Collapsin Response Mediator Protein 5 (CRMP5) Induces Mitophagy,Thereby Regulating Mitochondrion Numbers in Dendrites

Degradation of damaged mitochondria by mitophagy is an essential process to ensure cell homeostasis. Because neurons, which have a high energy demand, are particularly dependent on the mitochondrial dynamics, mitophagy represents a key mechanism to ensure correct neuronal function. Collapsin response mediator proteins 5 (CRMP5) belongs to a family of cytosolic proteins involved in axon guidance and neurite outgrowth signaling during neural development. CRMP5, which is highly expressed during brain development, plays an important role in the regulation of neuronal polarity by inhibiting dendrite outgrowth at early developmental stages. Here, we demonstrated that CRMP5 was present in vivo in brain mitochondria and is targeted to the inner mitochondrial membrane. The mitochondrial localization of CRMP5 induced mitophagy. CRMP5 overexpression triggered a drastic change in mitochondrial morphology, increased the number of lysosomes and double membrane vesicles termed autophagosomes, and enhanced the occurrence of microtubule-associated protein 1 light chain 3 (LC3) at the mitochondrial level. Moreover, the lipidated form of LC3, LC3-II, which triggers autophagy by insertion into autophagosomes, enhanced mitophagy initiation. Lysosomal marker translocates at the mitochondrial level, suggesting autophagosome-lysosome fusion, and induced the reduction of mitochondrial content via lysosomal degradation. We show that during early developmental stages the strong expression of endogenous CRMP5, which inhibits dendrite growth, correlated with a decrease of mitochondrial content. In contrast, the knockdown or a decrease of CRMP5 expression at later stages enhanced mitochondrion numbers in cultured neurons, suggesting that CRMP5 modulated these numbers. Our study elucidates a novel regulatory mechanism that utilizes CRMP5-induced mitophagy to orchestrate proper dendrite outgrowth and neuronal function.     

Sustained clinical efficacy of sulfadoxine-pyrimethamine for uncomplicated falciparum malaria in Malawi after 10 years as first line treatment: five year prospective study

Objective To measure the efficacy of sulfadoxine-pyrimethamine treatment of falciparum malaria in Malawi from 1998 to 2002, after a change from chloroquine to sulfadoxine-pyrimethamine as first line treatment in that country in 1993.Design Prospective open label drug efficacy study.Setting Health centre in large peri-urban township adjacent to Blantyre, Malawi.Participants People presenting to a health centre with uncomplicated Plasmodium falciparum malaria.Main outcome measures Therapeutic efficacy and parasitological resistance to standard sulfadoxine-pyrimethamine treatment at 14 days and 28 days of follow up.Results Therapeutic efficacy remained stable, with adequate clinical response rates of 80% or higher throughout the five years of the study. Analysis of follow up to 28 days showed modest but significant trends towards diminishing clinical and parasitological efficacy over time within the study period.Conclusion Contrary to expectations, sulfadoxine-pyrimethamine has retained good efficacy after 10 years as the first line antimalarial drug in Malawi. African countries with very low chloroquine efficacy, high sulfadoxine-pyrimethamine efficacy, and no other immediately available alternatives may benefit from interim use of sulfadoxine-pyrimethamine while awaiting implementation of combination antimalarial treatments.     

Female behaviour prior to additional sperm receipt in the hermaphroditic pond snail Lymnaea stagnalis

In many species, sexual resistance by females can fluctuate depending on different physiological and/or ecological conditions. Based on theory, we decided to test whether the simultaneous hermaphrodite Lymnaea stagnalis becomes more reluctant to mate in the female role after receipt of an ejaculate. We aimed to test whether receipt of an ejaculate affects the expression of recipient behaviours, and whether this can have a potential effect on the outcome of mating interactions. We found that the expression of Crawl-out and Biting behaviours increased significantly in recently inseminated snails. Our data also indicated that an increase in expression of these latter behaviours can have direct effects on the outcome of the mating interactions. These findings suggest that under the influence of recent insemination, this hermaphroditic snail is more reluctant to accept another mating in the female role and may try to actively discourage the mounting snail (sperm donor). Hence, despite the fact that these simultaneous hermaphrodites express both sexual functions, there can still be variation in motivation to mate in either role.     

Wound healing applications of biogenic colloidal silver and gold nanoparticles: recent trends and future prospects

Applied Microbiology and Biotechnology - Nanotechnology has emerged as a prominent scientific discipline in the technological revolution of this millennium. The scientific community has focused on...     

