young Italian DS patients. Allele frequencies of DS subjects did not differ from those in the general population. However, we found a significantly faster rate of decline in intellectual ability in the subgroup of DS patients carrying at least one V allele compared with the M/M DS subjects. An additive deleterious effect of apolipoprotein E epsilon 4 allele was detected after stratifying by APOE gene status. Our findings provide evidence that variability of the PRNP gene at codon 129 might contribute to accelerating the rate of earlier cognitive decline in DS subjects.

Adolescent↗

PubMed2003

Face-selective processing and the effect of pleasant and unpleasant emotional expressions on ERP correlates.

Previous studies have revealed that decoding of facial-expressions starts very early in the brain ( approximately 180 ms post-stimulus) and might be processed separately from the basic stage of face perception. In order to explore brain potentials (ERPs) related to decoding of facial-expressions and the effect of emotional valence of the stimulus, we analyzed 18 normal subjects. Faces with five basic emotional expressions (fear, anger, surprise, happiness, sadness) and neutral stimulus were presented in random order. The results demonstrated that an emotional face elicited a negative peak at approximately 230 ms (N230), distributed mainly over the posterior site for each emotion. The electrophysiological activity observed may represent specific cognitive processing underlying the decoding of emotional facial-expressions. Nevertheless, differences in peak amplitude were observed for high-arousal negative expressions compared with positive (happiness) and low-arousal expressions (sadness). N230 amplitude increased in response to anger, fear and surprise, suggesting that subjects' ERP variations are affected by experienced emotional intensity, related to arousal and unpleasant value of the stimulus.

Adult↗

PubMed2003

Comparative analysis of vertebrate dystrophin loci indicate intron gigantism as a common feature.

The human DMD gene is the largest known to date, spanning > 2000 kb on the X chromosome. The gene size is mainly accounted for by huge intronic regions. We sequenced 190 kb of Fugu rubripes (pufferfish) genomic DNA corresponding to the complete dystrophin gene (FrDMD) and provide the first report of gene structure and sequence comparison among dystrophin genomic sequences from different vertebrate organisms. Almost all intron positions and phases are conserved between FrDMD and its mammalian counterparts, and the predicted protein product of the Fugu gene displays 55% identity and 71% similarity to human dystrophin. In analogy to the human gene, FrDMD presents several-fold longer than average intronic regions. Analysis of intron sequences of the human and murine genes revealed that they are extremely conserved in size and that a similar fraction of total intron length is represented by repetitive elements; moreover, our data indicate that intron expansion through repeat accumulation in the two orthologs is the result of independent insertional events. The hypothesis that intron length might be functionally relevant to the DMD gene regulation is proposed and substantiated by the finding that dystrophin intron gigantism is common to the three vertebrate genes.

Amino Acid Sequence↗

PubMed2002

Relevance of sequence and structure elements for deletion events in the dystrophin gene major hot-spot.

Large intragenic deletions within the DMD locus account for about 60% of Duchenne and Becker muscular dystrophy patients. Two deletion hot-spots have been described in the dystrophin gene, but the mechanisms that determine chromosome breaks in these regions are unknown, and the huge dimensions of the gene have hampered the description of a consistent number of breakpoint sequences. A long-distance polymerase chain reaction strategy was used to amplify 20 deletion junctions involving the major hot-spot and to describe breakpoint position at the sequence level. These junctions were analyzed together with previously reported breakpoint locations so as to increase the sample number and possibly provide a comprehensive study. Minisatellite core sequences, chi elements, translin-binding sites, Pur elements, and matrix attachment regions were sought over the whole gene. Sequence-dependent DNA curvature and duplex stability were also calculated throughout the gene, and their cumulative frequency distribution was evaluated. No association with either sequence or structure elements involved in known illegitimate recombination mechanisms was identified. This study highlights the importance of a whole gene approach to rule out the presumptive role of specific features that, when locally analyzed, might suggest involvement in gene rearrangements.

Base Sequence↗

PubMed2002

Comparative analysis of the human dystrophin and utrophin gene structures.

We present analysis of intronic sequences in the human DMD and UTRN genes. In both genes accumulation of repeated elements could account for intron expansion. Out-of-frame rod-domain exons have stronger splice sites and are separated by significantly longer introns as compared to in-frame exons. These features are unique for the two homologs and not shared by other spectrin superfamily genes.

Cytoskeletal Proteins↗