Identification of mitochondrial carriers in Saccharomyces cerevisiae by transport assay of reconstituted recombinant proteins

The inner membranes of mitochondria contain a family of carrier proteins that are responsible for the transport in and out of the mitochondrial matrix of substrates, products, co-factors and biosynthetic precursors that are essential for the function and activities of the organelle. This family of proteins is characterized by containing three tandem homologous sequence repeats of approximately 100 amino acids, each folded into two transmembrane alpha-helices linked by an extensive polar loop. Each repeat contains a characteristic conserved sequence. These features have been used to determine the extent of the family in genome sequences. The genome of Saccharomyces cerevisiae contains 34 members of the family. The identity of five of them was known before the determination of the genome sequence, but the functions of the remaining family members were not. This review describes how the functions of 15 of these previously unknown transport proteins have been determined by a strategy that consists of expressing the genes in Escherichia coli or Saccharomyces cerevisiae, reconstituting the gene products into liposomes and establishing their functions by transport assay. Genetic and biochemical evidence as well as phylogenetic considerations have guided the choice of substrates that were tested in the transport assays. The physiological roles of these carriers have been verified by genetic experiments. Various pieces of evidence point to the functions of six additional members of the family, but these proposals await confirmation by transport assay. The sequences of many of the newly identified yeast carriers have been used to characterize orthologs in other species, and in man five diseases are presently known to be caused by defects in specific mitochondrial carrier genes. The roles of eight yeast mitochondrial carriers remain to be established.     

Tyrosine phosphorylated proteins in different tissues during chick embryo development

A high affinity polyclonal antibody specific for phosphotyrosyl residues has been used in immunoblotting experiments to survey developing embryonic chicken tissues for the presence and characteristics of tyrosine phosphorylated proteins. Proteins phosphorylated on tyrosine were found to be present in all the embryonic tissues examined, including heart, thigh, gizzard, intestine, lung, liver, kidney, brain, and lens, from 7 to 21 d of development in ovo, but were greatly reduced or absent in the same tissues taken from adult chickens. A limited number of major tyrosine phosphorylated proteins were seen in all the tissues examined and they ranged in molecular mass from 35 to 220 kD. Most of the tissues contained proteins phosphorylated on tyrosine with apparent molecular masses of 120, 70, 60, and 35 kD, suggesting that the substrates of tyrosine protein kinases in different tissues may be related proteins. One-dimensional peptide mapping of the 120- and 70-kD protein bands indicated a close structural relationship among the phosphotyrosine-containing proteins of 120 kD, and similarly among those of 70 kD, from the different tissues.     

Soluble Guanylate Cyclase α1–Deficient Mice: A Novel Murine Model for Primary Open Angle Glaucoma

Primary open angle glaucoma (POAG) is a leading cause of blindness worldwide. The molecular signaling involved in the pathogenesis of POAG remains unknown. Here, we report that mice lacking the α₁ subunit of the nitric oxide receptor soluble guanylate cyclase represent a novel and translatable animal model of POAG, characterized by thinning of the retinal nerve fiber layer and loss of optic nerve axons in the context of an open iridocorneal angle. The optic neuropathy associated with soluble guanylate cyclase α₁–deficiency was accompanied by modestly increased intraocular pressure and retinal vascular dysfunction. Moreover, data from a candidate gene association study suggests that a variant in the locus containing the genes encoding for the α₁ and β₁ subunits of soluble guanylate cyclase is associated with POAG in patients presenting with initial paracentral vision loss, a disease subtype thought to be associated with vascular dysregulation. These findings provide new insights into the pathogenesis and genetics of POAG and suggest new therapeutic strategies for POAG.     

Cats and Dogs Down the Tree: The Tempo and Mode of Evolution in the Lower Carnassial of Fossil and Living Carnivora

Teeth are usually considered the primer to understand mammalian evolution. Their morphology is the result of the complex interplay between adaptation to food processing and somewhat fixed developmental programs. We analyse the morphometry of the first lower molar (m1 = lower carnassial) in 199 carnivore species (64 extant and 135 extinct) in order to clarify the tempo and mode of evolution of two different and possibly related traits, tooth size and tooth angular height. We applied a set of comparative methods and produced disparity through time plots by using four alternative phylogenetic trees. Three of them include fossil species, and were used in order to estimate what an effect the inclusion of fossil species has on the patterns we found for extant species. Our analyses on living species show that m1 size exhibits a weak phylogenetic signal, and that its variance occurs more within than among clades. The opposite applies to m1 angular height, which evolved in an adaptive radiation-like fashion, showing very strong phylogenetic signal and great among-clade variation. These results are thoroughly confirmed after the inclusion of fossil data. Major differences in m1 shape in Carnivora were clearly established early in their evolutionary history, probably as a result of rapid morphospace occupation after the clade radiation. Successive variation in tooth size allowed species to avoid potential overlap in feeding morphologies as a mechanism to limit interspecific competitive interaction within subclades.     