A virus-derived short hairpin RNA confers resistance against sugarcane mosaic virus in transgenic sugarcane

RNA interference (RNAi) is commonly used to produce virus tolerant transgenic plants. The objective of the current study was to generate transgenic sugarcane plants expressing a short hairpin RNAs (shRNA) targeting the coat protein (CP) gene of sugarcane mosaic virus (SCMV). Based on multiple sequence alignment, including genomic sequences of four SCMV strains, a conserved region of ~ 456 bp coat protein (CP) gene was selected as target gene and amplified through polymerase chain reaction (PCR). Subsequently, siRNAs2 and siRNA4 were engineered as stable short hairpin (shRNA) transgenes of 110 bp with stem and loop sequences derived from microRNA (sof-MIR168a; an active regulatory miRNA in sugarcane). These transgenes were cloned in independent RNAi constructs under the control of the polyubiquitin promoter. The RNAi constructs were delivered into two sugarcane cultivars ‘SPF-234 and NSG-311 in independent experiments using particle bombardment. Molecular identification through PCR and Southern blot revealed anti-SCMV positive transgenic lines. Upon mechanical inoculation of transgenic and non-transgenic sugarcane lines with SCMV, the degree of resistance was found variable among the two sugarcane cultivars. For sugarcane cultivar NSG-311, the mRNA expression of the CP–SCMV was reduced to 10% in shRNA2-transgenic lines and 80% in shRNA4-transgenic lines. In sugarcane cultivar SPF-234, the mRNA expression of the CP–SCMV was reduced to 20% in shRNA2-transgenic lines and 90% in shRNA4 transgenic lines, revealing that transgenic plants expressing shRNA4 were almost immune to SCMV infection.     

The effect of slow-growth strategy on a production of Petunia × hybrida Vilm. microcuttings

Petunia × hybrida Vilm. is a fast-growing ornamental plant that was cultured under varying storage conditions to address seasonal fluctuations in microcutting demand. The effects of storage period (16 to 32 wk), temperature (15 to 23°C), low and high light intensity, sucrose (1 to 5% w/v), and mannitol (0 to 4% w/v) in factorial arrangements were analyzed. Stored and non-stored shoots were compared for microcutting production, harvested twice at 3-wk intervals, and were subsequently transferred to the greenhouse for 17 d. Nearly all plants from 16-wk storage survived well at all treatment conditions and the quality and quantity of microcuttings were enhanced from shoots stored at 15°C and 3% (w/v) sucrose, without mannitol for all storage periods. Another experiment tested 11- to 25-wk storage period with Petunia hybrida ‘Ragtime’ and ‘Suncatcher’ at 12°C and low light intensity. Repeated cycles of microcutting at 2-wk intervals were extended with ex vitro rooting in the greenhouse for 15 d. More and better quality microcuttings were obtained from the second and third cutting cycles than from the first or fourth cycles. By reducing temperature and light intensity, Petunia hybrida was successfully stored for 32 wk (without mannitol). A seasonal schedule with a short production window, followed by cutting large numbers of high-quality shoot-tips, could be affected and efficiently managed through storage. The value of stored shoots was enhanced by extending the number of times a shoot could be cut over repeated cutting cycles with a gain in microcutting quality.

     

Extracellular identification of a processed type II ComR/ComS pheromone of Streptococcus mutans

The competence-stimulating peptide (CSP) and the sigX-inducing peptide (XIP) are known to induce Streptococcus mutans competence for genetic transformation. For both pheromones, direct identification of the native peptides has not been accomplished. The fact that extracellular XIP activity was recently observed in a chemically defined medium devoid of peptides, as mentioned in an accompanying paper (K. Desai, L. Mashburn-Warren, M. J. Federle, and D. A. Morrison, J. Bacteriol. 194:3774-3780, 2012), provided ideal conditions for native XIP identification. To search for the XIP identity, culture supernatants were filtered to select for peptides of less than 3 kDa, followed by C(18) extraction. One peptide, not detected in the supernatant of a comS deletion mutant, was identified by tandem mass spectrometry (MS/MS) fragmentation as identical to the ComS C-terminal sequence GLDWWSL. ComS processing did not require Eep, a peptidase involved in processing or import of bacterial small hydrophobic peptides, since eep deletion had no inhibitory effect on XIP production or on synthetic XIP response. We investigated whether extracellular CSP was also produced. A reporter assay for CSP activity detection, as well as MS analysis of supernatants, revealed that CSP was not present at detectable levels. In addition, a mutant with deletion of the CSP-encoding gene comC produced endogenous XIP levels similar to those of a nondeletion mutant. The results indicate that XIP pheromone production is a natural phenomenon that may occur in the absence of natural CSP pheromone activity and that the heptapeptide GLDWWSL is an extracellular processed form of ComS, possibly the active XIP pheromone. This is the first report of direct identification of a ComR/ComS pheromone.     

