The protective effect of dietary fish oil on murine lupus

Dietary marine lipids markedly reduce the severity of glomerulonephritis and its associated mortality in inbred strains of mice developing autoimmune disease, a model for human systemic lupus erythematosus. We report here the influence of varying the dose of menhaden oil and the timing of its administration on the mortality of female (NZB x NZW) F1 mice. After ingesting 25 wt% menhaden oil (MO) for periods of 1.5 weeks to 12 months, there was a stable content of tissue n-3 fatty acids, with total n-3 fatty acids of 28% and 35% in spleen and liver, respectively. The extent of protection from mortality was dependent on the dose of MO with marked protection at doses of 11 to 25%, marginal protection at 5.5% and no protection at 2.5% MO. Delay in the institution of MO until ages 5 or 7 months still resulted in large reductions of mortality. Conversely, institution of a MO diet from 6 weeks until ages 5 to 7 months followed by a change to beef tallow resulted in little protection. Serum levels of 4 cyclooxygenase products were reduced ranging from 26 to 76% in mice fed MO diets, compared to mice fed beef tallow, based on radioimmunoassay. The degree of reduction of mortality on different doses of MO was correlated best with tissue levels of C22:5, and levels of C20:5 and C22:6 were similar at high and low doses of MO, suggesting that levels of 22:5 may be related to the protective effects of marine lipids on autoimmune disease.     

Codon usage in the vertebrate hemoglobins and its implications

A study of codon usage in vertebrate hemoglobins revealed an evolutionary trend toward elevated numbers of CpG codon boundary pairs in mammalian hemoglobin alpha genes. Selection for CpG codon boundaries countering the generally observed CpG suppression is strongly suggested by these data. These observations parallel recently published experimental results that indicate that constitutive expression of the human alpha-globin gene appears to be determined by regulatory information encoded within the structural gene. The possibility is raised that, in the absence of selection, CpG decay can be used to date the evolutionary origin of a mammalian alpha pseudogene from its active alpha gene.      

Identification of amino-terminal pro-C-type natriuretic peptide in human plasma

We report the first identification of a circulating peptide from the amino-terminal end of proCNP. A specific radioimmunoassay was established based on antisera to the synthetic peptide proCNP(1-15). Extracts of plasma, drawn from patients with congestive heart failure or from sheep with experimental heart failure, were subjected to size exclusion and reverse-phase high-pressure liquid chromatography (HPLC) coupled to radioimmunoassay (RIA). These studies revealed the presence of an immunoreactive peptide with a molecular weight (M(r) approximately 5 kDa) similar to that expected for NT-proCNP(1-50), a potential fragment released during processing of pro(CNP). The same material was isolated from extracts of homogenized ovine pituitary, a tissue known to be a relatively enriched source of CNP. Plasma NT-proCNP levels in 22 patients with congestive heart failure (9.7 +/- 0.5 pmol/L, mean +/- SEM, range 5.4-13.7 pmol/L) were raised (P = 0.003) compared to those in 16 healthy volunteers (7.4 +/- 0.3 pmol/L, range 5.7-10.7 pmol/L) and were higher than levels reported for CNP in similar subjects. This first identification of circulating NT-proCNP opens the possibility of studying the factors regulating CNP production and metabolism in vivo.     

Overlapping binding sites in protein phosphatase 2A for association with regulatory A and alpha-4 (mTap42) subunits

Diverse functions of protein Ser/Thr phosphatases depend on the distribution of the catalytic subunits among multiple regulatory subunits. In cells protein phosphatase 2A catalytic subunit (PP2Ac) mostly binds to a scaffold subunit (A subunit or PR65); however, PP2Ac alternatively binds to alpha-4, a subunit related to yeast Tap42 protein, which also associates with phosphatases PP4 or PP6. We mapped alpha-4 binding to PP2Ac to the helical domain, residues 19-165. We mutated selected residues and transiently expressed epitope-tagged PP2Ac to assay for association with A and alpha-4 subunits by co-precipitation. The disabling H118N mutation at the active site or the presence of the active site inhibitor microcystin-LR did not interfere with binding of PP2Ac to either the A subunit or alpha-4, showing that these are allosteric regulators. Positively charged side chains Lys(41), Arg(49), and Lys(74) on the back surface of PP2Ac are unique to PP2Ac, compared with phosphatases PP4, PP6, and PP1. Substitution of one, two, or three of these residues with Ala produced a progressive loss of binding to the A subunit, with a corresponding increase in binding to alpha-4. Conversely, mutation of Glu(42) in PP2Ac essentially eliminated PP2Ac binding to alpha-4, with an increase in binding to the A subunit. Reciprocal changes in binding because of mutations indicate competitive distribution of PP2Ac between these regulatory subunits and demonstrate that the mutated catalytic subunits retained a native conformation. Furthermore, neither the Lys(41)-Arg(49)-Lys(74) nor Glu(42) mutations affected the phosphatase-specific activity or binding to microcystin-agarose. Binding of PP2Ac to microcystin and to alpha-4 increased with temperature, consistent with an activation energy barrier for these interactions. Our results reveal that the A subunit and alpha-4 (mTap42) require charged residues in separate but overlapping surface regions to associate with the back side of PP2Ac and modulate phosphatase activity.     