Entacapone Reduces Cortical Activation in Parkinson's Disease with Wearing-Off: A f-MRI Study

Background and Purpose

Wearing-off is one of the most frequent problems encountered by levodopa-treated patients. Entacapone, a peripheral inhibitor of catechol-O-methyltransferase (COMT), reduces this motor complication by prolonging the effect of levodopa. We sought to understand the impact of COMT-inhibition on movement execution in PD patients with wearing-off by comparing functional magnetic resonance imaging (f-MRI) activation patterns prior to and during entacapone treatment. Our hypothesis was to determine whether changes in cortical activation are associated to COMT-inhibitor treatment.

Methods

Nine levodopa-treated non-demented PD patients with wearing-off were prospectively studied in two f-MRI session, prior to and during entacapone treatment. A group of control subjects were also studied for comparison.

Results

The patients significantly improved under COMT-inhibitor treatment based on home diaries. F-MRI results showed that at baseline the patients presented a bilateral activation of the primary motor, controlateral premotor cortex and supplementary motor area, as well as ipsilateral cerebellum. During treatment with entacapone, PD patients showed reductions in the activations of these cortical areas and a decreased activation in the ipsilateral cerebellum.

Conclusions

Our preliminary findings indicate that f-MRI is able to detect cortical activation changes during long-term modulation of dopaminergic treatment in PD patients with wearing-off, and thus, this technique could be further investigated in advanced PD patients.     

Cardioprotective effect of ischemic preconditioning is preserved in food-restricted senescent rats

Ischemic preconditioning (PC) has been proposed as an endogenous form of protection against-ischemia reperfusion injury. We have shown that PC does not prevent postischemic dysfunction in the aging heart. This phenomenon could be due to the reduction of cardiac norepinephrine release, and it has also been previously demonstrated that age-related decrease of norepinephrine release from cardiac adrenergic nerves may be restored by caloric restriction. We investigated the effects on mechanical parameters of PC against 20 min of global ischemia followed by 40 min of reperfusion in isolated hearts from adult (6 mo) and "ad libitum"-fed and food-restricted senescent (24 mo) rats. Norepinephrine release in coronary effluent was determined by high-performance liquid chromatography. Final recovery of percent developed pressure was significantly improved after PC in adult hearts versus unconditioned controls (85.2 +/- 19% vs. 51.5 +/- 10%, P < 0.01). The effect of PC on developed pressure recovery was absent in ad libitum-fed rats, but it was restored in food-restricted senescent hearts (66.6 +/- 13% vs. 38.3 +/- 11%, P < 0.05). Accordingly, norepinephrine release significantly increased after PC in both adult and in food-restricted senescent hearts, and depletion of myocardial norepinephrine stores by reserpine abolished the PC effect in both adult and in food-restricted senescent hearts. We conclude that PC reduces postischemic dysfunction in the hearts from adult and food-restricted but not in ad libitum-fed senescent rats. Despite the possibility of multiple age-related mechanisms, the protection afforded by PC was correlated with increased norepinephrine release, and it was blocked by reserpine in both adult and food-restricted senescent hearts. Thus caloric restriction may restore PC in the aging heart probably via increased norepinephrine release.     

Combined control of Lasioderma serricorne (F.) and Ephestia elutella (Hbn.) in a tobacco processing facility by attracticide method

Results on application of attracticide method in the combined control of cigarette beetle, Lasioderma serricorne and tobacco moth, Ephestia elutella, infesting a tobacco processing facility are reported. Experimental devices with cross-panel stripes treated with microencapsulated permethrin at 6% (a.i.) and baited at same time with (4,6-dimethyl-7-hydroxynonan-3-one) serricornin and (Z,E)-9,12-tetradecadien-1 ol acetate (TDA) were used for 2 years (2017 and 2018) in an Italian tobacco processing facility. The experiment demonstrated that the presence of both serricornin and TDA dispensers on cross-panel striped devices had no affect on the numbers of L. serricorne and E. elutella male adults caught. After 2 years, the densities of adult males were clearly suppressed in room treated with attracticide compared to control room. Thus, the 2017–2018 experiment suggests that attracticide devices with two pheromone lures and microencapsulated permethrin at 6% (a.i.) can be effective suppression tools for managing combined pest populations of L. serricorne and E. elutella in tobacco facilities. The reduction of male infestation by L. serricorne ranged from 36% to 87%, and in E. elutella, the reduction of infestation ranged from 63% to 91%.     