A strategy for mitigating avian colibacillosis disease using plant growth promoting rhizobacteria and green synthesized zinc oxide nanoparticles

Avian colibacillosis caused by the zoonotic pathogen Escherichia coli is a common bacterial infection that causes major losses in the poultry sector. Extracts of different medicinal plants and antibiotics have been used against poultry bacterial pathogens. However, overuse of antibiotics and extracts against pathogenic strains leads to the proliferation of multi-drug resistant bacteria. Due to their environmentally friendly nature, nanotechnology and beneficial bacterial strains can be used as effective strategies against poultry infections. Green synthesis of zinc oxide nanoparticles (ZnO-NPs) from Eucalyptus globulus leaves was carried out in this study. Their characterization was done by UV–vis spectroscopy, X-ray diffraction (XRD), and Fourier transmission infrared spectroscopy (FT-IR) which confirmed their synthesis, structure, and size. In vitro, antimicrobial activities of plant leaf extract, ZnO-NPs, and plant growth-promoting rhizobacteria (PGPR) were checked against E. coli using well diffusion as well as disc diffusion method. Results proved that the antimicrobial activity of ZnO-NPs and PGPR strains was more enhanced when compared to eucalyptus leaf extract at 36 h. The maximum relative inhibition shown by ZnO-NPs, PGPR strains and eucalyptus leaf extracts was 88%, 67% and 58%, respectively. The effectiveness of ZnO-NPs was also increased with an increase in particle dose and treatment time. The 90 mg/ml of ZnO-NPs was more effective. PGPR strains from all over the tested strains, Pseudomonas sp. (HY8N) exhibited a strong antagonism against the E. coli strain as compared to other PGPR strains used in this study. However, combined application of PGPR (Pseudomonas sp. (HY8N)) and ZnO-NPs augment antagonistic effects and showed maximum 69% antagonism. The study intends to investigate the binding affinity of ZnO-NPs with the suitable receptor of the bacterial pathogen by in silico methods. The binding site conformations showed that the ligand ZnO binds with conserved binding site of penicillin-binding protein 6 (PBP 6) receptor. According to the interactions, ZnO-NPs form the same interaction pattern with respect to other reported ligands, hence it can play a significant role in the inhibition of PBP 6. This research also found that combining ZnO-NPs with Pseudomonas sp. (HY8N) was a novel and effective technique for treating pathogenic bacteria, including multidrug-resistant bacteria.     

Heparan sulfate/heparin oligosaccharides protect stromal cell-derived factor-1 (SDF-1)/CXCL12 against proteolysis induced by CD26/dipeptidyl peptidase IV

Stromal cell-derived factor-1 (SDF-1) is a CXC chemokine that is constitutively expressed in most tissues and displayed on the cell surface in association with heparan sulfate (HS). Its numerous biological effects are mediated by a specific G protein-coupled receptor, CXCR4. A number of cells inactivate SDF-1 by specific processing of the N-terminal domain of the chemokine. In particular, CD26/dipeptidyl peptidase IV (DPP IV), a serine protease that co-distributes with CXCR4 at the cell surface, mediates the selective removal of the N-terminal dipeptide of SDF-1. We report here that heparin and HS specifically prevent the processing of SDF-1 by DPP IV expressed by Caco-2 cells. The level of processing increases with the level of differentiation of these cells, which correlates with an increase of DPP IV activity. A mutant SDF-1 that does not interact with HS is readily cleaved by DPP IV, a process that is not inhibited by HS, demonstrating that a productive interaction between HS and SDF-1 is required for the protection to take place. Moreover, we found that protection depends on the degree of polymerization of the HS sulfated S-domains. Finally a structural model of SDF-1, in complex with HS oligosaccharides of defined length, rationalizes the experimental data. The mechanisms by which HS regulates SDF-1 may thus include, in addition to its ability to locally concentrate the chemokine at the cell surface, a control of selective protease cleavage events that directly affect the chemokine activity.