Evolutionary interactions between N-linked glycosylation sites in the HIV-1 envelope

The addition of asparagine (N)-linked polysaccharide chains (i.e., glycans) to the gp120 and gp41 glycoproteins of human immunodeficiency virus type 1 (HIV-1) envelope is not only required for correct protein folding, but also may provide protection against neutralizing antibodies as a “glycan shield.” As a result, strong host-specific selection is frequently associated with codon positions where nonsynonymous substitutions can create or disrupt potential N-linked glycosylation sites (PNGSs). Moreover, empirical data suggest that the individual contribution of PNGSs to the neutralization sensitivity or infectivity of HIV-1 may be critically dependent on the presence or absence of other PNGSs in the envelope sequence. Here we evaluate how glycan–glycan interactions have shaped the evolution of HIV-1 envelope sequences by analyzing the distribution of PNGSs in a large-sequence alignment. Using a “covarion”-type phylogenetic model, we find that the rates at which individual PNGSs are gained or lost vary significantly over time, suggesting that the selective advantage of having a PNGS may depend on the presence or absence of other PNGSs in the sequence. Consequently, we identify specific interactions between PNGSs in the alignment using a new paired-character phylogenetic model of evolution, and a Bayesian graphical model. Despite the fundamental differences between these two methods, several interactions are jointly identified by both. Mapping these interactions onto a structural model of HIV-1 gp120 reveals that negative (exclusive) interactions occur significantly more often between colocalized glycans, while positive (inclusive) interactions are restricted to more distant glycans. Our results imply that the adaptive repertoire of alternative configurations in the HIV-1 glycan shield is limited by functional interactions between the N-linked glycans. This represents a potential vulnerability of rapidly evolving HIV-1 populations that may provide useful glycan-based targets for neutralizing antibodies.     

Mapping protease inhibitor resistance to human immunodeficiency virus type 1 sequence polymorphisms within patients

Resistance genotyping provides an important resource for the clinical management of patients infected with human immunodeficiency virus type 1 (HIV-1). However, resistance to protease (PR) inhibitors (PIs) is a complex phenotype shaped by interactions among nearly half of the residues in HIV-1 PR. Previous studies of the genetic basis of PI resistance focused on fixed substitutions among populations of HIV-1, i.e., host-specific adaptations. Consequently, they are susceptible to a high false discovery rate due to founder effects. Here, we employ sequencing “mixtures” (i.e., ambiguous base calls) as a site-specific marker of genetic variation within patients that is independent of the phylogeny. We demonstrate that the transient response to selection by PIs is manifested as an excess of nonsynonymous mixtures. Using a sample of 5,651 PR sequences isolated from both PI-naive and -treated patients, we analyze the joint distribution of mixtures and eight PIs as a Bayesian network, which distinguishes residue-residue interactions from direct associations with PIs. We find that selection for resistance is associated with the emergence of nonsynonymous mixtures in two distinct groups of codon sites clustered along the substrate cleft and distal regions of PR, respectively. Within-patient evolution at several positions is independent of PIs, including those formerly postulated to be involved in resistance. These positions are under strong positive selection in the PI-naive patient population, implying that other factors can produce spurious associations with resistance, e.g., mutational escape from the immune response.     

Altered mitochondrial oxidative phosphorylation capacity in horses suffering from polysaccharide storage myopathy

Polysaccharide storage myopathy (PSSM) is a widely described cause of exertional rhabdomyolysis in horses. Mitochondria play a central role in cellular energetics and are involved in human glycogen storage diseases but their role has been overlooked in equine PSSM. We hypothesized that the mitochondrial function is impaired in the myofibers of PSSM-affected horses. Nine horses with a history of recurrent exercise-associated rhabdomyolysis were tested for the glycogen synthase 1 gene (GYS1) mutation: 5 were tested positive (PSSM group) and 4 were tested negative (horses suffering from rhabdomyolysis of unknown origin, RUO group). Microbiopsies were collected from the gluteus medius (gm) and triceps brachii (tb) muscles of PSSM, RUO and healthy controls (HC) horses and used for histological analysis and for assessment of oxidative phosphorylation (OXPHOS) using high-resolution respirometry. The modification of mitochondrial respiration between HC, PSSM and RUO horses varied according to the muscle and to substrates feeding OXPHOS. In particular, compared to HC horses, the gm muscle of PSSM horses showed decreased OXPHOS- and electron transfer (ET)-capacities in presence of glutamate&malate&succinate. RUO horses showed a higher OXPHOS-capacity (with glutamate&malate) and ET-capacity (with glutamate&malate&succinate) in both muscles in comparison to the PSSM group. When expressed as ratios, our results highlighted a higher contribution of the NADH pathway (feeding electrons into Complex I) to maximal OXPHOS or ET-capacity in both rhabdomyolysis groups compared to the HC. Specific modifications in mitochondrial function might contribute to the pathogenesis of PSSM and of other types of exertional rhabdomyolyses.     

Transient EPR: using spin polarization in sequential radical pairs to study electron transfer in photosynthesis

The light induced electron transfer in photosynthesis generates a series of sequential spin polarized radical pairs, and transient electron paramagnetic resonance (TREPR) is ideally suited to study the lifetimes and physical and electronic structures of these radical pairs. In this article, the basic principles of TREPR are outlined with emphasis on the electron spin polarization (ESP) that develops during the electron transfer process. Examples of the analysis of TREPR data are given to illustrate the information that can be obtained. Recent applications of the technique to study the functionality of reaction centers, light-induced structural changes, and protein–cofactor interactions are reviewed.     

