Molecular characterization of phytoplasmas in lilies with fasciation in the Czech Republic

Lilium spp. with symptoms of severe fasciation were observed in Southern and central Bohemia during the period 1999-2003. Nucleic acids extracted from symptomatic and asymptomatic plants were used in nested-PCR assays with primers amplifying 16S-23S rRNA sequences specific for phytoplasmas. The subsequent nested-PCR with phytoplasma group-specific primers followed by RFLP analyses and the 16S ribosomal gene sequencing, allowed classification of the detected phytoplasmas in the aster yellows group, subgroups 16SrI-B and 16SrI-C alone, and in mixed infection. Samples infected by 16SrI-C phytoplasmas showed different overlapping RFLP profiles after TruI digestion of R16F2/R2 amplicons. Two of these amplicons were sequenced, one of them directly and the other after cloning; sequence analyses and blast alignment confirmed the presence of two different overlapping patterns in samples studied. The sequences obtained were closely related, respectively, to operon A and operon B ribosomal sequences of the clover phyllody phytoplasma. Direct PCR followed by RFLP analyses of the tuf gene with two restriction enzymes showed no differences from reference strain of subgroup 16SrI-C. Infection with aster yellows phytoplasmas of 16SrI-B subgroup in asymptomatic lilies cv. Sunray was also detected.     

A soluble carotenoid protein involved in phycobilisome-related energy dissipation in cyanobacteria

Photosynthetic organisms have developed multiple protective mechanisms to survive under high-light conditions. In plants, one of these mechanisms is the thermal dissipation of excitation energy in the membrane-bound chlorophyll antenna of photosystem II. The question of whether or not cyanobacteria, the progenitor of the chloroplast, have an equivalent photoprotective mechanism has long been unanswered. Recently, however, evidence was presented for the possible existence of a mechanism dissipating excess absorbed energy in the phycobilisome, the extramembrane antenna of cyanobacteria. Here, we demonstrate that this photoprotective mechanism, characterized by blue light-induced fluorescence quenching, is indeed phycobilisome-related and that a soluble carotenoid binding protein, ORANGE CAROTENOID PROTEIN (OCP), encoded by the slr1963 gene in Synechocystis PCC 6803, plays an essential role in this process. Blue light is unable to quench fluorescence in the absence of phycobilisomes or OCP. The fluorescence quenching is not DeltapH-dependent, and it can be induced in the absence of the reaction center II or the chlorophyll antenna, CP43 and CP47. Our data suggest that OCP, which strongly interacts with the thylakoids, acts as both the photoreceptor and the mediator of the reduction of the amount of energy transferred from the phycobilisomes to the photosystems. These are novel roles for a soluble carotenoid protein.     

Aromatase inhibitors in ovarian stimulation

The selective estrogen receptor modulator, clomiphene citrate (CC), has been the principal drug used for induction of ovulation in women with polycystic ovarian syndrome (PCOS). CC is associated with adverse side effects and low pregnancy rates attributed to long-lasting estrogen receptor depletion. Anastrozole and letrozole are potent, non-steroidal, reversible aromatase inhibitors, developed for postmenopausal breast cancer therapy. We hypothesized that aromatase inhibitors could mimic the action of CC in reducing estrogen negative feedback on follicle stimulating hormone (FSH) secretion, without depleting estrogen receptors. In a series of preliminary studies, we reported the success of aromatase inhibition in inducing ovulation in anovulatory women with PCOS. Moreover, we showed that concomitant use of aromatase inhibitors resulted in a significant reduction of the FSH dose needed for controlled ovarian hyperstimulation. We suggest that aromatase inhibitors act through an increase in endogenous gonadotropin secretion as well as through increased intraovarian androgen levels that may increase ovarian FSH receptors. Recently, we demonstrated the safety of aromatase inhibitors in pregnancy outcome studies examining spontaneous pregnancy loss, multiple pregnancy rates and congenital anomalies compared to a control group of infertility patients treated with CC.     

