Devaraj S et al. (2018) Biodegradation "Anthelmintic efficacy of glycolipid biosurfactant produced by Pseudomonas plecoglossicida: an insight from mutant and transgenic forms of Caenorhabditis elegans."

Kathirvel P, Amirthalingam M, Devaraj S, Govindan S, Palanisamy S, Sabapathy PC

[Biodegradation, 2018]

The current research focuses on the production and characterization of glycolipid biosurfactant (GB) from Pseudomonas plecoglossicida and its anthelmintic activity against Caenorhabditis elegans. The GB was purified and characterized by Fourier Transform Infrared Spectroscopy (FTIR) and Gas Chromatography and Mass Spectrometry (GC-MS) analysis. Anthelmintic activity of GB was studied at six different pharmacological doses from 10 to 320g/mL on C. elegans. Exposure of different developmental stages (L1, L2, L3, L4 and adult) of C. elegans to the GB reduced the survivability of worms in a dose and time-dependent manner. Adult and L4 worms were least susceptible, while L1, L2 and L3 were more susceptible to GB when compared to the untreated control. An increased exposure period drastically reduced the survival rate of worms and reduction in LC₅₀ value. The GB significantly inhibited the development of C. elegans with an IC₅₀ value of 53.14g/mL and even reduced the adult body length and egg hatching. Fecundity rate of the worms treated with GB at 20, 40 and 80g/mL decreased from 261.90+/-3.21 to 239.70+/-5.58, 164.20+/-5.94 and 44.80+/-6.22 eggs per worm, respectively. Besides the toxicological effects, prolonged exposure to GB significantly decreased (p0.0001) the lifespan of wild type worms under standard laboratory conditions. Additionally, GB was found to be lethal towards ivermectin and albendazole resistant C. elegans strains. Overall, the data indicated that the GB extracted from P. plecoglossicida could be utilized for the control of non-susceptible and resistant gastrointestinal nematodes towards broad spectrum anthelmintic drugs, ivermectin and albendazole.

Lucio D et al. (2017) Eur J Pharm Biopharm "Optimization and evaluation of zein nanoparticles to improve the oral delivery of glibenclamide. In vivo study using C. elegans."

Irache JM, Martinez-Oharriz MC, Gonzalez-Navarro CJ, Aranaz P, Jaras G, Radulescu A, Lucio D

[Eur J Pharm Biopharm, 2017]

The aim of this work was to evaluate the capability of zein nanoparticles as oral carriers for glibenclamide (GB). Nanoparticles were prepared by a desolvation procedure in the presence of lysine as stabilizer. A central composite design was used to optimize this preparative process. Under the selected conditions, nanoparticles displayed a size of about 190 nm, a surface charge of -37 mV and a payload of 45 g GB/mg. Small-angle neutron scattering and X-ray diffraction techniques suggested an internal fractal-like structure, based on the repetition of spherical blocks of zein units (about 20 nm) grouped to form the nanoparticles. This structure, stabilized by lysine molecules located at the surface, would determine the release of GB (molecularly trapped into the nanoparticles) by a pure diffusion mechanism. Moreover, GB-loaded nanoparticles induced a significant hypolipidemic effect with a reduction of about 15% in the fat content of C. elegans worms. In addition, did not induce any significant modification in the lifespan of worms. In summary, the employment of zein nanoparticles as delivery systems of glibenclamide may be an interesting approach to develop new oral formulations of this antidiabetic drug.

Lucio D et al. (2017) Colloids Surf B Biointerfaces "Coencapsulation of cyclodextrins into poly(anhydride) nanoparticles to improve the oral administration of glibenclamide. A screening on C. elegans."

