PPS, a Large Multidomain Protein, Functions with Sex-Lethal to Regulate Alternative Splicing in Drosophila

Alternative splicing controls the expression of many genes, including the Drosophila sex determination gene Sex-lethal (Sxl). Sxl expression is controlled via a negative regulatory mechanism where inclusion of the translation-terminating male exon is blocked in females. Previous studies have shown that the mechanism leading to exon skipping is autoregulatory and requires the SXL protein to antagonize exon inclusion by interacting with core spliceosomal proteins, including the U1 snRNP protein Sans-fille (SNF). In studies begun by screening for proteins that interact with SNF, we identified PPS, a previously uncharacterized protein, as a novel component of the machinery required for Sxl male exon skipping. PPS encodes a large protein with four signature motifs, PHD, BRK, TFS2M, and SPOC, typically found in proteins involved in transcription. We demonstrate that PPS has a direct role in Sxl male exon skipping by showing first that loss of function mutations have phenotypes indicative of Sxl misregulation and second that the PPS protein forms a complex with SXL and the unspliced Sxl RNA. In addition, we mapped the recruitment of PPS, SXL, and SNF along the Sxl gene using chromatin immunoprecipitation (ChIP), which revealed that, like many other splicing factors, these proteins bind their RNA targets while in close proximity to the DNA. Interestingly, while SNF and SXL are specifically recruited to their predicted binding sites, PPS has a distinct pattern of accumulation along the Sxl gene, associating with a region that includes, but is not limited to, the SxlPm promoter. Together, these data indicate that PPS is different from other splicing factors involved in male-exon skipping and suggest, for the first time, a functional link between transcription and SXL–mediated alternative splicing. Loss of zygotic PPS function, however, is lethal to both sexes, indicating that its role may be of broad significance

Human Cryptochrome-1 Confers Light Independent Biological Activity in Transgenic Drosophila Correlated with Flavin Radical Stability

Cryptochromes are conserved flavoprotein receptors found throughout the biological kingdom with diversified roles in plant development and entrainment of the circadian clock in animals. Light perception is proposed to occur through flavin radical formation that correlates with biological activity in vivo in both plants and Drosophila. By contrast, mammalian (Type II) cryptochromes regulate the circadian clock independently of light, raising the fundamental question of whether mammalian cryptochromes have evolved entirely distinct signaling mechanisms. Here we show by developmental and transcriptome analysis that Homo sapiens cryptochrome - 1 (HsCRY1) confers biological activity in transgenic expressing Drosophila in darkness, that can in some cases be further stimulated by light. In contrast to all other cryptochromes, purified recombinant HsCRY1 protein was stably isolated in the anionic radical flavin state, containing only a small proportion of oxidized flavin which could be reduced by illumination. We conclude that animal Type I and Type II cryptochromes may both have signaling mechanisms involving formation of a flavin radical signaling state, and that light independent activity of Type II cryptochromes is a consequence of dark accumulation of this redox form in vivo rather than of a fundamental difference in signaling mechanism

Evidence of mother-child transmission of Helicobacter pylori infection Evidência da transmissão mãe-filho da infecção por Helicobacter pylori

BACKGROUND: Low socioeconomical status is a major risk factor for natural acquisition of Helicobacter pylori (H. pylori) infection in developing countries. Its transmission route is unknown but studies suggest person-to-person transmission. AIM: To evaluate seropositivity of anti-H. pylori antibodies in family members of infected symptomatic index patients as compared to family members of symptomatic uninfected index patients. PATIENTS AND METHODS: One hundred and twelve family members of 38 patients who underwent endoscopy to exclude peptic disease were studied. Patients were deemed H. pylori infected or not infected when rapid urease test and histology were both positive or both negative. The family members underwent ELISA serology using the Cobas Core II Kit (Roche) and were classified into three groups: I - 29 family members of 10 H. pylori (+) duodenal ulcer index patients; II - 57 family members of 17 H. pylori (+) index patients without duodenal ulcer; III - 26 family members of 11 H. pylori (-) index patients. RESULTS: Seropositivity of group I and II (infected patients) was higher than the control group, 83% vs 38%, specially in mothers, 81% vs 18%, and in siblings 76% vs 20%. Differences between fathers' seropositivity was not statistically significant in the three groups: 100% vs 86% vs 70%. Seropositivity of all family members (mother, father and siblings) between infected group (I vs II) was similar. CONCLUSION: Prevalence of H. pylori infection was higher in family members of infected patients, but was similar among family members of infected patients with and without duodenal ulcer. H. pylori infection is more frequent in mothers and siblings of infected index children. A common source of infection cannot be excluded, but facts suggest that person-to-person transmission occurs, specially from mother to child.<br>O estrato socioeconômico baixo é o maior fator de risco para a aquisição natural da infecção por Helicobacter pylori em países em desenvolvimento. As vias de transmissão são desconhecidas embora estudos sugerem transmissão pessoa-pessoa. OBJETIVO: Avaliar a soropositividade de anticorpos anti H. pylori em familiares de pacientes sintomáticos infectados comparados a de pacientes não infectados. CASUÍSTICA E MÉTODOS: Foram estudados 112 familiares de 38 pacientes encaminhados para afastar doença péptica. Os pacientes foram submetidos a exame endoscópico, sendo realizadas quatro biopsias gástricas para pesquisa de H. pylori: duas para teste rápido da urease e duas para histologia (HE/Giemsa). Foi considerado infectado por H. pylori quando ambos os exames resultaram positivos. Nos familiares foi realizada sorologia com método ELISA, utilizando-se o Kit Cobas Core II (Roche), sendo considerado resultado positivo a titulação 7U/mL. Os familiares foram divididos em três grupos: grupo I: 29 familiares de 10 pacientes com úlcera duodenal H. pylori+; grupo II: 57 familiares de 17 pacientes sem úlcera duodenal H. pylori+; grupo III: 26 familiares de 11 pacientes H. pylori-. Foi testada a associação entre grupos e positividade através de uma extensão do teste exato de Fisher (método de Montecarlo SPSS), sendo analisada a soropositividade em cada um dos membros da família: pai, mãe, irmãos e o binômio mãe/pai e para a avaliação de múltiplas variáveis utilizou-se ANOVA. RESULTADOS: Os familiares de pacientes H. pylori+ apresentaram maior soropositividade comparado com o grupo controle, 83% vs 38%, sendo maior nas mães 81% vs 18% e irmãos 76% vs 20%. A soropositividade do pai não foi estatiscamente significante, quando comparados os três grupos de pacientes: 100% vs 86% vs 70%. A soropositividade de todos os membros da família, mãe, pai e irmãos nos grupos de úlcera duodenal H. pylori+ e sem úlcera duodenal H. pylori+ foram semelhantes. CONCLUSÃO: Familiares de pacientes infectados apresentam mais infecção por H. pylori. A soropositividade foi semelhante entre os familiares dos pacientes infectados com e sem úlcera duodenal. Infecção por H. pylori é mais freqüente em mães e irmãos de pacientes infectados; ao contrário, nos pais não houve diferença estatisticamente significante nos três grupos. As crianças apresentam mais infecção quando ambos os pais são H. pylori+ e existe uma concordância do resultado da sorologia entre os cônjuges. Não podemos afastar uma fonte comum de infecção, mas os fatos nos sugerem que a transmissão acontece de pessoa a pessoa e especialmente da mãe para filho e entre os irmãos