Application of the osmotic virial equation in cryobiology

The multisolute osmotic virial equation is the only multisolute thermodynamic solution theory that has been derived from first principles and can make predictions of multisolute solution behaviour in the absence of multisolute solution data. Other solution theories either (i) include simplifying assumptions that do not take into account the interactions between different types of solute molecules or (ii) require fitting to multisolute data to obtain empirical parameters. The osmotic virial coefficients, which are obtained from single-solute data, can be used to make predictions of multisolute solution osmolality. The osmotic virial coefficients for a range of solutes of interest in cryobiology are provided in this paper, for use with concentration units of both molality and mole fraction, along with an explanation of the background and theory necessary to implement the multisolute osmotic virial equation.     

C-type Natriuretic Peptide: a novel biomarker of steroid induced bone toxicity in children with acute lymphoblastic leukemia (ALL)

Impaired bone growth and mineralization, and osteonecrosis are significant and common long-term sequelae of chemotherapy for childhood acute lymphoblastic leukemia (ALL). Here we have evaluated the relationship between linear bone growth during chemotherapy for ALL and bone derived C-type Natriuretic Peptide (CNP). CNP is known to be critical to normal endochondral bone growth in both rodents and humans, and plasma concentration of the amino terminal pro CNP (NTproCNP) is strongly correlated with concurrent height velocity in children. Plasma NTproCNP and CNP were measured by radio-immunoassay in 12 children aged 2-9 years during induction and maintenance chemotherapy for children with ALL. Height velocity was calculated from stadiometer readings at intervals of 3-12 months and related to plasma NTproCNP during each growth interval. Plasma NTproCNP was markedly suppressed in all subjects during induction chemotherapy. Brief periods of NTproCNP decline and rapid rebound during maintenance treatment coincided with the use of dexamethasone but not with other chemotherapeutics. Height velocity was markedly reduced during ALL induction but unaffected in maintenance phase, and these changes in growth were strongly correlated with plasma NTproCNP concentration. Plasma NTproCNP has potential use as a biomarker of glucocorticoid-induced bone toxicity.     

The integrated use of chemical insecticides and the entomopathogenic nematode,Steinernema carpocapsae (Nematoda: Steinernematidae), for the control of sweetpotato whitefly,Bemisia tabaci (Hemiptera: Aleyrodidae)

The integration of chemical insecticides and infective juveniles of the entomopathogenic nematode Steinernema carpocapsae (Wesier) (Nematoda: Steinernematidae), to control second instars of the sweetpotato whitefly, Bemisia tabaci Gennadius (Hemiptera: Aleyrodidae) was investigated. Using a sand bioassay, the effects of direct exposure of S. carpocapsae for 24 h to field rate dilutions of four insecticides (spiromesifen, thiacloprid, imidacloprid and pymetrozine) on infectivity to Galleria mellonella larvae were tested. Although all chemicals tested, except spiromesifen, produced acceptable nematode infectivity rates, they were all significantly less than the water control. The effect of insecticide treatment (dry residues of spiromesifen, thiacloprid and pymetrozine and soil drench of imidacloprid) on the efficacy of the nematode against B. tabaci was also investigated. Nematodes in combination with thiacloprid and spiromesifen gave higher B. tabaci mortality (86.5% and 94.3% respectively) compared to using nematodes alone (75.2%) on tomato plants. There was no significant difference in B. tabaci mortality when using the chemicals imidacloprid, pymetrozine and spiromesifen in conjunction with nematodes compared to using the chemicals alone. However, using thiacloprid in combination with the nematodes produced significantly higher B. tabaci mortality than using the chemical alone. The integration of S. carpocapsae and these chemical agents into current integrated pest management programmes for the control of B. tabaci is discussed.     

The structure of Tap42/alpha4 reveals a tetratricopeptide repeat-like fold and provides insights into PP2A regulation

Physiological functions of protein phosphatase 2A (PP2A) are determined via the association of its catalytic subunit (PP2Ac) with diverse regulatory subunits. The predominant form of PP2Ac assembles into a heterotrimer comprising the scaffolding PR65/A subunit together with a variable regulatory B subunit. A distinct population of PP2Ac associates with the Tap42/alpha4 subunit, an interaction mutually exclusive with that of PR65/A. Tap42/alpha4 is also an interacting subunit of the PP2Ac-related phosphatases, PP4 and PP6. Tap42/alpha4, an essential protein in yeast and suppressor of apoptosis in mammals, contributes to critical cellular functions including the Tor signaling pathway. Here, we describe the crystal structure of the PP2Ac-interaction domain of Saccharomyces cerevisiae Tap42. The structure reveals an all alpha-helical protein with striking similarity to 14-3-3 and tetratricopeptide repeat (TPR) proteins. Mutational analyses of structurally conserved regions of Tap42/alpha4 identified a positively charged region critical for its interactions with PP2Ac. We propose a scaffolding function for Tap42/alpha4 whereby the interaction of PP2Ac at its N-terminus promotes the dephosphorylation of substrates recruited to the C-terminal region of the molecule.