Cohesiveness tunes assembly and morphology of FG nucleoporin domain meshworks – Implications for nuclear pore permeability

Nuclear pore complexes control the exchange of macromolecules between the cytoplasm and the nucleus. A selective permeability barrier that arises from a supramolecular assembly of intrinsically unfolded nucleoporin domains rich in phenylalanine-glycine dipeptides (FG domains) fills the nuclear pore. There is increasing evidence that selective transport requires cohesive FG domain interactions. To understand the functional roles of cohesive interactions, we studied monolayers of end-grafted FG domains as a bottom-up nanoscale model system of the permeability barrier. Based on detailed physicochemical analysis of the model films and comparison of the data with polymer theory, we propose that cohesiveness is tuned to promote rapid assembly of the permeability barrier and to generate a stable and compact pore-filling meshwork with a small mesh size. Our results highlight the functional importance of weak interactions, typically a few k_BT per chain, and contribute important information to understand the mechanism of size-selective transport.     

Pollen morphology of Sorghum Moench – Sections Eu-sorghum and Para-sorghum

Pollen morphological studies have been carried out by SEM on 23 species of Sorghum (Gramineae) in order to resolve the exine surface patterns in sections Eu-sorghum (subsection Arundinacea-series Spontanea and Sativa and subsection Halepensia), and Para-sorghum. Basically, two exine ornamentation types have been observed viz. granular and insular. In section Eu-sorghum, series Spontanea and Sativa (of subsection Arundinacea) are heterogeneous having both types of exine pattern. In the same section, subsection Halepensia is characterised by having only a granular exine. Section Para-sorghum shows a marked pollen morphological similarity with subsection Halepensia of section Eu-sorghum. Snowden's concept that the two series Spontanea and Sativa are closely related and that the cultivated Sorghum (series Sativa) might have evolved from the wild Sorghum (series Spontanea), is supported by the present observations.     

Mitochondrial permeability transition pore: a promising target for the treatment of Parkinson’s disease

Among the neurodegenerative diseases (ND), Parkinson’s disease affects 6.3 million people worldwide characterized by the progressive loss of dopaminergic neurons in substantia nigra. The mitochondrial permeability transition pore (mtPTP) is a non-selective voltage-dependent mitochondrial channel whose opening modifies the permeability properties of the mitochondrial inner membrane. It is recognized as a potent pharmacological target for diseases associated with mitochondrial dysfunction and excessive cell death including ND such as Parkinson’s disease (PD). Imbalance in Ca²⁺ concentration, change in mitochondrial membrane potential, overproduction of reactive oxygen species (ROS), or mutation in mitochondrial genome has been implicated in the pathophysiology of the opening of the mtPTP. Different proteins are released by permeability transition including cytochrome c which is responsible for apoptosis. This review aims to discuss the importance of PTP in the pathophysiology of PD and puts together different positive as well as negative aspects of drugs such as pramipexole, ropinirole, minocyclin, rasagilin, and safinamide which act as a blocker or modifier for mtPTP. Some of them may be detrimental in their neuroprotective nature.     

Presence in Legionella pneumophila of a mammalian-like mitochondrial permeability transition pore?

The genome of Legionella pneumophila reveals the presence of a large number of genes coding for eukaryotic-like proteins. By using database searches and homology investigations, we identified three proteins in L. pneumophila whose sequences share similarities with that of eukaryotic polypeptides (lpg0211, lpg1974 and lpg1982). In eukaryotes, the corresponding proteins (PBR, peripheral benzodiazepine receptor; VDAC, voltage-dependant anion channel; and CypD, cyclophilin D) participate in the formation of the mammalian mitochondrial permeability transition pore (MPTP), a complex involved in cell apoptosis. Intriguingly, the presence of these proteins has never been reported in the same bacterium and constitutes, up to now, a unique feature of L. pneumophila. In Legionella, we hypothesize that these proteins are recruited in a multiprotein complex close to the MPTP that may regulate intracellular survival and/or proliferation.     

