p23H implicated as cis/trans regulators of AlaXp directed editing for mammalian cell homeostasis

doi:10.1073/pnas.1019400108

The toxicity of mistranslation of serine for alanine appears  to be universal and is prevented in part by the editing activities of alanyl-tRNA synthetases (AlaRSs), which remove serine from mischarged tRNA Ala. The problem of serine mistranslation is so acute that free standing, genome- encoded fragments of the editing domain of AlaRSs are found throughout evolution. These AlaXps are thought to provide functional redundancy of editing. Indeed, archaeal versions rescue the conditional lethality of bacterial cells harboring an editing inactive AlaRs. In mammals, AlaXps are encoded by a gene that fuses coding sequences of a homology of the HSP90 cochaperone p23 (p23 H) to those of AlaXp, to create p23H AlaXp. Not known is whether this fusion protein or various potential splice variants are expressed as editing proficient proteins in mammalian cells. Here we show that both p23H AlaXp and AlaXp alternative splice variants can be detected as proteins in mammalian cells. The variant that ablated p23H and encoded just AlaXp was active in vitro. In contrast, neither the p23H AlaXp fusion protein, nor the mixture of free p23H with AlaXp, was active. Further experiments in a mammalian cell based system showed that RNAi directed suppression of sequences encoding AlaXp led to a serine sensitive increase in the accumulation of misfolded proteins. The results demonstrate the dependence of mammalian cell homeostasis on AlaXp and implicate p23H as a cis and trans acting regulator of its activity.

Crystal structure of the synergistic antibiotic pair, lankamycin and lankacidin, in complex with the large ribosomal subunit

doi:10.1073/pnas.1019406108

The structures of the large ribosomal subunit of Deinococcus radiodurans (D50S) in complex with the antibiotic lankamycin (3.2 Angastrom) and a double antibiotic complex of lankamycin and lankacidin C (3.45 Angastrom) have been determined, in continuation of previous crystallographic studies on lankacidin-D50S complex. These two drugs have been previously reported to inhibit ribosomal function with mild synergistic effect. Lankamycin, a member of the macrolide family, binds in a similar manner to erythromycin. However, when in complex with lankacidin, lankamycin is loacted so that it can form interactions with lankacidin in the adjacent ribosomal binding site. When compared to the well documented synergistic antibiotics, Streptogramins A and B, the pair of lankacidin and lankamycin bind in similar sites, the peptidyl transferase center and nascent peptide exit tunnel, respectively. Herein, we discuss the structural basis for antibiotic synergism and highlight the key factors involved in ribosomal inhibition.

Crystal structures of two active proliferating cell nuclear antigens (PCNAs) encoded by Thermococcus kodakaraensis

doi:10.1073/pnas.1019179108

Proliferating cell nuclear antigen (PCNA) is a ring shaped protein that encircles duplex DNA and plays an essential role in many DNA metabolic processes in archaea and eukarya. The eukaryotic and euryarchaea genomes contain a single gene encoding for PCNA. Interestingly, the genome of the euryarchaeon Thermococcus kodakaraensis contains two PCNA encoding genes (TK0535 and TK0582), making it unique among the euryarchaea kingdom. It is shown here that the two T. kodakaraensis PCNA proteins support processive DNA synthesis by the polymerase. Both proteins from trimeric structures with characterisitcs similar to those of other archeal and eukaryal PCNA proteins. One of the notable differences between the TK0535 and TK0582 rings is that the interfaces are different, resulting in different stabilities for the two trimers. The possible implications of these observations for PCNA functions are discussed.

pH induced metal ligand cross links inspired by mussel yield self healing polymer networks with near covalent elastic moduli

doi:10.1073/pnas.1015862108

Growing evidence supports a critical role of metal ligand coordination in many attributes of biological materials including adhesion, self assembly, toughness, and hardness without mineralization. Coordination between Fe and catechol ligands has recently been correlated to the hardness and high extensibility of the cuticle mussel byssal threads and proposed to endow self healing properties. Inspired by the pH jump experienced by proteins during maturation of a mussel byssus secretion, we have developed a simple method to control catechol Fe 3+ interpolymer cross linking via pH. The resonance Raman signature of catechol-Fe3+ cross linked polymer gels at high pH was similar to that from native mussel thread cuticle and the gels displayed elastic moduli that approach covalently cross linked gels as well as self healing properties.