Three paradigms of airway smooth muscle hyperresponsiveness in young guinea pigs

Evidence for contributions of airway smooth muscle (ASM) to the hyperresponsiveness of newborn and juvenile airways continues to accumulate. In our laboratory, 3 novel paradigms of hyperresponsiveness of newborn and young ASM have recently emerged using a guinea pig model of maturation in 3 age groups: 1 week (newborn), 3 weeks (juvenile), and 2-3 months (adult). The first paradigm includes evidence for a natural decline after newborn and juvenile life of the velocity of ASM shortening associated with a decrease in regulatory myosin light chain phosphorylation and a parallel decline in the content of myosin light chain kinase. Associated with the decrease in ASM shortening with age is an increase in the internal resistance to shortening. Dynamic stiffness is shown to relate inversely to the expression of myosin light chain kinase. This suggests that developmental changes in shortening relate inversely to the stiffness of the ASM early in shortening, suggesting a dynamic role for the cytoskeleton in facilitating and opposing ASM shortening. This relationship can be approximated as (dP/dt)max approximately (dP/dL)passive x (dL/dt)max (the maximal rate of increase of active stress generation approximately to the passive stiffness x the maximal shortening velocity). The second paradigm demonstrates that newborn ASM, unlike that in adults, does not relax during prolonged electric field stimulation. The impaired relaxation is related to changes in prostanoid synthesis and acetylcholinesterase function. The third paradigm demonstrates that, whereas oscillatory strain serves to transiently relax adult ASM, in newborns it induces (after the initial relaxation) a sustained potentiation of active stress. This is related to developmental changes in the prostanoid release. Together, these paradigms demonstrate that ASM contributes by multiple mechanisms to the natural hyperresponsiveness of newborn and juvenile airways. Future studies will elaborate the mechanisms and extend these paradigms to ASM hyperresponsiveness following sensitization in early life.     

Eph receptors in the adult brain

The Eph receptors are a large family of receptor tyrosine kinases with important roles in the establishment of neuronal and vascular networks during embryonic development. The functions of Eph receptors in the adult brain have only recently been investigated, and the results are forcing us to amend the conventional view that these molecules function predominantly in a developmental context. This review summarizes this rapidly expanding new area of research, which has shown that the Eph receptors regulate the structure and physiological function of excitatory synapses through multiple mechanisms, and might thus play a significant role in higher brain functions.     

Geographic differences in genetic susceptibility to IgA nephropathy: GWAS replication study and geospatial risk analysis

IgA nephropathy (IgAN), major cause of kidney failure worldwide, is common in Asians, moderately prevalent in Europeans, and rare in Africans. It is not known if these differences represent variation in genes, environment, or ascertainment. In a recent GWAS, we localized five IgAN susceptibility loci on Chr.6p21 (HLA-DQB1/DRB1, PSMB9/TAP1, and DPA1/DPB2 loci), Chr.1q32 (CFHR3/R1 locus), and Chr.22q12 (HORMAD2 locus). These IgAN loci are associated with risk of other immune-mediated disorders such as type I diabetes, multiple sclerosis, or inflammatory bowel disease. We tested association of these loci in eight new independent cohorts of Asian, European, and African-American ancestry (N = 4,789), followed by meta-analysis with risk-score modeling in 12 cohorts (N = 10,755) and geospatial analysis in 85 world populations. Four susceptibility loci robustly replicated and all five loci were genome-wide significant in the combined cohort (P = 5×10⁻³²–3×10⁻¹⁰), with heterogeneity detected only at the PSMB9/TAP1 locus (I² = 0.60). Conditional analyses identified two new independent risk alleles within the HLA-DQB1/DRB1 locus, defining multiple risk and protective haplotypes within this interval. We also detected a significant genetic interaction, whereby the odds ratio for the HORMAD2 protective allele was reversed in homozygotes for a CFHR3/R1 deletion (P = 2.5×10⁻⁴). A seven–SNP genetic risk score, which explained 4.7% of overall IgAN risk, increased sharply with Eastward and Northward distance from Africa (r = 0.30, P = 3×10⁻¹²⁸). This model paralleled the known East–West gradient in disease risk. Moreover, the prediction of a South–North axis was confirmed by registry data showing that the prevalence of IgAN–attributable kidney failure is increased in Northern Europe, similar to multiple sclerosis and type I diabetes. Variation at IgAN susceptibility loci correlates with differences in disease prevalence among world populations. These findings inform genetic, biological, and epidemiological investigations of IgAN and permit cross-comparison with other complex traits that share genetic risk loci and geographic patterns with IgAN.