Novel 1,3-dipropyl-8-(1-heteroarylmethyl-1H-pyrazol-4-yl)-xanthine derivatives as high affinity and selective A2B adenosine receptor antagonists

A series of new 1,3-dipropyl-8-(1-heteroarylmethyl-1H-pyrazol-4-yl)-xanthine derivatives as A(2B)-AdoR antagonists have been synthesized and evaluated for their binding affinities for the A(2B), A(1), A(2A), and A(3)-AdoRs. 8-(1-((3-phenyl-1,2,4-oxadiazol-5-yl)methyl)-1H-pyrazol-4-yl)-1,3-dipropyl-1H-purine-2,6(3H,7H)-dione (4) displayed high affinity (K(i)=1 nM) and selectivity for the A(2B)-AdoR versus A(1), A(2A), and A(3)-AdoRs (A(1)/A(2B), A(2A)/A(2B), and A(3)/A(2B) selectivity ratios of 370, 1100, and 480, respectively). The synthesis and SAR of this novel class of compounds are presented herein.     

Comparison of loaded and unloaded jump squat training on strength/power performance in college football players

The purpose of this study was to explore the effects of 5 weeks of eccentrically loaded and unloaded jump squat training in experienced resistance-trained athletes during the strength/ power phase of a 15-week periodized off-season resistance training program. Forty-seven male college football players were randomly assigned to 1 of 3 groups. One group performed the jump squat exercise using both concentric and eccentric phases of contraction (CE; n = 15). A second group performed the jump squat exercise using the concentric phase only (n = 16), and a third group did not perform the jump squat exercise and served as control (CT; n = 16). No significant differences between the groups were seen in power, vertical jump height, 40-yd sprint speed and agility performance. In addition, no differences between the groups were seen in integrated electromyography activity during the jump squat exercise. Significant differences between the CE and CT groups were seen in Delta 1RM squat (65.8 and 27.5 kg, respectively) and Delta 1RM power clean (25.9 and 3.8 kg, respectively). No other between-group differences were observed. Results of this study provide evidence of the benefits of the jump squat exercise during a short-duration (5-week) training program for eliciting strength and power gains. In addition, the eccentric phase of this ballistic movement appears to have important implications for eliciting these strength gains in college football players during an off-season training program. Thus, coaches incorporating jump squats (using both concentric and eccentric phases of contraction) in the off-season training programs of their athletes can see significant performance improvements during a relatively short duration of training.     

Molecular distinction between true centric fission and pericentric duplication-fission

Centromere (centric) fission, also known as transverse or lateral centric misdivision, has been defined as the splitting of one functional centromere of a metacentric or submetacentric chromosome to produce two derivative centric chromosomes. It has been observed in a range of organisms and has been ascribed an important role in karyotype evolution; however, the underlying mechanisms remain unknown. We have investigated four cases of apparent centric fission in humans. Two cases show a missing chromosome 22 or 18 that is replaced by two centric ring products, a third case shows two chromosome-10-derived telocentric chromosomes, whereas a fourth case involves the formation of two chromosome-18-derived isochromosomes. In all four cases, results of gross cytogenetic and fluorescence in situ hybridisation analyses were consistent with a simple centric fission event. However, detailed molecular analyses provided evidence in support of centromere duplication as a predisposing mechanism for the observed chromosomal breakage in two of the cases. Results for the third case are consistent with direct centric fission not involving centromere pre-duplication as the likely mechanism. Insufficient material has precluded the further study of the fourth case. The data provide the first molecular evidence for centromere pre-duplication as a possible mechanism to explain the classically assumed simple centric fission events in clinical cytogenetics, karyotype evolution and speciation.     

Forensic genetic identification of abalone (<Emphasis Type=Italic>Haliotis</Emphasis> spp.) of the northeastern Pacific Ocean

International trade in abalone meats, exclusive of their shells, is a lucrative business based upon both legally and illegally harvested abalone from many jurisdictions. The inability to visually identify abalone meat to species in the absence of the shell impedes enforcement efforts to reduce the illegal exploitation of the world’s abalone resources. We describe species-specific DNA sequences for the gamete recognition proteins, sperm lysin and vitelline egg receptor for lysin, and their use in forensic species identification among abalone of the northeastern Pacific Ocean. Some commercially relevant abalone species from the Northern and Southern hemispheres can be differentiated on the basis of the length of the second intron of the sperm lysin gene. The seven North American species of abalone that occupy the waters of Mexico, the USA and Canada can be distinguished based on sequence differentiation in the first three repeats of the vitelline receptor gene.     

Structure-based design of 2,6,7-trisubstituted-7H-pyrrolo[2,3-d]pyrimidines as Aurora kinases inhibitors

This Letter reports the optimization of a pyrrolopyrimidine series as dual inhibitors of Aurora A/B kinases. This series derived from a pyrazolopyrimidine series previously reported as inhibitors of aurora kinases and CDKs. In an effort to improve the selectivity of this chemotype, we switched to the pyrrolopyrimidine core which allowed functionalization on C-2. In addition, the modeling rationale was based on superimposing the structures of Aurora-A kinase and CDK2 which revealed enough differences leading to a path for selectivity improvement. The synthesis of the new series of pyrrolopyrimidine analogs relied on the development of a different route for the two key intermediates 7 and 19 which led to analogs with both tunable activity against CDK1 and maintained cell potency.