Quantification of Collagen Ultrastructure after Penetrating Keratoplasty – Implications for Corneal Biomechanics

Purpose

To quantify long-term changes in stromal collagen ultrastructure following penetrating keratoplasty (PK), and evaluate their possible implications for corneal biomechanics.

Methods

A pair of 16 mm post-mortem corneo-scleral buttons was obtained from a patient receiving bilateral penetrating keratoplasty 12 (left)/28 (right) years previously. Small-angle x-ray scattering quantified collagen fibril spacing, diameter and spatial order at 0.5 mm or 0.25 mm intervals along linear scans across the graft margin. Corresponding control data was collected from two corneo-scleral buttons with no history of refractive surgery. Wide-angle x-ray scattering quantified collagen fibril orientation at 0.25 mm (horizontal)×0.25 mm (vertical) intervals across both PK specimens. Quantification of orientation changes in the graft margin were verified by equivalent analysis of data from a 13 year post-operative right PK specimen obtained from a second patient in a previous study, and comparison made with new and published data from normal corneas.

Results

Marked changes to normal fibril alignment, in favour of tangentially oriented collagen, were observed around the entire graft margin in all PK specimens. The total number of meridional fibrils in the wound margin was observed to decrease by up to 40%, with the number of tangentially oriented fibrils increasing by up to 46%. As a result, in some locations the number of fibrils aligned parallel to the wound outnumbered those spanning it by up to five times. Localised increases in fibril spacing and diameter, with an accompanying reduction in matrix order, were also evident.

Conclusions

Abnormal collagen fibril size and spatial order within the PK graft margin are indicative of incomplete stromal wound remodelling and the long term persistence of fibrotic scar tissue. Lasting changes in collagen fibril orientation in and around PK wounds may alter corneal biomechanics and compromise the integrity of the graft-host interface in the long term.     

The multifaceted activity of VEGF in angiogenesis – Implications for therapy responses

Vascular endothelial growth factor (VEGF) is a key growth factor driving angiogenesis (i.e. the formation of new blood vessels) in health and disease. Pharmacological blockade of VEGF signaling to inhibit tumor angiogenesis is clinically approved but the survival benefit is limited as patients invariably acquire resistance. This is partially mediated by the intrinsic flexibility of tumor cells to adapt to VEGF-blockade. However, it has become clear that tumor stromal cells also contribute to the resistance. Originally, VEGF was thought to specifically target endothelial cells (ECs) but it is now clear that many stromal cells also respond to VEGF signaling, making anti-VEGF therapy more complex than initially anticipated. A more comprehensive understanding of the complex responses of stromal cells to VEGF-blockade might inform the design of improved anti-angiogenic agents.     

Dysregulation of cellular microRNAs by human oncogenic viruses – Implications for tumorigenesis

Infection with certain animal and human viruses, often referred to as tumor viruses, induces oncogenic processes in their host. These viruses can induce tumorigenesis through direct and/or indirect mechanisms, and the regulation of microRNAs expression has been shown to play a key role in this process. Some human oncogenic viruses can express their own microRNAs; however, they all can dysregulate the expression of cellular microRNAs, facilitating their respective life cycles. The modulation of cellular microRNAs expression brings consequences to the host cells that may lead to malignant transformation, since microRNAs regulate the expression of genes involved in oncogenic pathways. This review focus on the mechanisms used by each human oncogenic virus to dysregulate the expression of cellular microRNAs, and their impact on tumorigenesis.     

The gastrointestinal tract of Adélie penguins – morphology and function

The aim of this study was to provide data on the morphology of the gastrointestinal tract of Adélie penguins (Pygoscelis adeliae). It was found to consist of a long oesophagus, a two-chambered stomach, a small intestine measuring only 5.22body length, two rudimentary caeca and a short colon, typical of carnivorous birds. The stomach comprised a glandular proventriculus and a muscular gizzard that frequently contained grit. An acidic pH was recorded in both chambers. Ultrastructural studies of the small intestinal mucosal membrane revealed epithelial cells with elongated, irregular microvilli and high affinity for toluidine blue, absorptive intestinal epithelial cells and goblet cells. Numerous large lymphocyte-like cells were observed close to the brush border of the epithelium, and empty spaces on the epithelial surface reflected normal cell loss in the small intestine. The rudimentary caeca and colon provide relatively little volume and time for symbiotic bacteria to aid the digestion of crustacean chitin.     

