Structural and functional studies of RegB, a new member of a family of sequence-specific ribonucleases involved in mRNA inactivation on the ribosome

The RegB endoribonuclease participates in the bacteriophage T4 life cycle by favoring early messenger RNA breakdown. RegB specifically cleaves GGAG sequences found in intergenic regions, mainly in translation initiation sites. Its activity is very low but can be enhanced up to 100-fold by the ribosomal 30 S subunit or by ribosomal protein S1. RegB has no significant sequence homology to any known protein. Here we used NMR to solve the structure of RegB and map its interactions with two RNA substrates. We also generated a collection of mutants affected in RegB function. Our results show that, despite the absence of any sequence homology, RegB has structural similarities with two Escherichia coli ribonucleases involved in mRNA inactivation on translating ribosomes: YoeB and RelE. Although these ribonucleases have different catalytic sites, we propose that RegB is a new member of the RelE/YoeB structural and functional family of ribonucleases specialized in mRNA inactivation within the ribosome.     

Listeria monocytogenes ferritin protects against multiple stresses and is required for virulence

In this study, the role of Listeria monocytogenes ferritin was investigated. The fri gene encoding the ferritin was deleted and the phenotype of the mutant was analyzed demonstrating that ferritin is necessary for optimal growth in minimal medium in both presence and absence of iron, as well as after cold- and heat-shock. We also showed that ferritin provides protection against reactive oxygen species and is essential for full virulence of L. monocytogenes. A comparative proteomic analysis revealed an effect of the fri deletion on the levels of listeriolysin O and several stress proteins. Together, our study demonstrates that fri has multiple roles that contribute to Listeria virulence.     

Possible role of region 152-156 in the structural duality of a peptide fragment from sheep prion protein

The conformational conversion of the nonpathogenic "cellular" prion isoform into a pathogenic "scrapie" protease-resistant isoform is a fundamental event in the onset of transmissible spongiform encephalopathies (TSE). During this pathogenic conversion, helix H1 and its two flanking loops of the normal prion protein are thought to undergo a conformational transition into a beta-like structure. A peptide spanning helix H1 and beta-strand S2 (residues 142-166 in human numbering) was studied by circular dichroism and nuclear magnetic resonance spectroscopies. This peptide in aqueous solution, in contrast to many prion fragments studied earlier (1) is highly soluble and (2) does not aggregate until the millimolar concentration range, and (3) exhibits an intrinsic propensity to a beta-hairpin-like conformation at neutral pH. We found that this peptide can also fold into a helix H1 conformation when dissolved in a TFE/PB mixture. The structures of the peptide calculated by MD showed solvent-dependent internal stabilizing forces of the structures and evidenced a higher mobility of the residues following the end of helix H1. These data suggest that the molecular rearrangement of this peptide in region 152-156, particularly in position 155, could be associated with the pathogenic conversion of the prion protein.     

Shrub cover in northern Nunavik: can herbivores limit shrub expansion?

Recent climate changes have increased the primary productivity of many Arctic and subarctic regions. Erected shrub has been shown to increase in abundance over the last decades in northern regions in response to warmer climate. At the same time, caribou herds are declining throughout the circumboreal regions. Based on observation of heavy browsing on shrubs at Deception Bay (Nunavik, Canada), we hypothesized that the densification of shrubs observed in nearby locations did not occur at our study site despite of observed warming because of a recent peak of the Rivière-aux-Feuilles caribou herd. To assess shrub cover changes, we compared a 1972 mosaic of aerial photos to a 2010 satellite image over a 5 km² area, divided into 56 grids of 100 30 m × 30 m cells. Most cells (n = 4,502) did not show any changes in the cover of shrubs but those who did were as likely to increase as to decrease. The relative cover of shrubs in cells who changed was not higher in 2010 (6.1 ± 0.2 %) than in 1972 (7.3 ± 0.4 %). More than 70 % of birch and willow had more than 50 % of their shoot browsed, suggesting that caribou may limit shrub expansion at this site. We cannot rule out that abiotic factors also contribute to the inertia in shrub cover. Increases in shrub abundance reported in Nunavik and elsewhere were located closer to the tree line or in discontinuous permafrost, whereas our site is characterized by herbaceous arctic tundra, continuous permafrost and relatively low annual precipitation.     

