A cryptic promoter in the first exon of the SPG4 gene directs the synthesis of the 60-kDa spastin isoform

<p>Abstract</p> <p>Background</p> <p>Mutations in <it>SPG4 </it>cause the most common form of autosomal dominant hereditary spastic paraplegia, a neurodegenerative disease characterized by weakness and spasticity of the lower limbs due to degeneration of the corticospinal tract. <it>SPG4 </it>encodes spastin, a microtubule-severing ATPase belonging to the AAA family. Two isoforms of spastin, 68 and 60 kDa, respectively, are variably abundant in tissues, show different subcellular localizations and interact with distinct molecules. The isoforms arise through alternative initiation of translation from two AUG codons in exon 1; however, it is unclear how regulation of their expression may be achieved.</p> <p>Results</p> <p>We present data that rule out the hypothesis that a cap-independent mechanism may be involved in the translation of the 60-kDa spastin isoform. Instead, we provide evidence for a complex transcriptional regulation of <it>SPG4 </it>that involves both a TATA-less ubiquitous promoter and a cryptic promoter in exon 1. The cryptic promoter covers the 5'-UTR and overlaps with the coding region of the gene. By using promoter-less constructs in various experimental settings, we found that the cryptic promoter is active in HeLa, HEK293 and motoneuronal NSC34 cells but not in SH-SY-5Y neuroblastoma cells. We showed that the cryptic promoter directs the synthesis of a <it>SPG4 </it>transcript that contains a shorter 5'-UTR and translates the 60-kDa spastin isoform selectively. Two polymorphisms (S44L and P45Q), leading to an early onset severe form of hereditary spastic paraplegia when present in heterozygosity with a mutant allele, fall a few nucleotides downstream of the novel transcriptional start site, opening up the possibility that they may exert their modifier effect at the transcriptional level. We provide evidence that at least one of them decreases the activity of the cryptic promoter in luciferase assays.</p> <p>Conclusion</p> <p>We identified a cryptic promoter in exon 1 of the <it>SPG4 </it>gene that selectively drives the expression of the 60-kDa spastin isoform in a tissue-regulated manner. These data may have implications for the understanding of the biology of spastin and the pathogenic basis of hereditary spastic paraplegia.</p

Microcephaly and macrocephaly. A study on anthropometric and clinical data from 308 subjects

Head circumference is the auxological parameter that most correlates with developmental anomalies in childhood. Head circumference (HC) two standard deviations (SD) below or above the mean defines microcephaly and macrocephaly, respectively. The aim of this retrospective study was to explore anthropometric parameters and clinical characteristics among subjects with abnormalities in HC who had been referred for developmental assessment. One hundred and sixty four subjects with microcephaly and 144 subjects with macrocephaly were enrolled from birth to 18 months of age. Head circumference at birth and the association with variables related to maternal health status, gestational age, growth pattern, brain imaging and clinical characteristics were analyzed. In some cases, an etiological diagnosis was made. In the two considered conditions, we found different anthropometric and clinical associations, some of which were statistically significant, with implications for ongoing neurodevelopmental surveillance

Rejection of randomly coinciding events in ZnMoO4_4 scintillating bolometers

Random coincidence of events (particularly from two neutrino double beta decay) could be one of the main sources of background in the search for neutrinoless double beta decay with cryogenic bolometers due to their poor time resolution. Pulse-shape discrimination by using front edge analysis, mean-time and $\chi^2$ methods was applied to discriminate randomly coinciding events in ZnMoO$_4$ cryogenic scintillating bolometers. These events can be effectively rejected at the level of 99% by the analysis of the heat signals with rise-time of about 14 ms and signal-to-noise ratio of 900, and at the level of 92% by the analysis of the light signals with rise-time of about 3 ms and signal-to-noise ratio of 30, under the requirement to detect 95% of single events. These rejection efficiencies are compatible with extremely low background levels in the region of interest of neutrinoless double beta decay of $^{100}$Mo for enriched ZnMoO$_4$ detectors, of the order of $10^{-4}$ counts/(y keV kg). Pulse-shape parameters have been chosen on the basis of the performance of a real massive ZnMoO$_4$ scintillating bolometer. Importance of the signal-to-noise ratio, correct finding of the signal start and choice of an appropriate sampling frequency are discussed

Understanding the purchasing behavior of consumers in response to sustainable marketing practices: An empirical analysis in the food domain

Sustainability has become an important driver in defining business strategies, affecting most critical corporate functions and changing the way in which value is created, communicated, and distributed. This is increasingly impacting marketing practices, in particular, through promoting the development of sustainable marketing in the food sector. In line with this, our study aimed to investigate if and how sustainable marketing practices affect consumer loyalty to a specific brand. To answer our research questions, we relied on the results of a survey submitted to a sample of 907 Italian consumers of biscuits. Results showed that the consumers’ attention to sustainable issues (in the absence of adequate information that can guide them in choosing a brand) did not result in brand loyalty. The same outcome was found when consumers were overloaded by marketing campaigns, which had the effect of confusing users and making them unfaithful. Ultimately, when consumers showed both engagement with sustainable concerns and sensitivity to marketing initiatives (i.e., they are sensitive to sustainable marketing practices), a positive effect on brand loyalty was observed. Our results contribute to the emerging stream of literature discussing the relevance and potential impact of sustainable marketing

Turning to God to forgive: More than meets the eye.

