The serine protease Omi/HtrA2 regulates apoptosis by binding XIAP through a reaper-like motif

The inhibitor-of-apoptosis proteins (IAPs) play a critical role in the regulation of apoptosis by binding and inhibiting caspases. Reaper family proteins and Smac/DIABLO use a conserved amino-terminal sequence to bind to IAPs in flies and mammals, respectively, blocking their ability to inhibit caspases and thus promoting apoptosis. Here we have identified the serine protease Omi/HtrA2 as a second mammalian XIAP-binding protein with a Reaper-like motif. This protease autoprocesses to form a protein with amino-terminal homology to Smac/DIABLO and Reaper family proteins. Full-length Omi/HtrA2 is localized to mitochondria but fails to interact with XIAP. Mitochondria also contain processed Omi/HtrA2, which, following apoptotic insult, translocates to the cytosol, where it interacts with XIAP. Overexpression of Omi/HtrA2 sensitizes cells to apoptosis, and its removal by RNA interference reduces cell death. Omi/HtrA2 thus extends the set of mammalian proteins with Reaper-like function that are released from the mitochondria during apoptosis.     

Coordinate repression of cholesterol biosynthesis and cytoplasmic 3-hydroxy-3-methylglutaryl coenzyme A synthase by glucocorticoids in HeLa cells.

The stability constants for the calcium and magnesium complexes of rhodanese are >10⁵m^?1 at both high and low substrate concentrations. The stoichiometry of alkaline earth metal ion binding totals close to 1 per 18,500 molecular weight. The usual assay reagents contain sufficient amounts of these metal ions to maintain added enzyme in its metal-complexed form. When reaction mixtures are treated with oxalate to remove calcium ions, inhibition of rhodanese activity is virtually complete under circumstances such that the contribution of magnesium ion is low.Zinc and a number of transition metal ions are inhibitors of rhodanese activity. Studies of the concentration dependence of these effects with zinc, copper, and nickel showed that: 1) Some cyanide complexes of these metals are competitive with the donor substrate, thiosulfate ion. The binding of the copper and zinc complexes is mutually competitive. 2) Another cyanide species of copper appears to combine with the free enzyme to form a functionally active complex. 3) The zinc cyanide species with a net positive charge is an inhibitor competitive with the acceptor substrate, cyanide ion.All of these observations are consistent with a model in which metal ions serve as the electrophilic site of rhodanese.     

Longitudinal Analysis of CCR5 and CXCR4 Usage in a Cohort of Antiretroviral Therapy-Na?ve Subjects with Progressive HIV-1 Subtype C Infection

HIV-1 subtype C (C-HIV) is responsible for most HIV-1 cases worldwide. Although the pathogenesis of C-HIV is thought to predominantly involve CCR5-restricted (R5) strains, we do not have a firm understanding of how frequently CXCR4-using (X4 and R5X4) variants emerge in subjects with progressive C-HIV infection. Nor do we completely understand the molecular determinants of coreceptor switching by C-HIV variants. Here, we characterized a panel of HIV-1 envelope glycoproteins (Envs) (n = 300) cloned sequentially from plasma of 21 antiretroviral therapy (ART)-naïve subjects who experienced progression from chronic to advanced stages of C-HIV infection, and show that CXCR4-using C-HIV variants emerged in only one individual. Mutagenesis studies and structural models suggest that the evolution of R5 to X4 variants in this subject principally involved acquisition of an “Ile-Gly” insertion in the gp120 V3 loop and replacement of the V3 “Gly-Pro-Gly” crown with a “Gly-Arg-Gly” motif, but that the accumulation of additional gp120 “scaffold” mutations was required for these V3 loop changes to confer functional effects. In this context, either of the V3 loop changes could confer possible transitional R5X4 phenotypes, but when present together they completely abolished CCR5 usage and conferred the X4 phenotype. Our results show that the emergence of CXCR4-using strains is rare in this cohort of untreated individuals with advanced C-HIV infection. In the subject where X4 variants did emerge, alterations in the gp120 V3 loop were necessary but not sufficient to confer CXCR4 usage.     

Population genetics,demographic and evolutionary history of the Dudley’s lousewort (Pedicularis dudleyi), a rare redwood forest specialist

Pedicularis dudleyi (Dudley’s Lousewort, Orobanchaceae) is an extremely rare plant endemic to the redwood forests of Central California. Until recently, the species was known only from three extant natural populations. However, in 2019, one of those populations was described as a novel species (P. rigginsiae D.J. Keil) based on morphological and ecological data leaving only two populations described as P. dudleyi. While little is known about the past distribution of the species, historical records have led to speculation that the species was once more widespread and may have suffered from habitat destruction as a result of widespread logging during the early twentieth century. We utilized a combination of ddRAD SNP and Sanger sequencing data to: (1) Test the morphological hypothesis that P. rigginsiae is distinct from P. dudleyi; (2) Describe the genetic diversity and population structure of P. dudleyi and; (3) Test the hypothesis that the species underwent a bottleneck corresponding with increased logging of redwood forests in the early twentieth century. Our results support the recognition of P. rigginsiae as distinct from P. dudleyi, increasing the conservation priority of both species. Genetic diversity statistics and analyses of genetic structure suggest that both populations of P. dudleyi are highly differentiated from each other with one population exhibiting unexpected substructure. Finally, demographic modeling supports a scenario where the contemporary rarity of the species is explained by a recent bottleneck.