Martinez-Oharriz MC, Radulescu A, Navarro-Herrera D, Irache JM, Gonzalez-Navarro CJ, Lucio D, Gonzalez-Gaitano G

[Colloids Surf B Biointerfaces, 2017]

This work describes the feasibility of poly(anhydride) nanoparticles as carriers for the oral administration of glibenclamide (GB) as well as the in vivo evaluation of their hypolipidemic effect in a C. elegans model. For this purpose, and in order to increase the GB payload, the drug was encapsulated in nanoparticles in presence of cyclodextrins (either CD or HPCD). The optimized nanoparticles displayed a size of about 220nm and a negative zeta potential (-40mV), with a drug loading up to 52g/mg. Small-angle neutron scattering studies suggested an internal fractal-like structure, based on the repetition of spherical blocks of polymeric units (about 5nm) grouped to form the nanoparticle. X-ray diffraction study confirmed the absence of crystalline GB molecules due to its dispersion into the nanoparticles, either entrapped in the polymer chains and/or included into cyclodextrin cavities. GB-loaded nanoparticles induced a significant reduction in the fat content of C. elegans. This hypolipidemic effect was slightly higher for the nanoparticles prepared with coencapsulated HPCD (8.2%) than for those prepared with CD (7.9%) or in the absence of cyclodextrins (7.0%). In summary, the coencapsulation of cyclodextrins into poly(anhydride) nanoparticles could be an interesting strategy to develop new oral formulations of glibenclamide.

Lucio D et al. (2018) Int J Pharm "Cyclodextrin-grafted poly(anhydride) nanoparticles for oral glibenclamide administration. In vivo evaluation using C. elegans."

Irache JM, Aranaz P, Gu Z, Martinez-Oharriz MC, Lucio D, He Y, Vizmanos JL

[Int J Pharm, 2018]

The aim of this work was to prepare and evaluate cyclodextrins-modified poly(anhydride) nanoparticles to enhance the oral administration of glibenclamide. A conjugate polymer was synthesized by incorporating hydroxypropyl--cyclodextrin to the backbone of poly(methylvinyl ether-co-maleic anhydride) via Steglich reaction. The degree of substitution of anhydride rings by cyclodextrins molecules was calculated to be 4.9% using H-NMR spectroscopy. A central composite design of experiments was used to optimize the preparative process. Under the optimal conditions, nanoparticles displayed a size of about 170 nm, a surface charge of -47 mV and a drug loading of 69 g GB/mg. X-ray diffraction studies confirmed the loss of the crystalline structure of GB due to its dispersion into the nanoparticles, either included into cyclodextrin cavities or entrapped in the polymer chains. Glibenclamide was mainly release by Fickian-diffusion in simulated intestinal fluid. GB-loaded nanoparticles produced a hypolipidemic effect over C. elegans N2 wild-type and daf-2 mutant. The action mechanism included daf-2 and daf-28 genes, both implicated in the insulin signaling pathway of C. elegans. In summary, the covalent linkage of cyclodextrin to the poly(anhydride) backbone could be an interesting strategy to prepare nanoparticles for the oral administration of glibenclamide.

Zeev-Ben-Mordehai T et al. (2014) Structure "Extracellular vesicles: a platform for the structure determination of membrane proteins by Cryo-EM."

Grunewald K, Siebert CA, Whittle C, Zeev-Ben-Mordehai T, Vasishtan D

[Structure, 2014]

Membrane protein-enriched extracellular vesicles (MPEEVs) provide a platform for studying intact membrane proteins natively anchored with the correct topology in genuine biological membranes. This approach circumvents the need to conduct tedious detergent screens for solubilization, purification, and reconstitution required in classical membrane protein studies. We have applied this method to three integral type I membrane proteins, namely the Caenorhabditis elegans cell-cell fusion proteins AFF-1 and EFF-1 and the glycoprotein B (gB) from Herpes simplex virus type 1 (HSV1). Electron cryotomography followed by subvolume averaging allowed the 3D reconstruction of EFF-1 and HSV1 gB in the membrane as well as an analysis of the spatial distribution and interprotein interactions on the membrane. MPEEVs have many applications beyond structural/functional investigations, such as facilitating the raising of antibodies, for protein-protein interaction assays or for diagnostics use, as biomarkers, and possibly therapeutics.