Mapping changes to vegetation pattern in a restoring wetland: Finding pattern metrics that are consistent across spatial scale and time

Tidal salt marshes in the San Francisco Estuary region display heterogeneous vegetation patterns that influence wetland function and provide adequate habitat for native or endangered wildlife. In addition to analyzing the extent of vegetation, monitoring the dynamics of vegetation pattern within restoring wetlands can offer valuable information about the restoration process. Pattern metrics, derived from classified remotely sensed imagery, have been used to measure composition and configuration of patches and landscapes, but they can be unpredictable across scales, and inconsistent across time. We sought to identify pattern metrics that are consistent across spatial scale and time – and thus robust measures of vegetation and habitat configuration – for a restored tidal marsh in the San Francisco Bay, CA, USA. We used high-resolution (20 cm) remotely sensed color infrared imagery to map vegetation pattern over 2 years, and performed a multi-scale analysis of derived vegetation pattern metrics. We looked at the influence on metrics of changes in grain size through resampling and changes in minimum mapping unit (MMU) through smoothing. We examined composition, complexity, connectivity and heterogeneity metrics, focusing on perennial pickleweed (Sarcocornia pacifica), a dominant marsh plant. At our site, pickleweed patches grew larger, more irregularly shaped, and closely spaced over time, while the overall landscape became more diverse. Of the two scale factors examined, grain size was more consistent than MMU in terms of identifying relative change in composition and configuration of wetland marsh vegetation over time. Most metrics exhibited unstable behavior with larger MMUs. With small MMUs, most metrics were consistent across grain sizes, from fine (e.g. 0.16 m²) to relatively large (e.g. 16 m²) pixel sizes. Scale relationships were more variable at the landcover class level than at the landscape level (across all classes). This information may be useful to applied restoration practitioners, and adds to our general understanding of vegetation change in a restoring marsh.     

Genome-wide DNA methylation indicates silencing of tumor suppressor genes in uterine leiomyoma

Background

Uterine leiomyomas, or fibroids, represent the most common benign tumor of the female reproductive tract. Fibroids become symptomatic in 30% of all women and up to 70% of African American women of reproductive age. Epigenetic dysregulation of individual genes has been demonstrated in leiomyoma cells; however, the in vivo genome-wide distribution of such epigenetic abnormalities remains unknown.

Principal Findings

We characterized and compared genome-wide DNA methylation and mRNA expression profiles in uterine leiomyoma and matched adjacent normal myometrial tissues from 18 African American women. We found 55 genes with differential promoter methylation and concominant differences in mRNA expression in uterine leiomyoma versus normal myometrium. Eighty percent of the identified genes showed an inverse relationship between DNA methylation status and mRNA expression in uterine leiomyoma tissues, and the majority of genes (62%) displayed hypermethylation associated with gene silencing. We selected three genes, the known tumor suppressors KLF11, DLEC1, and KRT19 and verified promoter hypermethylation, mRNA repression and protein expression using bisulfite sequencing, real-time PCR and western blot. Incubation of primary leiomyoma smooth muscle cells with a DNA methyltransferase inhibitor restored KLF11, DLEC1 and KRT19 mRNA levels.

Conclusions

These results suggest a possible functional role of promoter DNA methylation-mediated gene silencing in the pathogenesis of uterine leiomyoma in African American women.     

Characterization of a model cyanobacterium Synechocystis sp. PCC 6803 autotrophic growth in a flat‐panel photobioreactor

We characterized the photoautotrophic growth of glucose‐tolerant Synechocystis sp. PCC 6803 in a flat‐panel photobioreactor running on a semicontinuous regime under various lights, temperatures, and influx carbon dioxide concentrations. The maximum reached growth rate was 0.135 h^?1, which corresponds to a doubling time of 5.13 h—a growth speed never reported for Synechocystis before. Saturating red light intensity for the strain was 220–360 μmol(photons) m^?2 s^?1, and we did not observe any photoinhibition up to 660 μmol(photons) m^?2 s^?1. Synechocystis was able to grow under red light only; however, photons of wavelengths 405–585 and 670–700 nm further improved its growth. Optimal growth temperature was 35°C. Below 32°C, the growth rates decreased linearly with temperature coefficient (Q₁₀) 1.70. Semicontinuous cultivation is known to be efficient for growth characterization and optimization. However, the assumption of correct growth rates calculation—culture exponential growth—is often not fulfilled. The semicontinuous setup in this study was operated as a turbidostat. Accurate online OD measurements with high time‐resolution allowed fast and reliable growth rates determination. Repeating diluting frequencies (up to 18 dilutions per day) were essential for rapid growth stability evaluation. The presented setup provides improvement to previously published semicontinuous characterization strategies by decreasing experimental time requirements and maintaining the culture in exponential growth phase throughout the entire characterization procedure.