Control of Dead end localization and activity – Implications for the function of the protein in antagonizing miRNA function

Dead end (dnd) is a vertebrate-specific component of the germ plasm and germ-cell granules that is crucial for germ-cell development in zebrafish and mouse. Dnd counteracts the inhibitory function of miRNAs, thereby facilitating the expression of proteins such as Nanos and Tdrd7 in the germ cells. Here, we show that cis-acting elements within dnd mRNA and the RNA recognition motive (RRM) of the protein are essential for targeting protein expression to the germ cells and to the perinuclear granules, respectively. We demonstrate that as it executes its function, Dnd translocates between the germ-cell nucleus and germ-cell granules. This phenomenon is not observed in proteins mutated in the RRM motif, correlating with loss of function of Dnd. Based on molecular modeling, we identify the putative RNA binding domain of Dnd as a canonical RRM and propose that this domain is important for protein subcellular localization and function.     

ER membrane–bending proteins are necessary for de novo nuclear pore formation

Nucleocytoplasmic transport occurs exclusively through nuclear pore complexes (NPCs) embedded in pores formed by inner and outer nuclear membrane fusion. The mechanism for de novo pore and NPC biogenesis remains unclear. Reticulons (RTNs) and Yop1/DP1 are conserved membrane protein families required to form and maintain the tubular endoplasmic reticulum (ER) and the postmitotic nuclear envelope. In this study, we report that members of the RTN and Yop1/DP1 families are required for nuclear pore formation. Analysis of Saccharomyces cerevisiae prp20-G282S and nup133Δ NPC assembly mutants revealed perturbations in Rtn1–green fluorescent protein (GFP) and Yop1-GFP ER distribution and colocalization to NPC clusters. Combined deletion of RTN1 and YOP1 resulted in NPC clustering, nuclear import defects, and synthetic lethality with the additional absence of Pom34, Pom152, and Nup84 subcomplex members. We tested for a direct role in NPC biogenesis using Xenopus laevis in vitro assays and found that anti-Rtn4a antibodies specifically inhibited de novo nuclear pore formation. We hypothesize that these ER membrane–bending proteins mediate early NPC assembly steps.     

Chromatin-bound NLS proteins recruit membrane vesicles and nucleoporins for nuclear envelope assembly via importin-α/β

The mechanism for nuclear envelope (NE) assembly is not fully understood. Importin-β and the small GTPase Ran have been implicated in the spatial regulation of NE assembly process. Here we report that chromatin-bound NLS (nuclear localization sequence) proteins provide docking sites for the NE precursor membrane vesicles and nucleoporins via importin-α and -β during NE assembly in Xenopus egg extracts. We show that along with the fast recruitment of the abundant NLS proteins such as nucleoplasmin and histones to the demembranated sperm chromatin in the extracts, importin-α binds the chromatin NLS proteins rapidly. Meanwhile, importin-β binds cytoplasmic NE precursor membrane vesicles and nucleoporins. Through interacting with importin-α on the chromatin NLS proteins, importin-β targets the membrane vesicles and nucleoporins to the chromatin surface. Once encountering Ran-GTP on the chromatin generated by RCC1, importin-β preferentially binds Ran-GTP and releases the membrane vesicles and nucleoporins for NE assembly. NE assembly is disrupted by blocking the interaction between importin-α and NLS proteins with excess soluble NLS proteins or by depletion of importin-β from the extract. Our findings reveal a novel molecular mechanism for NE assembly in Xenopus egg extracts.     