     

Relative feeding and development of armyworm on switchgrass and corn, and its potential effects on switchgrass grown for biomass

To help assess the potential for damage by armyworms [Mythimna (Pseudaletia) unipuncta (Haworth) (Lepidoptera: Noctuidae)] to switchgrass (Panicum virgatum L.) and surrounding crops, survival and development were evaluated for larvae reared on leaves of switchgrass, corn (Zea mays L.), and miscanthus (Miscanthus x giganteus Greef and Deuter ex Hodkinson and Renvoize). Additional tests assessed the relationship between leaf position and the concentration of saponins (plant compounds which can provide protection from insect herbivores) and examined the effect of defoliation on switchgrass dry mass. Survival to adulthood was similar when larvae were reared on field-grown leaves of switchgrass and corn. However, lower larval mass (10 d) and delayed development of M. unipuncta (to pupation, adult emergence) suggest switchgrass is an inferior host relative to corn. When fed field-grown miscanthus, no larvae survived 10 d. Few differences were noted between switchgrass and corn grown under controlled (laboratory) conditions, but M. unipuncta survival seemed to decline rapidly when larvae were fed the fourth and fifth leaves of switchgrass. Switchgrass leaf samples collected from different leaf positions and stages of tiller maturity showed up to 10-fold differences in the concentration of the saponin protodioscin, with the greatest concentrations in the fourth and fifth leaves. However, other saponins showed an opposite pattern, indicating the role of protodioscin on insect development should be tested in isolation (e.g., by addition of the purified compound to an artificial diet). Defoliation trials indicated that extremely high M. unipuncta populations may be necessary to cause any significant reduction in switchgrass biomass. Collectively, results suggest M. unipuncta may not present a significant risk to biomass production in switchgrass, but that the spring emergence of switchgrass provides an alternate host for M. unipuncta before colonizing annual food and feed crops.     

Role of the active-site cysteine of Pseudomonas aeruginosa azurin. Crystal structure analysis of the CuII(Cys112Asp) protein

Replacement of the cysteine at position 112 of Pseudomonas aeruginosa azurin with an aspartic acid residue results in a mutant (Cys112Asp) protein that retains a strong copper-binding site. Cu^II(Cys112Asp) azurin can be reduced by excess [Ru^II(NH₃)₆]²⁺, resulting in a Cu^I protein with an electronic absorption spectrum very similar to that of wild-type Cu^I azurin. Cys112Asp azurin exhibits reversible interprotein electron-transfer reactivity with P. aeruginosa cytochrome c ₅₅₁ (μ?=?0.1?M sodium phosphate (pH?7.0);E°(Cu^II/I)?=?180 mV vs NHE); this redox activity indicates that electrons can still enter and exit the protein through the partially solvent-exposed imidazole ring of His117. The structure of Cu^II(Cys112Asp) azurin at 2.4-Å resolution shows that the active-site copper is five coordinate: the pseudo-square base of the distorted square-pyramidal structure is defined by the imidazole N^δ atoms of His46 and His117 and the oxygen atoms of an asymmetrically-bound bidentate carboxylate group of Asp112; the apical position is occupied by the oxygen atom of the backbone carbonyl group of Gly45. The Cu^II–Asp112 interaction is distinguished by an approximately 1.2-Å displacement of the metal center from the plane defined by the Asp112 carboxylate group.     

Prevalence of obese patients in a primary care setting

Objective: Previous research has addressed the relationship between BMI and chronic disease in primary care; however, little has been done with regard to the association between obesity and depression in primary care. The purpose of this paper is to assess the relationship between obesity and chronic conditions including depression. Research Methods and Procedures : Data from primary care patients seen at a university‐based family medicine clinic in the southeastern United States were extracted for the time between January 1, 1999 and January 1, 2002. Data extracted included most recent height and weight, age, ethnicity, pregnancy status, number of office visits, blood pressure, cholesterol, hemoglobin A_1C, current diagnoses, and medications. Results : A total of 8197 patients were included in the analysis. Sixty‐nine percent of patients seen in a 3‐year period were either overweight or obese. Comparing blood pressure, cholesterol, diagnoses, and medications between BMI groups found differences in virtually all categories. Diagnoses of high cholesterol, hypertension, diabetes, and depression significantly increased for obese patients. Discussion : Obese patients are over‐represented in primary care, and this over‐representation of obesity correlates with several diagnoses, including depression. Depression is a chronic disease that may interfere with health‐related behaviors and must be addressed within the health care system.