Genome-wide RNA polymerase II profiles and RNA accumulation reveal kinetics of transcription and associated epigenetic changes during diurnal cycles.

Interactions of cell-autonomous circadian oscillators with diurnal cycles govern the temporal compartmentalization of cell physiology in mammals. To understand the transcriptional and epigenetic basis of diurnal rhythms in mouse liver genome-wide, we generated temporal DNA occupancy profiles by RNA polymerase II (Pol II) as well as profiles of the histone modifications H3K4me3 and H3K36me3. We used these data to quantify the relationships of phases and amplitudes between different marks. We found that rhythmic Pol II recruitment at promoters rather than rhythmic transition from paused to productive elongation underlies diurnal gene transcription, a conclusion further supported by modeling. Moreover, Pol II occupancy preceded mRNA accumulation by 3 hours, consistent with mRNA half-lives. Both methylation marks showed that the epigenetic landscape is highly dynamic and globally remodeled during the 24-hour cycle. While promoters of transcribed genes had tri-methylated H3K4 even at their trough activity times, tri-methylation levels reached their peak, on average, 1 hour after Pol II. Meanwhile, rhythms in tri-methylation of H3K36 lagged transcription by 3 hours. Finally, modeling profiles of Pol II occupancy and mRNA accumulation identified three classes of genes: one showing rhythmicity both in transcriptional and mRNA accumulation, a second class with rhythmic transcription but flat mRNA levels, and a third with constant transcription but rhythmic mRNAs. The latter class emphasizes widespread temporally gated posttranscriptional regulation in the mouse liver.

PAX5 activates the transcription of the human telomerase reverse transcriptase gene in B cells.

Telomerase is an RNA-dependent DNA polymerase that synthesizes telomeric DNA. Its activity is not detectable in most somatic cells but it is reactivated during tumorigenesis. In most cancers, the combination of hTERT hypermethylation and hypomethylation of a short promoter region is permissive for low-level hTERT transcription. Activated and malignant lymphocytes express high telomerase activity, through a mechanism that seems methylation-independent. The aim of this study was to determine which mechanism is involved in the enhanced expression of hTERT in lymphoid cells. Our data confirm that in B cells, some T cell lymphomas and non-neoplastic lymph nodes, the hTERT promoter is unmethylated. Binding sites for the B cell-specific transcription factor PAX5 were identified downstream of the ATG translational start site through EMSA and ChIP experiments. ChIP assays indicated that the transcriptional activation of hTERT by PAX5 does not involve repression of CTCF binding. In a B cell lymphoma cell line, siRNA-induced knockdown of PAX5 expression repressed hTERT transcription. Moreover, ectopic expression of PAX5 in a telomerase-negative normal fibroblast cell line was found to be sufficient to activate hTERT expression. These data show that activation of hTERT in telomerase-positive B cells is due to a methylation-independent mechanism in which PAX5 plays an important role.

Signature polymorphisms in the internal transcribed spacer region relevant for the differentiation of zoophilic and anthropophilic strains of Trichophyton interdigitale and other species of T. mentagrophytes sensu lato.

Summary Background Dermatophytes are the main cause of superficial mycoses in humans and animals. Molecular research has given useful insights into the phylogeny and taxonomy of the dermatophytes to overcome the difficulties with conventional diagnostics. Objectives The Trichophyton mentagrophytes complex consists of anthropophilic as well as zoophilic species. Although several molecular markers have been developed for the differentiation of strains belonging to T. mentagrophytes sensu lato, correct identification still remains problematic, especially concerning the delineation of anthropophilic and zoophilic strains of T. interdigitale. This differentiation is not academic but is essential for selection of the correct antimycotic therapy to treat infected patients. Methods One hundred and thirty isolates identified by morphological characteristics as T. mentagrophytes sensu lato were investigated using restriction fragment length polymorphism (RFLP) and sequence analysis of the polymerase chain reaction-amplified internal transcribed spacer (ITS) region of the rDNA. Results Species of this complex produced individual RFLP patterns obtained by the restriction enzyme MvaI. Subsequent sequence analysis of the ITS1, 5.8S and ITS2 region of all strains, but of T. interdigitale in particular, revealed single unique polymorphisms in anthropophilic and zoophilic strains. Conclusions Signature polymorphisms were observed to be useful for the differentiation of these strains and epidemiological data showed a host specificity among zoophilic strains of T. interdigitale/Arthroderma vanbreuseghemii compared with A. benhamiae as well as characteristic clinical pictures in humans when caused by zoophilic or anthropophilic strains. The delineation is relevant because it helps in determining the correct treatment and provides clues regarding the source of the infection.

Effect of humeral stem design on humeral position and range of motion in reverse shoulder arthroplasty.

PURPOSE: The impacts of humeral offset and stem design after reverse shoulder arthroplasty (RSA) have not been well-studied, particularly with regard to newer stems which have a lower humeral inclination. The purpose of this study was to analyze the effect of different humeral stem designs on range of motion and humeral position following RSA. METHODS: Using a three-dimensional computer model of RSA, a traditional inlay Grammont stem was compared to a short curved onlay stem with different inclinations (155°, 145°, 135°) and offset (lateralised vs medialised). Humeral offset, the acromiohumeral distance (AHD), and range of motion were evaluated for each configuration. RESULTS: Altering stem design led to a nearly 7-mm change in humeral offset and 4 mm in the AHD. Different inclinations of the onlay stems had little influence on humeral offset and larger influence on decreasing the AHD. There was a 10° decrease in abduction and a 5° increase in adduction between an inlay Grammont design and an onlay design with the same inclination. Compared to the 155° model, the 135° model improved adduction by 28°, extension by 24° and external rotation of the elbow at the side by 15°, but led to a decrease in abduction of 9°. When the tray was placed medially, on the 145° model, a 9° loss of abduction was observed. CONCLUSIONS: With varus inclination prostheses (135° and 145°), elevation remains unchanged, abduction slightly decreases, but a dramatic improvement in adduction, extension and external rotation with the elbow at the side are observed.