Deorowicz S et al. (2013) BMC Bioinformatics "Disk-based k-mer counting on a PC."

Grabowski S, Debudaj-Grabysz A, Deorowicz S

[BMC Bioinformatics, 2013]

BACKGROUND: The k-mer counting problem, which is to build the histogram of occurrences of every k-symbol long substring in a given text, is important for many bioinformatics applications. They include developing de Bruijn graph genome assemblers, fast multiple sequence alignment and repeat detection. RESULTS: We propose a simple, yet efficient, parallel disk-based algorithm for counting k-mers. Experiments show that it usually offers the fastest solution to the considered problem, while demanding a relatively small amount of memory. In particular, it is capable of counting the statistics for short-read human genome data, in input gzipped FASTQ file, in less than 40 minutes on a PC with 16 GB of RAM and 6 CPU cores, and for long-read human genome data in less than 70 minutes. On a more powerful machine, using 32 GB of RAM and 32 CPU cores, the tasks are accomplished in less than half the time. No other algorithm for most tested settings of this problem and mammalian-size data can accomplish this task in comparable time. Our solution also belongs to memory-frugal ones; most competitive algorithms cannot efficiently work on a PC with 16 GB of memory for such massive data. CONCLUSIONS: By making use of cheap disk space and exploiting CPU and I/O parallelism we propose a very competitive k-mer counting procedure, called KMC. Our results suggest that judicious resource management may allow to solve at least some bioinformatics problems with massive data on a commodity personal computer.

Dakshinamoorthy G et al. (2012) PLoS One "Biochemical characterization and evaluation of a Brugia malayi small heat shock protein as a vaccine against lymphatic filariasis."

Dakshinamoorthy G, Samykutty AK, Kalyanasundaram R, Nutman T, Reddy MV, Munirathinam G, Shinde GB

[PLoS One, 2012]

Filarial nematodes enjoy one of the longest life spans of any human pathogen due to effective immune evasion strategies developed by the parasite. Among the various immune evasion strategies exhibited by the parasite, Interleukin 10 (IL-10) productions and IL-10 mediated immune suppression has significant negative impact on the host immune system. Recently, we identified a small heat shock protein expressed by Brugia malayi (BmHsp12.6) that can bind to soluble human IL-10 receptor alpha (IL-10R) and activate IL-10 mediated effects in cell lines. In this study we show that the IL-10R binding region of BmHsp12.6 is localized to its N-terminal region. This region has significant sequence similarity to the receptor binding region of human IL-10. In vitro studies confirm that the N-terminal region of BmHsp12.6 (N-BmHsp12.6) has IL-10 like activity and the region containing the alpha crystalline domain and C-terminus of BmHsp12.6 (BmHsp12.6c) has no IL-10 like activity. However, BmHsp12.6c contains B cell, T cell and CTL epitopes. Members of the sHSP families are excellent vaccine candidates. Evaluation of sera samples from putatively immune endemic normal (EN) subjects showed IgG1 and IgG3 antibodies against BmHsp12.6c and these antibodies were involved in the ADCC mediated protection. Subsequent vaccination trials with BmHsp12.6c in a mouse model using a heterologous prime boost approach showed that 83% protection can be achieved against B. malayi L3 challenge. Results presented in this study thus show that the N-BmHsp12.6 subunit of BmHsp12.6 has immunoregulatory function, whereas, the BmHsp12.6c subunit of BmHsp12.6 has significant vaccine potential.

Kornfeld K et al. (1995) Cell "The ksr-1 gene encodes a novel protein kinase involved in Ras-mediated signaling in C. elegans."