MIcroworm optode sensors limit particle diffusion to enable in vivo measurements

doi:10.1073/pnas.1015544108

There have been a variety of nanoparticles created for in vivo uses ranging from gene and drug delivery to tumor imaging and physiological monitoring. The use of nanoparticles to measure physiological conditions while being fluorescently addressed through the skin provides an ideal method toward minimally invasive health monitoring. Here we create unique particles that have all the necessary physical characteristics to serve as in vivo reporters, but with minimized diffusion from the point of injection. These particles, called microworms, have a cylindrical shape coated with a biocompatible porous membrane that possesses a large surface area to volume ration while maintaining a large hydrodynamic radius. We use these microworms to create fluorescent sodium sensors for use as in vivo sodium conceration detectors after subcutaneous injection. However, the microworm concept has the potential to extend to the immobilization of other types of polymers for continuous physiological detection or delivery of molecules.

Substantial expression of mature elastin in arterial constructs

, doi:10.1073/pnas.1017834108

Mature elastin synthesis is a key challenge in arterial tissue engineering. Most engineered vessels lack elastic fibers in the medial layer and those present are poorly organized. The objective of this study is to increase mature elastin synthesis in small diameter arterial constructs. Adult primary baboon smooth muscle cells (SMC) were seeded in the lumen of porous tubular scaffolds fabricated from a biodegradable elastomer, poly (glycerol sebacate) (PGS) and cultured in a pulsatile flow bioreactor for 3 weeks. We tested the effect of pore sizes on construct properties by histological, biochemical, and mechanical evaluations. Histological analysis revealed circumferentially organized extracellular matrix proteins including elastin and the presence of multilayered SMCs expressing calponin and alpha smooth muscle actin. Biochemical analysis demonstrated that the constructs contained mature elastin equivalent to 19% of the native arteries. Mechanical tests indicated that the constructs could withstand up to 200 mmHg burst pressure and exhibited compliance comparable to native arteries. These results show that nontransfected cells in PGS scaffolds in unsupplemented medium produced a substantial  amount of mature elastin within 3 week and the elastic fibers has similar orientation as those in native arteries. The 25-32 micro meter pore size supported cell organization and elastin synthesis more than larger pore sizes. To our knowledge, there was no prior report of the synthesis of mature and organized elastin in arterial constructs without exogenous factors or viral transduction.

An extinct monkey from Haiti and the origins of the Greater Antillean primates

doi:10.1073/pnas.1009161108

A new extinct Late Quaternary platyrrhine from Haiti, Insulacebus toussaintiana, is described here from the most complete Caribbean subfossil primate dentition yet  recorded, demonstrating the likely coexistence of two primate species on Hispaniola. Like other Caribbean platyrrhines, l.toussaintiana exhibits primitive features resembling early Middle Miocene Patagonian fossils, reflecting an early derivation before the Amazonian community of modern New World anthropoids was configured. This , in combination with the young age of the fossils, provides a unique opportunity to examine a different parallel radiation of platyrrhines that survived into modern times, but is only distantly related to extant mainland forms. Their ecological novelty is indicated by their unique dental proportions, and by their relatively large estimated body weights, possibly and island effect, which places the group in a size class not exploited by mainland South American monkeys. Several features tie the new species to the extinct Jamaican monkey Xenothrix mcgregori, perhaps providing additional evidence for an inter Antillean clade.

The vulnerable developing brain

Dino A. Giussani, PNAS, doi:10.1073/pnas.1019726108

During healthy pregnancy, appropriate levels of fetal nutrition and oxygenation are indispensable for the optimal growth and development of the fetal organs. In pregnancy complicated by decreased nutrient and oxygen delivery to the fetus, current dogma is that the growth and development of key organs, such as the brain, are “spared” at the expense of nonessential organs served by peripheral vascular beds, for instance the kidneys, pancreas and skeletal muscle. The article by Antonow Schlorke et al. in PNAS challenges this concept and shows that the developing brain is more vulnerable than previously thought, even to moderate reductions in maternal nutrition during early pregnancy. The study is particularly relevant because it was undertaken in nonhuman primates rather than in rodents. In contrast to primates, including the human in rodent species many stages of brain development continue to occur after birth.