Adults (N = 89; 59% female) recruited from divorce records reported levels of turning to God to forgive (TGF) themselves, their ex-spouse, and God for their divorce when it occurred (Time 1, Tl) and one year later (Time 2, T2). Seventy-five percent of participants reported TGF. T1 TGF predicted higher levels of T2 positive spiritual emotions, T2 verbal aggression by the participant and ex-spouse, and T2 demonization of ex-spouse. Participants were grouped according to pattern of TGF over time (Resolved, Chronic, Delayed, and Low). Repeated measures ANOVAs showed that the Resolved group reported greater declines in demonizing the divorce relative to other groups. Multiple main effects for TGF pattern also emerged. This highlights the need to consider the potentially desirable and undesirable psychosocial and spiritual factors associated with TGF

5-(carbamoylmethylene)-oxazolidin-2-ones as a promising class of heterocycles inducing apoptosis triggered by increased ROS levels and mitochondrial dysfunction in breast and cervical cancer

Oxazolidinones are antibiotics that inhibit protein synthesis by binding the 50S ribosomal subunit. Recently, numerous worldwide researches focused on their properties and possible involvement in cancer therapy have been conducted. Here, we evaluated in vitro the antiproliferative activity of some 5-(carbamoylmethylene)-oxazolidin-2-ones on MCF-7 and HeLa cells. The tested compounds displayed a wide range of cytotoxicity on these cancer cell lines, measured by MTT assay, exhibiting no cytotoxicity on non-tumorigenic MCF-10A cells. Among the nine tested derivatives, four displayed a good anticancer potential. Remarkably, OI compound showed IC50 values of 17.66 and 31.10 µM for MCF-7 and HeLa cancer cells, respectively. Furthermore, we assessed OI effect on the cell cycle by FACS analysis, highlighting a G1 phase arrest after 72 h, supported by a low expression level of Cyclin D1 protein. Moreover, mitochondrial membrane potential was reduced after OI treatment driven by high levels of ROS. These findings demonstrate that OI treatment can inhibit MCF-7 and HeLa cell proliferation and induce apoptosis by caspase-9 activation and cytochrome c release in the cytosol. Hence, 5-(carbamoylmethylene)-oxazolidin-2-ones have a promising anticancer activity, in particular, OI derivative could represent a good candidate for in vivo further studies and potential clinical use

Cellule somatiche in latte ovino: analisi e applicazione delle curve ROC

Negli ovini da latte le mastiti costituiscono una delle principali cause di perdite economiche per la ridotta produzione e soprattutto per la scarsa qualità tecnologica del latte. Diversi studi hanno dimostrato che il contenuto in cellule somatiche (SCC) del latte può essere impiegato sia come metodo diagnostico sia come criterio di selezione nei programmi di miglioramento genetico per la resistenza alle mastiti. Obiettivo di questo lavoro è stato applicare la metodologia delle Receiver Operating Characteristics curves (ROC) per valutare la capacità diagnostica del SCC nel discriminare le mammelle infette da quelle sane e per individuare un valore soglia discriminante in ovini da latte. In totale sono stati analizzati 14.072 campioni di latte individuale da 720 pecore di razza Valle del Belice in 5 allevamenti. L’isolamento e l’identificazione batterica sono stati effettuati tramite semina di 10 μl di latte su Agar Sangue incubato a 37°C per 24 - 48h in aerofilia e, se necessario, in microaerofilia e la determinazione delle SCC mediante citometria di flusso. In base al risultato dell’esame batteriologico sono stati identificati 9 gruppi. La media aritmetica del SCS per i test negativi e positivi è risultata rispettivamente di 5.17 ± 0.008 e 5.80 ± 0.010, la media geometrica del SCC per i test negativi e positivi è risultata rispettivamente di 146.96 x 103 e 630.85 x 103 cells/ml. Complessivamente sono risultati positivi all’esame batteriologico 6207 campioni con una prevalenza pari al 44,1%. L’AUC è risultata pari a 0.73 ± 0.004 (P 0.0001). Il POO è risultato essere >427x103 cells/ml, in corrispondenza di tale punto la sensibilità è pari al 59,2% mentre la specificità si attesta all’80,9%. In corrispondenza di tale valore il 71% dei soggetti positivi ed il 72% dei soggetti negativi sono classificati correttamente. L’approccio di tipo non parametrico per la stima delle ROC ha dimostrato che il SCC può essere validamente utilizzato per discriminare gli animali infetti da quelli sani. Tuttavia ulteriori studi si ritengono necessari per una più precisa valutazione economica dei costi delle mastiti in considerazione del fatto che il POO sulle ROC tende a spostarsi verso valori con più alta Se e più bassa Sp man mano che i costi dei falsi positivi si riducono rispetto a quelli dei falsi negativi o quando aumenta il valore della prevalenza

