Regulation of photosensation by hydrogen peroxide and antioxidants in C. elegans

The eyeless C. elegans exhibits robust phototaxis behavior in response to short-wavelength light, particularly UV light. C. elegans senses light through LITE-1, a unique photoreceptor protein that belongs to the invertebrate taste receptor family. However, it remains unclear how LITE-1 is regulated. Here, we performed a forward genetic screen for genes that when mutated suppress LITE-1 function. One group of lite-1 suppressors are the genes required for producing the two primary antioxidants thioredoxin and glutathione, suggesting that oxidization of LITE-1 inhibits its function. Indeed, the oxidant hydrogen peroxide (H₂O₂) suppresses phototaxis behavior and inhibits the photoresponse in photoreceptor neurons, whereas other sensory behaviors are relatively less vulnerable to H₂O₂. Conversely, antioxidants can rescue the phenotype of lite-1 suppressor mutants and promote the photoresponse. As UV light illumination generates H₂O₂, we propose that upon light activation of LITE-1, light-produced H₂O₂ then deactivates LITE-1 to terminate the photoresponse, while antioxidants may promote LITE-1’s recovery from its inactive state. Our studies provide a potential mechanism by which H₂O₂ and antioxidants act synergistically to regulate photosensation in C. elegans.     

Isolation and characterization of a monoclonal antibody containing an extra heavy-light chain Fab arm

Isolation and characterization of monoclonal antibody (mAb) variants to understand the impact of their structure on function is a typical activity during early-stage candidate selection that contributes to derisking clinical development. In particular, efforts are devoted to characterizing oligomeric variants, owing to their potential immunogenic nature. We report here a mAb variant consisting of a canonical mAb monomer associated in a non-covalent fashion with an antigen-binding fragment (Fab) arm amputated from its Fc domain. The truncated heavy chain is encoded in the cell line genome and is the likely product of a genomic recombination during cell line generation. The addition of the Fab arm results in severe loss of potency, indicating its interaction with the Fab domain of the monomer. The presence of such a variant can easily be mitigated by an adequate purification step.     

Detection of quantitative trait loci for reproduction and production traits in Large White and French Landrace pig populations (Open Access publication)

A genome-wide scan was performed in Large White and French Landrace pig populations in order to identify QTL affecting reproduction and production traits. The experiment was based on a granddaughter design, including five Large White and three French Landrace half-sib families identified in the French porcine national database. A total of 239 animals (166 sons and 73 daughters of the eight male founders) distributed in eight families were genotyped for 144 microsatellite markers. The design included 51 262 animals recorded for production traits, and 53 205 litter size records were considered. Three production and three reproduction traits were analysed: average backfat thickness (US_M) and live weight (LWGT) at the end of the on-farm test, age of candidates adjusted at 100 kg live weight, total number of piglets born per litter, and numbers of stillborn (STILLp) and born alive (LIVp) piglets per litter. Ten QTL with medium to large effects were detected at a chromosome-wide significance level of 5% affecting traits US_M (on SSC2, SSC3 and SSC17), LWGT (on SSC4), STILLp (on SSC6, SSC11 and SSC14) and LIVp (on SSC7, SSC16 and SSC18). The number of heterozygous male founders varied from 1 to 3 depending on the QTL.     

An assessment of the impact of Raman based glucose feedback control on CHO cell bioreactor process development

Process analytical technology (PAT) tools such as Raman Spectroscopy have become established tools for real time measurement of CHO cell bioreactor process variables and are aligned with the QbD approach to manufacturing. These tools can have a significant impact on process development if adopted early, creating an end-to-end PAT/QbD focused process. This study assessed the impact of Raman based feedback control on early and late phase development bioreactors by using a Raman based PLS model and PAT management system to control glucose in two CHO cell line bioreactor processes. The impact was then compared to bioreactor processes which used manual bolus fed methods for glucose feed delivery. Process improvements were observed in terms of overall bioreactor health, product output and product quality. Raman controlled batches for Cell Line 1 showed a reduction in glycation of 43.4% and 57.9%, respectively. Cell Line 2 batches with Raman based feedback control showed an improved growth profile with higher VCD and viability and a resulting 25% increase in overall product titer with an improved glycation profile. The results presented here demonstrate that Raman spectroscopy can be used in both early and late-stage process development and design for consistent and controlled glucose feed delivery.     