The concept of fetal brain sparing in adverse pregnancy is based on overwhelming evidence obtained from epidemiological studies in humans and experimental studies in animals, which have revealed that malnourished or hypoxic pregnancy yields disproportionately growth retarded offspring. The “thrifty phenotype” baby has an increased head diameter to body length or weight ration, with a reduced ponderal index, being thin of its length. In experimental animals, asymmetric intrauterine growth restriction is also represented by increased fetal brain weight relative to body weight or an increase in the………….

The significance of self control

Angela L Duckworth, PNAS, doi:10.1073/pnas.1019725108

Self control is among the most widely studied constructs in the social sciences. For instance, more than 3% of peer reviewed psychology articles in the past year were referenced by the key word “self control” or closely related terms. The report by Moffitt et al. in PNAS substantially advances this growing literature by demonstrating robust predictive associations between childhood self control and a wide range of consequential life outcomes in a large, nationally representative sample of New Zealanders.

Monikers for self control vary widely and include delay of gratification, effortful control, willpower, executive control, time preference, self discipline, self regulation and ego strength. Moffitt et al. use the term self control synonymously with conscientiousness, a large class of personality traits that includes responsibility, industriouness and orderliness. The common thread running through diverse conceptualizations of self control is the idea of effortful regulation of the self by the self. Self controlled individuals are more adept than their impulsive counterparts at regulating their behavioral, emotional and attentional impulses to achieve long term goals.

The notion of effortful self governance presumes as internal conflict between mutually exclusive responses. One cannot, alas, have one’s cake later and eat is now too. Critical to situations that call upon self control is that one response….

Toll in the vessel wall for better or worse?

Goran K. Hansson and Anna M. Lundberg, PNAS,doi:10.1073/pnas.1019722108

Pattern recognition receptors detect danger signals such as pathogen associated molecular patterns. Among such detectors, the membrane bound receptors of the Toll like receptor (TLR) family can detect all kinds of pathogens and play an important role in host defense. Not surprisingly, TLRs are also involved in chronic inflammatory diseases such as arthritis, colitis and atherosclerosis.

Atherosclerosis, the cause of myocardial infraction and stroke is a chronic inflammatory disease elicited by cholesterol accumulation in the artery wall. Innate as well as adaptive immunity contributes to inflammation in atherosclerosis and a spectrum of TLRs are expressed in the human atherosclerotic plaque. Experiments in gene targeted models show that certain TLRs impact on atherosclerosis in hypercholesterolemic mice. Specifically, TLR2 and TLR4 ligations promote disease development whereas ablation of these receptors reduces lesion size. Thus, one could envisage that endotoxins released during infections with Gram negative bacteria would ligate TLR4 in lesions, leading to accelerated atherosclerosis. Similarly, peptidoglycans of Gram positive bacteria could promote atherosclerosis by ligating TLR2. These findings are of obvious interest, because certain bacterial infections have been suggested to promote cardiovascular disease.

Viruses can be recognized by TLR3, which ligates dsRNA, TLR7 and TLR8, which bind certain ssRNA species and TLR9 which is stimulated by unmethylated CpG DNA motifs. The effects of such signals are also of interest for cardiovascular disease, because certain viral infections have been associated…..

Reverse and flick: Hybrid locomotion in bacteria

Roman Stocker, PNAS, doi:10.1073/pnas.1019199108

Many bacteria are motile. They use one or more helical flagella as propellers, rotating them like the corkscrew on a wine bottle opener. Despite the limited morphological repertoire of the propulsive system, radically different movement strategies have evolved, likely reflecting the diversity of physicochemical conditions among bacterial habitats. In PNAS, Xie et al. report on a newly discovered mechanism for turning used by Vibrio alginolyticus, an inhabitant of the coastal ocean: These monotrichous (single haired) bacteria change direction with a “flick” of their flagellum. Intriguingly, Xie et al. show that less can be more when it comes to bacterial flagella: With its single flagellum, V.alginolyticus outperforms the multiflagellated Esherichia coli in climbing nutrient gradients (chemotaxis), suggesting that the flick is part of an advanced chemotaxis system.