Demonization of Divorce: Prevalence Rates and Links to Postdivorce Adjustment

The meaning-making process can be crucial to individuals as they adjust to their divorce. Demonization is a negative coping response (also known as spiritual struggle) that involves appraising someone or something as related to demonic forces. Individuals may cognitively frame a divorce as the work of Satan in order to understand suffering while maintaining beliefs in a just world or benevolent God. In this study, nearly half (48%) of the community sample (N = 100) endorsed some form of demonization related to their recent divorce. Differences were observed in psychological postdivorce adjustment (post-traumatic stress, depression, anger, and positive/negative spiritual emotions) among groups with differing levels of demonization of divorce, demonization of ex-spouse, and demonization of self (none, low, and high). Implications for practitioners and researchers are discussed

Spiritual stress and coping model of divorce: A longitudinal study.

This study represents the first longitudinal effort to use a spiritual stress and coping model to predict adults\u27 psychosocial adjustment following divorce. A community sample of 89 participants completed measures at the time of their divorce and 1 year later. Though the sample endorsed slightly lower levels of religiosity than the general U.S. population, most reported spiritual appraisals and positive and negative religious coping tied to divorce. Hierarchical regression analyses controlling general religiousness and nonreligious forms of coping indicated that (a) appraising divorce as a sacred loss or desecration at the time it occurred predicted more depressive symptoms and dysfunctional conflict tactics with the ex-spouse 1 year later; (b) positive religious coping reported about the year following divorce predicted greater posttraumatic growth 1 year after divorce; and (c) negative religious coping reported about the year following divorce predicted more depressive symptoms 1 year after the divorce. Bootstrapping mediation analyses indicated that negative religious coping fully mediated links between appraising the divorce as a sacred loss or desecration at the time it occurred and depressive symptoms 1 year later. In addition, moderation analyses revealed that negative religious coping is more strongly associated with depressive symptoms among those who form high versus low appraisals of their divorce as a sacred loss or desecration. These findings are relevant to divorce education and intervention provided by professionals in legal, family, mental health, and clerical roles. Implications are discussed for clinical and counseling psychology and religious communities

Cellule somatiche ed isolamento di agenti mastidogeni in ovini di razza Valle del Belice

Il conteggio delle cellule somatiche nel latte ovino si è dimostrato un valido strumento per la determinazione dello stato sanitario della mammella. Oggetto del presente studio è stato valutare eventuali relazioni fra il contenuto in cellule somatiche (SCC) e le infezioni mammarie (IMI) negli ovini. L’indagine è stata condotta nel periodo 2006-2012 su n.14072 campioni di latte individuale. In base al tenore i campioni sono state inseriti in 4 classi: 300.000; da 301.000 a 500.000; da 501.000 a 1.000.000 e ≥1.001.000 cellule/ml. Per l’isolamento colturale sono stati seminati 10 μl di latte su piastre di Agar Sangue incubate per 24-48 h a 37 °C in aerofilia e, se necessario, in microaerofilia. Sono stati distinti 10 gruppi: nessun isolamento, isolamento di stafilococchi coagulasi positivi (S. aureus, S. intermedius e S. hyicus), stafilococchi coagulasi negativi (CNS), streptococchi patogeni (S. agalactiae, S. dysgalactiae e S. uberis), altri streptococchi o enterococchi (S. acidominimus,S. canis, S. viridans, S. mitis, S. mitior ed E. faecalis), Corynebacterium, Mannheimia haemolithyca, Bacillus spp, altri germi (Proteus spp., E.coli e Pseudomonas spp.) ed associazione di CNS ed altri streptococchi. Per ogni gruppo è stata calcolata la media geometrica del SCC. Il 44% dei campioni analizzati mostrava positività all’esame microbiologico con percentuali crescenti in funzione della classe di SCC d’appartenenza. Si riportano i microrganismi isolati e le relative prevalenze: CNS 80%, stafilococchi coagulasi positivi 11,76% di cui 677 S. aureus (10,9%), 43 S. intermedius (0,69%) e 10 S. hyicus (0,16%), Streptococcus spp. 3,09%, streptococchi patogeni 2,64%, M. haemolytica 0.63%, Corynebacterium 0,18%, Pseudomonas 0,10%, Bacillus 0,68%, E. coli 0,06% e Proteus 0,34%. La media geometrica del SCC mostra differenze statisticamente significative fra i vari gruppi di batteri. Nell’allevamento ovino gli CNS si confermano i principali agenti di mastite subclinica con tendenza alla cronicizzazione pertanto ulteriori approfondimenti sono necessari per indagare il loro ruolo patogenetic