Horvitz HR, Hom DB, Kornfeld K

[Cell, 1995]

By screening for mutations that suppress the vulval defects caused by a constitutively active let gb ras gene, we identified six loss-of-function alleles of ksr-1, a novel C. elegans gene. Our genetic analysis showed ksr-1 positively mediates Ras signaling and functions downstream of or in parallel to let-60. In the absence of ksr-1 function, normal Ras signaling is impaired only slightly, suggesting ksr-1 may act to modulate, or in a branch that diverges from, the main signaling pathway. The predicted KSR-1 protein has a protein kinase domain and is most similar to a recently identified Drosophila protein involved in Ras signaling. We propose that the function of ksr-1 is evolutionarily conserved.

Patikoglou GA et al. (2000) East Coast Worm Meeting "G-protein beta-2 regulates the activity of two RGS proteins"

Chase DL, Koelle MR, Patikoglou GA

[East Coast Worm Meeting, 2000]

Heterotrimeric G proteins are signaling molecules that permit cells to adapt to changing extracellular conditions. Termination of the G protein signal is stimulated by a class of GTPase activating proteins known as regulators of G protein signaling or RGS proteins. We want to understand how RGS proteins select their proper G protein target in vivo. The Ga proteins GOA-1 and EGL-30, while normally expressed in the same cells of the C. elegans nervous system, have opposite effects on several behaviors including egg laying. Genetic analysis has identified two RGS proteins, EGL-10 and EAT-16, that negatively regulate GOA-1 and EGL-30, respectively. Such specificity of RGS proteins for Ga is not generally observed in vitro and encouraged further study of these signaling pathways. EGL-10 and EAT-16 share a region of similarity to Gg proteins known as the G-gamma-like or GGL domain. Mammalian GGL-containing RGS proteins have been shown to bind to the unique Gb protein, Gb5. To determine if the C. elegans ortholog of Gb5, GPB-2, plays a role in G protein signaling, or if GPB-2 contributes to signaling specificity, we isolated a deletion allele of gpb-2. This allele, missing the first two exons and 500 bases of promoter sequence (and thus a presumptive null allele), has little affect on egg-laying behavior in a wild-type background apparently because it has equal and opposing effects through EGL-10 and EAT-16. In mutants of either egl-10 or eat-16 in which we can isolate one RGS protein in the absence of the other, the gpb-2 deletion allele has dramatic effects. These effects are consistent with GPB-2 being required for full activity of these RGS proteins. If EGL-10 and EAT-16 physically interact with GPB-2 there may exist novel types of heterotrimeric complexes consisting of Ga, GPB-2 and RGS protein. Activation of these complexes by a receptor would then liberate GTP-bound Ga from the Gb-RGS complex. One might imagine that the proximity of the active Ga and Gb-RGS molecules could lead to a dramatically increased rate of GTP hydrolysis and hence termination of the Ga signal. We are currently using in vitro biochemical analyses of purified proteins to independently establish the interaction of GPB-2 with both EGL-10 and EAT-16. If these complexes form we will also determine the effect of GPB-2 binding on RGS activity and Ga subunit selection.

Rizk G et al. (2014) Bioinformatics "MindTheGap: integrated detection and assembly of short and long insertions."

Lemaitre C, Rizk G, Gouin A, Chikhi R

[Bioinformatics, 2014]

MOTIVATION: Insertions play an important role in genome evolution. However, such variants are difficult to detect from short-read sequencing data, especially when they exceed the paired-end insert size. Many approaches have been proposed to call short insertion variants based on paired-end mapping. However, there remains a lack of practical methods to detect and assemble long variants. RESULTS: We propose here an original method, called MindTheGap, for the integrated detection and assembly of insertion variants from re-sequencing data. Importantly, it is designed to call insertions of any size, whether they are novel or duplicated, homozygous or heterozygous in the donor genome. MindTheGap uses an efficient k-mer-based method to detect insertion sites in a reference genome, and subsequently assemble them from the donor reads. MindTheGap showed high recall and precision on simulated datasets of various genome complexities. When applied to real Caenorhabditis elegans and human NA12878 datasets, MindTheGap detected and correctly assembled insertions >1 kb, using at most 14 GB of memory.