Out understanding of bacterial locomotion has long been driven and biased by the wealth of knowledge on E.coli, commonly found in animal intestines. E.coli is peritrichous, having four to eight flagella emerging from random points on its 2x1 micro meter hotdog shaped body. Each flagellum is powered by a reversible rotary motor. When all motors spin counterclockwise, hydrodynamic interactions cause the flagella to form a bundle that propels E.coli forward in a nearly straight “run” at ~30 micro meter/second. When one or more motors switch direction, the bundle comes apart, causing a change in direction (tumble) before a new run begins. The angle of reorientation during a tumble is nearly random, with the new run only slightly biased in the direction of the old one. This “run and tumble” movement pattern is common among peritrichous bacteria, including the pathogen Salmonella typhimurium and soil dwelling Bacillus subtilis.

Other bacteria like V.alginolyticus, have single flagellum and thus lack E.coli’s tumbling mechanism….

Leading the dog of selection by its mutational nose

Daniel S. Fisher, PNAS, doi:10.1073/pnas.1100339108

There are two simple caricatures of evolutionary dynamics: the phenotypic caricature focuses on continuous and predictable selection on variability of quantitative traits, whereas the genotypic caricature focuses on discrete, stochastic mutations. Although the apparent contradictions between these pictures were reconciled long ago, our quantitative understanding of the interplay between them is still surprisingly primitive. Indeed, even the simplest models of the dynamics of large asexual populations in which many alleles and many new mutations contribute to the evolving fitness have resisted solution. The PNAS paper by Hallatschek is a substantial advance in the development of the mathematical methods needed to analyze these and more complex models.

What is the source of the basic difficulty? The Fundamental Theorem of Natural Selection describes, quantitatively, how a diverse population subject to selective pressure evolves. It states that the rate of change of the average of a quantitative trait is proportional to the hereditable part of the variance of that trait times the strength of selection for it. But what determines the variance of a trait that is under selection? Initially, the frequencies of the various alleles that contribute to it: if there are many of these, the sum of their positive and negative contributions will give rise to a roughly normal bell shaped distribution of the fitness. However, as selection proceeds, some of the alleles will rise to fixation and other die out. Thus, under steady selection, after a modest time, only the beneficial alleles will remain and there will be no variance at all: the increase in fitness then comes to a grinding halt. But of course there can be new beneficial mutations. If only one such mutation arose at a time, it would sweep through the population before another occurred: this is the simple population genetics caricature of discrete mutation selection events…

A Study Asks, Could E-Cigarettes Really Help Smokers Quit?- TIME Healthland

Electronic Cigarettes, the smokeless battery operated nicotine-delivery devices that look like real cigarettes, are becoming increasingly available online, with manufacturers marketing them largely to people who are trying to quit smoking? Question is : do they work?

At least one previous study said no, finding that e-cigarettes don’t deliver much nicotine and don’t reduce smokers’ cravings.

Now two new studies of e-cigs, published recently in the American Journal of Preventive Medicine, attempt to shed a little more light on the issue. The first study compared internet searchers for, and purchases of, e-cigarettes and other quit smoking products like nicotine gum from Jan 2008 to Sept. 2010 in the US, Britain, Canada and Australia. The authors didn’t look at the effectiveness of e-cigs, but did find that they were the most popular smoking alternatives or cessation products on the online market, according to a statement.

In another study, researchers at Boston University sent online surveys to 5,000 people who had bought Blu e-cigarettes for the first time during a two week period in 2009. The number of respondents was small just 222. They were mostly male and long time smokers who had tried and failed to quit several times before. Among them, 67% said they had cut down on the number of cigarettes they smoked six months after buying  Blu, and 31% had quit at the six- month mark; 49% also said they’d stopped smoking for some unspecified amount of time.

Of course, it’s entirely possible that smokers who were more successful at cutting down or quitting were more likely to respond to the survey, which would have biased the results.

“Neither of these two studies provides scientific evidence that e-cigarettes are effective in helping people to quit,” said John Pierce, a professor of cancer prevention at the Moores Cancer Center at the University of California, San Diego, in a statement. “It’s not clear to me that e-cigarettes aren’t harmful in some way. It’s not clear to the FDA, either.”

In Sept, 2010, the FDA announced it would start regulating e-cigarettes as drug delivery devices and cited five distributors for “violations of good manufacturing practices, making unsubstantiated drug claims and using the devices as delivery mechanisms for active pharmaceutical ingredients,” according to an agency press release.