Cucumber mosaic virus genome is encapsidated in alfalfa mosaic virus coat protein expressed in transgenic tobacco plants

The expression of viral coat protein (CP) in transgenic plants has been shown to be very effective in virus plant protection. However, the introduction of CP genes into plants presents the potential risk of the encapsidation of a superinfecting viral genome in the transgenic protein, an event which could change the epidemiology of the disease. To detect the potential heterologous encapsidation of the cucumber mosaic virus (CMV) genome by alfalfa mosaic virus (AIMV) CP expressed in transgenic tobacco plants, a system of immunocapture (IC) and amplification by polymerase chain reaction (PCR) was optimized. This provided high sensitivity and reliable selection of the heterologously encapsidated CMV genome in the presence of natural CMV particles. As little as 2 pg of virus could be detected by immunocapture/polymerase chain reaction (IC/PCR) technique. Evidence for heterologous encapsidation of the CMV genome was found in 11 of the 33 transgenic plants tested two weeks after CMV inoculation. This demonstrates a significant rate of heterologous encapsidation events between two unrelated viruses in transgenic plants. Since CP is involved in the interactions of the virus particle with its vector, the release in the field of such transgenic plants could alter the transmission properties of some important viruses.     

Estrone sulfate (E1S), a prognosis marker for tumor aggressiveness in prostate cancer (PCa)

Seeking insight into the possible role of estrogens in prostate cancer (PCa) evolution, we assayed serum E₂, estrone (E₁), and estrone sulfate (E₁S) in 349 PCa and 100 benign prostatic hyperplasia (BPH) patients, and in 208 control subjects in the same age range (50–74 years).

E₁ (pmol/L ± S.D.) and E₁S (nmol/L ± S.D.) in the PCa and BPH patients (respectively 126.1 ± 66.1 and 2.82 ± 1.78, and 127.8 ± 56.4 and 2.78 ± 2.12) were significantly higher than in the controls (113.8 ± 47.6 and 2.11 ± 0.96). E₂ was not significantly different among the PCa, BPH, and control groups. These assays were also carried out in PCa patients after partition by prognosis (PSA, Gleason score (GS), histological stage, and surgical margins (SM)). Significantly higher E₁S levels were found in PCa with: PSA > 10 ng/L (3.05 ± 1.92) versus PSA ≤ 10 ng/mL (2.60 ± 1.55), stage pT3-T4 (2.99 ± 1.80) versus pT2 (2.58 ± 1.58), and positive (3.26 ± 1.95) versus negative margins (2.52 ± 1.48). E₁ was higher in poor- than in better-prognosis PCa. E₂ was significantly higher in PCa with GS ≥ 4 + 3 (109.5 ± 43.8) versus GS ≤ 3 + 4 (100.6 ± 36.5) and increased significantly when GS increased from 3 + 3 to 4 + 4. Estrogens, especially E₁S appeared to be possible markers of PCa progression.

Attempting to identify potential sources of E₂ in PCa according to prognosis, as well as in BPH, we found a significant correlation coefficient between E₁S and E₂ (0.266–0.347) in poor-prognosis PCa and no correlation in BPH (0.026) and better-prognosis PCa (0.013–0.104).

It is as though during progression of PCa from good to poor prognosis there were a shift in the E₁ to E₂ metabolic pathway from predominantly oxidative to predominantly reductive.     

Multimodal optical analysis of meningioma and comparison with histopathology

Meningioma is the most frequent primary central nervous system tumor. The risk of recurrence and the prognosis are correlated with the extent of the resection that ideally encompasses the infiltrated dura mater and, if required, the infiltrated bone. No device can deliver real‐time intraoperative histopathological information on the tumor environment to help the neurosurgeon to achieve a gross total removal. This study assessed the abilities of nonlinear microscopy to provide relevant and real‐time data to help resection of meningiomas. Nine human meningioma samples (four World Health Organization Grade I, five Grade II) were analyzed using different optical modalities: spectral analysis and imaging, lifetime measurements, fluorescence lifetime imaging microscopy, fluorescence emitted under one‐ and two‐photon excitation and the second‐harmonic generation signal imaging using a multimodal setup. Nonlinear microscopy produced images close to histopathology as a gold standard. The second‐harmonic generation signal delineated the collagen background and two‐photon fluorescence underlined cell cytoplasm. The matching between fluorescence images and Hematoxylin and Eosin staining was possible in all cases. Grade I meningioma emitted less autofluorescence than Grade II meningioma and Grade II meningioma exhibited a distinct lifetime value. Autofluorescence was correlated with the proliferation rates and seemed to explain the observed differences between Grade I and II meningiomas. This preliminary multimodal study focused on human meningioma samples confirms the potential of tissue autofluorescence analysis and nonlinear microscopy in helping intraoperatively neurosurgeons to reach the actual boundaries of the tumor infiltration.

Correspondence between H&E staining (top pictures) and the two‐photon fluorescence imaging (bottom pictures)