Three founding ancestral genomes involved in the origin of sugarcane

Background and AimsModern sugarcane cultivars (Saccharum spp.) are high polyploids, aneuploids (2n = ~12x = ~120) derived from interspecific hybridizations between the domesticated sweet species Saccharum officinarum and the wild species S. spontaneum.MethodsTo analyse the architecture and origin of such a complex genome, we analysed the sequences of all 12 hom(oe)ologous haplotypes (BAC clones) from two distinct genomic regions of a typical modern cultivar, as well as the corresponding sequence in Miscanthus sinense and Sorghum bicolor, and monitored their distribution among representatives of the Saccharum genus.Key ResultsThe diversity observed among haplotypes suggested the existence of three founding genomes (A, B, C) in modern cultivars, which diverged between 0.8 and 1.3 Mya. Two genomes (A, B) were contributed by S. officinarum; these were also found in its wild presumed ancestor S. robustum, and one genome (C) was contributed by S. spontaneum. These results suggest that S. officinarum and S. robustum are derived from interspecific hybridization between two unknown ancestors (A and B genomes). The A genome contributed most haplotypes (nine or ten) while the B and C genomes contributed one or two haplotypes in the regions analysed of this typical modern cultivar. Interspecific hybridizations likely involved accessions or gametes with distinct ploidy levels and/or were followed by a series of backcrosses with the A genome. The three founding genomes were found in all S. barberi, S. sinense and modern cultivars analysed. None of the analysed accessions contained only the A genome or the B genome, suggesting that representatives of these founding genomes remain to be discovered.ConclusionsThis evolutionary model, which combines interspecificity and high polyploidy, can explain the variable chromosome pairing affinity observed in Saccharum. It represents a major revision of the understanding of Saccharum diversity.     

Ligand-binding properties of the glutathione-binding protein of the mussel, Mytilus edulis

The glutathione-binding protein of Mytilus edulis possesses only one tryptophan per polypeptide. Quenching of intrinsic fluorescence due to this residue was studied in the presence of glutathione S-transferase ligands; hematin, bilirubin, biliverdin, bromosulphophthalein, 1-anilino-8-naphthalene sulphonate, 1,2-dichloro-4-nitrobenzene, ethacrynic acid and sodium deoxycholate as well as in the presence of triethyltin bromide. K_d values were estimated from these experiments and were found to be 38–310 μM. Based on non-denaturing electrophoresis, the protein was found to have a native molecular weight of 50 kDa. Taken together with previously reported subunit molecular weights in the region of 25 kDa, this indicates that this protein has a dimeric quaternary structure.     

Consistent interethnic differences in the distribution of clinically relevant endothelial nitric oxide synthase genetic polymorphisms.

A maldistribution of endothelial nitric oxide synthase (eNOS) genetic variants may explain differences in NO-mediated effects and response to drugs among black and white subjects. While interethnic differences in the distribution of eNOS genetic variants exist in the American population, it is not known whether such interethnic differences exist in other populations. To test this possibility, we examined the distribution of genetic variants of three clinically relevant eNOS polymorphisms (T(-786)C in the promoter, the VNTR in intron 4, and the Glu298Asp variant in exon 7) in 136 black and 154 white subjects from a Brazilian population, which is very heterogeneous. We also estimated the haplotype frequency and evaluated associations between these variants. The Asp298 variant was more common in whites (32.8%) than in blacks (15.1%) (P < 0.004). Similarly, the C(-786) variant was more common in whites (41.9%) than in blacks (19.5%) (P < 0.0004). However, the 4a variant was more common in blacks (32.0%) than in whites (17.9%) (P < 0.003). The most common predicted haplotype in both ethnic groups combined only wild-type variants. While the second most common haplotype in blacks includes the variant 4a and the wild-type variants for the remaining polymorphisms, the second most common haplotype in whites includes the variants Asp298 and C(-786) and the wild-type variant for polymorphism in intron 4. The marked interethnic differences that we found in Brazilians are very similar to those previously reported in Americans. These findings strongly suggest a consistent difference in the distribution of eNOS genetic variants in blacks compared with whites and indicate that the interethnic differences do not vary with geographic origin.