In January, the FDA tried unsuccessfully to block e-cigarette importation. Several US states are now moving to ban or restrict their use.

http://healthland.time.com/2011/02/10/a-study-asks-could-e-cigarettes-really-help-smokers-quit/

Dog Sniffing Out Cancer May Lead to Early Detection Test

 The latest study demonstrating that dogs can sniff out cancer has confirmed the notion that a specific cancer smell does exist, and has added fuel to the idea of developing a test based on odor.

Previous studies have reported on dogs that can detect lung and breast cancer from breath samples, and there has been anecdotal evidence suggesting that dogs can detect melanoma, bladder, and ovarian cancers.

In this latest study, published online January 31 in Gut, a Labrador retriever was trained over several months to sniff out colorectal cancer in breath and watery stool samples.

Hideto Sonoda, MD, and colleagues from Kyushu University in Fukuoka, Japan, report that this dog was then tested with samples obtained from colorectal cancer patients and from volunteers, some of whom had gastrointestinal problems such as ulcers and inflammatory bowel disease.

The dog correctly identified cancer in 33 of 36 breath tests and in 37 of 38 stool tests. This equates to 95% accuracy overall for the breath test and 98% accuracy overall for the stool test, the researchers report.

The highest detection rates were among samples taken from patients with early-stage cancer, they add. Samples taken from smokers and from people with other gastrointestinal diseases, which might be expected to mask or interfere with cancer odors, did not appear to confuse the dog.

"This study shows that a specific cancer scent does indeed exist," the researchers conclude.

They are not suggesting using dogs in clinical practice, however. They point out that training the dog was expensive and time-consuming, and that ability and concentration vary between individual dogs and even the same dog on different days. The dog's concentration tends to decrease during the hot summer season; hence, they conducted their test between November and early June.

What they do propose is that this research could be used to develop cancer detection tests based on "odor materials."

This would involve identifying the cancer-specific volatile organic compounds (VOC) that are being detected by dogs using chemical analysis, and then developing an early cancer detection sensor that would substitute for the dog, they explain.

There has already been some work conducted on VOC in exhaled breath (using gas chromatography and mass spectroscopy) for the early detection of breast and lung cancer, they note, although they add that this work is still preliminary.

"We hope that the results of the present study will provide encouragement for the development of cancer detection and solving the biological character of cancer using odor material," Dr. Sonoda and colleagues conclude.

The researchers have disclosed no relevant financial relationships.

Gut. Published online January 31, 2011.

Mitochondrial Dysfunction Linked to Autism

Mitochondrial dysfunction (MD) is more common in children with autism and autism spectrum disorder (ASD) than the general population, a comprehensive systematic review and meta-analysis of relevant research confirms.

Mitochondrial dysfunction "may play a significant role in contributing to the symptoms of autism and is generally underrecognized in these children," Daniel A. Rossignol, MD, of the International Child Development Resource Center, Melbourne, Florida, told Medscape Medical News.

Testing for mitochondrial dysfunction is available, and early treatment might lead to better long-term developmental outcomes," said Dr. Rossignol, who coauthored the review with Richard E. Frye, MD, PhD, of the University of Texas in Houston.

The report was published online January 25 in Molecular Psychiatry.

Commenting on the study Cecilia Giulivi, PhD, professor of biochemistry and metabolic regulation, at the University of California, Davis, who was not involved in the analysis, said, "At this point, it looks like there is a higher incidence of mitochondrial disease in autism, much higher than we suspected."

She noted, however, that testing for MD "is not a trivial task [and] we need more research to come up with a consensus of diagnostic tests to run. In addition, maybe other metabolic syndromes should be looked into," Dr. Giulivi said.

The primary objectives of the analysis were to identify features of MD in the general population of children with ASD and compare characteristics of MD in children with ASD and concomitant significant and severe MD with that of ASD children without MD and non-ASD children with MD.

They included 68 relevant published articles in a qualitative synthesis, including 18 studies with a total of 112 children with ASD and MD.

Genetics Not the Culprit

The results showed the prevalence of MD in the general population of children with ASD is approximately 5% (95% confidence interval [CI], 3.2% – 6.9%), which is 500% higher than the general population prevalence of 0.01%. For a variety of reasons, "this 5% value is most likely an underestimation," Dr. Rossignol said.