Oxidative stress–induced assembly of PML nuclear bodies controls sumoylation of partner proteins

The promyelocytic leukemia (PML) protein organizes PML nuclear bodies (NBs), which are stress-responsive domains where many partner proteins accumulate. Here, we clarify the basis for NB formation and identify stress-induced partner sumoylation as the primary NB function. NB nucleation does not rely primarily on intermolecular interactions between the PML SUMO-interacting motif (SIM) and SUMO, but instead results from oxidation-mediated PML multimerization. Oxidized PML spherical meshes recruit UBC9, which enhances PML sumoylation, allow partner recruitment through SIM interactions, and ultimately enhance partner sumoylation. Intermolecular SUMO–SIM interactions then enforce partner sequestration within the NB inner core. Accordingly, oxidative stress enhances NB formation and global sumoylation in vivo. Some NB-associated sumoylated partners also become polyubiquitinated by RNF4, precipitating their proteasomal degradation. As several partners are protein-modifying enzymes, NBs could act as sensors that facilitate and confer oxidative stress sensitivity not only to sumoylation but also to other post-translational modifications, thereby explaining alterations of stress response upon PML or NB loss.     

The influence of the N-terminal region proximal to the core domain on the assembly and chaperone activity of αB-crystallin

αB-Crystallin (HSPB5) is a small heat-shock protein that is composed of dimers that then assemble into a polydisperse ensemble of oligomers. Oligomerisation is mediated by heterologous interactions between the C-terminal tail of one dimer and the core “α-crystallin” domain of another and stabilised by interactions made by the N-terminal region. Comparatively little is known about the latter contribution, but previous studies have suggested that residues in the region 54–60 form contacts that stabilise the assembly. We have generated mutations in this region (P58A, S59A, S59K, R56S/S59R and an inversion of residues 54–60) to examine their impact on oligomerisation and chaperone activity in vitro. By using native mass spectrometry, we found that all the αB-crystallin mutants were assembly competent, populating similar oligomeric distributions to wild-type, ranging from 16-mers to 30-mers. However, circular dichroism spectroscopy, intrinsic tryptophan and bis-ANS fluorescence studies demonstrated that the secondary structure differs to wild type, the 54–60 inversion mutation having the greatest impact. All the mutants exhibited a dramatic decrease in exposed hydrophobicity. We also found that the mutants in general were equally active as the wild-type protein in inhibiting the amorphous aggregation of insulin and seeded amyloid fibrillation of α-synuclein in vitro, except for the 54–60 inversion mutant, which was significantly less effective at inhibiting insulin aggregation. Our data indicate that alterations in the part of the N-terminal region proximal to the core domain do not drastically affect the oligomerisation of αB-crystallin, reinforcing the robustness of αB-crystallin in functioning as a molecular chaperone.     

Low Grade Gliomas in Eloquent Locations – Implications for Surgical Strategy,Survival and Long Term Quality of Life

Background

Surgical management of suspected LGG remains controversial. A key factor when deciding a surgical strategy is often the tumors’ perceived relationship to eloquent brain regions

Objective

To study the association between tumor location, survival and long-term health related quality of life (HRQL) in patients with supratentorial low-grade gliomas (LGG).

Methods

Adults (≥18 years) operated due to newly diagnosed LGG from 1998 through 2009 included from two Norwegian university hospitals. After review of initial histopathology, 153 adults with supratentorial WHO grade II LGG were included in the study. Tumors’ anatomical location and the relationship to eloquent regions were graded. Survival analysis was adjusted for known prognostic factors and the initial surgical procedure (biopsy or resection). In long-term survivors, HRQL was assessed with disease specific questionnaires (EORTC QLQ-C30 and BN20) as well as a generic questionnaire (EuroQol 5D).

Results

There was a significant association between eloquence and survival (log-rank, p<0.001). The estimated 5-year survival was 77% in non-eloquent tumors, 71% in intermediate located tumors and 54% in eloquent tumors. In the adjusted analysis the hazard ratio of increasing eloquence was 1.5 (95% CI 1.1–2.0, p = 0.022). There were no differences in HRQL between patients with eloquent and non-eloquent tumors. The most frequent self-reported symptoms were related to fatigue, cognition, and future uncertainty.

Conclusion

Eloquently located LGGs are associated with impaired survival compared to non-eloquently located LGG, but in long-term survivors HRQL is similar. Although causal inference from observational data should be done with caution, the findings illuminate the delicate balance in surgical decision making in LGGs, and add support to the probable survival benefits of aggressive surgical strategies, perhaps also in eloquent locations.