It also appears that one-third or more of children with autism may have some type of dysfunction in their mitochondria. On the basis of laboratory testing, the prevalence of abnormal biomarker values of MD, including lactate, pyruvate, carnitine, and ubiquinone, was high in children with ASD, much higher than the prevalence of MD. Some of these markers correlated with the severity of ASD.

Most of the 112 children with ASD and MD (79%) had no an identifiable genetic abnormality that could account for the MD.

"The mitochondrial dysfunction and disease reported in autism are related to a genetic abnormality in only 1 out of 5 children; meaning that a majority of these children have something else contributing to this dysfunction, which might include multiple environmental factors, such as toxins, oxidative stress, inflammation, and decreased levels of antioxidants," said Dr. Rossignol.

"Clearly, mitochondrial function is a ripe area of research when investigating the biological mechanism(s) of action of environmental toxicant exposures and indigenous abnormalities associated with ASD," the study authors write.

Loss of Social Skills

Children with ASD and MD had some distinct characteristics compared with the general population of children with ASD. In 12 studies, "children with autism and mitochondrial problems were more likely to lose acquired skills compared to children with autism in general," said Dr. Rossignol. However, it was not clear whether MD contributed to or caused the reported regression.

In addition to a higher prevalence of developmental regression (52%), seizures (41%), motor delay (51%), and gastrointestinal abnormalities (74%), such as reflux and constipation, also appear to be significantly more common in children with ASD and MD relative to children with just ASD.

Currently, "testing for mitochondrial problems in children with autism is rarely done, and we feel that testing should be routine, especially in children with regression or loss of skills," said Dr. Rossignol. "This is important because early recognition of mitochondrial problems in autism might lead to better outcomes in children with autism."

Dr. Rossignol and Dr. Frye note in their report that published studies looking at treatment for ASD and MD are limited. However, some studies have suggested that treatment with mitochondrial cofactor supplementation, including antioxidants, carnitine, coenzyme Q10, and B vitamins, may improve mitochondrial function and behavior in some children with ASD.

"A therapeutic trial of mitochondrial cofactors and antioxidants may be reasonable in children with ASD/MD," the study authors conclude. Carnitine, they say, may be particularly helpful in children with ASD because carnitine deficiency has been implicated in ASD, and some studies have reported improvements with the use of carnitine in ASD.

The researchers emphasize, however, that systematic studies documenting the efficacy of this and other potential treatments for MD in children with ASD are generally lacking.

Need for Longitudinal Studies

A small study by Dr. Giulivi and colleagues showed that impaired mitochondrial function and mitochondrial DNA abnormalities, including overreplication and deletions, were more common in children with autism than in typically developing children (Giulivi et al. JAMA. 2010;304:2389-2396).

"According to our study and those of others, there is evidence for MD in typical autism and ASD," Dr. Giulivi said. "However, I don't think that we know the role of this MD, meaning if it is related to the etiology (cause) or consequence of another underlying altered pathway.

"In any case [cause or consequence], a decline in mitochondrial function [below the threshold for a given tissue] and especially in highly aerobic tissues, such as brain, will have an impact on cellular energy and, in addition, maybe on other mitochondria-dependent pathways, such as heme metabolism," Dr. Giulivi noted.

She also made the point that most studies to date have been cross-sectional and therefore, "unless it is a longitudinal one, we cannot assess the role of MD in ASD and autism."

Dr. Rossignol and Dr. Frye point out that many of the studies they reviewed had a number of limitations, including "small sample sizes, referral or publication biases, and variability in protocols for selecting children for MD workup, collecting mitochondrial biomarkers, and defining MD." Only 39% of the ASD/MD studies reviewed noted the criterion used for diagnosing MD.

They agree with Dr. Giulivi that further studies are needed to further define the role of MD in ASD.

This research was funded in part by the Autism Research Institute and the Jane Botsford Johnson Foundation. Dr. Rossignol has 2 children with ASD and is a practicing primary care physician who treats ASD children with standard and integrative treatments. Dr. Frye provides expert testimony for children with MDs who may have been injured from vaccines. Funds from such testimony are used to support research on the biological basis of neurodevelopmental disorders. Dr. Giulivi has disclosed no relevant financial relationships.

Mol Psychiatry. Published online January 25, 2011.