Why "suboptimal" is optimal: Jensen's inequality and ectotherm thermal preferences

Body temperature (T(b)) profoundly affects the fitness of ectotherms. Many ectotherms use behavior to control T(b) within narrow levels. These temperatures are assumed to be optimal and therefore to match body temperatures (Trmax) that maximize fitness (r). We develop an optimality model and find that optimal body temperature (T(o)) should not be centered at Trmax but shifted to a lower temperature. This finding seems paradoxical but results from two considerations relating to Jensen's inequality, which deals with how variance and skew influence integrals of nonlinear functions. First, ectotherms are not perfect thermoregulators and so experience a range of T(b). Second, temperature-fitness curves are asymmetric, such that a T(b) higher than Trmax depresses fitness more than will a T(b) displaced an equivalent amount below Trmax. Our model makes several predictions. The magnitude of the optimal shift (Trmax - To) should increase with the degree of asymmetry of temperature-fitness curves and with T(b) variance. Deviations should be relatively large for thermal specialists but insensitive to whether fitness increases with Trmax ("hotter is better"). Asymmetric (left-skewed) T(b) distributions reduce the magnitude of the optimal shift but do not eliminate it. Comparative data (insects, lizards) support key predictions. Thus, "suboptimal" is optimal.     

Colorimetric determination of pure Mg(2+)-dependent phosphatidate phosphatase activity

The malachite green-molybdate reagent was used for a colorimetric assay of pure Mg2(+)-dependent phosphatidate phosphatase activity. This enzyme plays a major role in fat metabolism. Enzyme activity was linear with time and protein concentration, and with the concentration of water-soluble dioctanoyl phosphatidate. The colorimetric assay was used to examine enzyme inhibition by phenylglyoxal, propranolol, and dimethyl sulfoxide. Pure enzyme and a water-soluble phosphatidate substrate were required for the assay, which should be applicable to a well-defined large-scale screen of Mg2(+)-dependent phosphatidate phosphatise inhibitors (or activators).     

Entamoeba histolytica mitosomes: organelles in search of a function

It has been more than eight years since the discovery of mitosomes (mitochondrial remnant organelles) in the intestinal human pathogen Entamoeba histolytica. Despite detailed knowledge about the biochemistry of this parasite and the completion of the E. histolytica genome sequencing project no physiological function has yet been unequivocally assigned to these organelles. Entamoeba mitosomes seem to be the most degenerate of all endosymbiosis-derived organelles studied to date. They do not appear to participate in energy metabolism and may have dispensed completely with the proteins required for iron-sulphur cluster biosynthesis. However, the large number of mitosomes found in E. histolytica trophozoites hints at a significant biological role for these organelles in their natural environment. Identifying the protein complement of mitosomes will provide answers as to their biological significance and the reason(s) for their retention in this parasite.     

Dermacentor variabilis: regulation of fibroblast migration by tick salivary gland extract and saliva

We examined the effects of tick SGx and saliva on basal- and platelet-derived growth factor (PDGF)-stimulated cell migration and extracellular signal-regulated kinase (ERK) signaling in fibroblasts. Repair of injured monolayers was delayed by SGx pretreatment and was not associated with reductions in cell number. In migration assays, SGx suppressed both basal- and PDGF-stimulated fibroblast movement. Furthermore, SGx and saliva reduced PDGF-stimulated ERK activity. Thus, the delayed repair of monolayer injuries resulted from SGx inhibiting fibroblast migratory responses to chemotactic signals. SGx also suppressed injury- and growth factor-induced ERK activation in renal epithelial OK cells. Our data suggest that maintenance of the tick feeding lesion results, in part, from suppressing ERK signaling and fibroblast migration, events playing integral roles in the wound healing response. The effects of SGx on cells not involved in wound healing suggest that a constituent(s) in tick saliva has global effects on the ERK signaling pathway.     

Intestinal macrophage/epithelial cell-derived CCL11/eotaxin-1 mediates eosinophil recruitment and function in pediatric ulcerative colitis

Clinical studies have demonstrated a link between the eosinophil-selective chemokines, eotaxins (eotaxin-1/CCL11 and eotaxin-2/CCL24), eosinophils, and the inflammatory bowel diseases, Crohn's disease and ulcerative colitis (UC). However, the cellular source and individual contribution of the eotaxins to colonic eosinophilic accumulation in inflammatory bowel diseases remain unclear. In this study we demonstrate, by gene array and quantitative PCR, elevated levels of eotaxin-1 mRNA in the rectosigmoid colon of pediatric UC patients. We show that elevated levels of eotaxin-1 mRNA positively correlated with rectosigmoid eosinophil numbers. Further, colonic eosinophils appeared to be degranulating, and the levels positively correlated with disease severity. Using the dextran sodium sulfate (DSS)-induced intestinal epithelial injury model, we show that DSS treatment of mice strongly induced colonic eotaxin-1 and eotaxin-2 expression and eosinophil levels. Analysis of eosinophil-deficient mice defined an effector role for eosinophils in disease pathology. DSS treatment of eotaxin-2(-/-) and eotaxin-1/2(-/-) mice demonstrated that eosinophil recruitment was dependent on eotaxin-1. In situ and immunofluorescence analysis-identified eotaxin-1 expression was restricted to intestinal F4/80(+)CD11b(+) macrophages in DSS-induced epithelial injury and to CD68(+) intestinal macrophages and the basolateral compartment of intestinal epithelial cells in pediatric UC. These data demonstrate that intestinal macrophage and epithelial cell-derived eotaxin-1 plays a critical role in the regulation of eosinophil recruitment in colonic eosinophilic disease such as pediatric UC and provides a basis for targeting the eosinophil/eotaxin-1 axis in UC.     

Creosote growth rate and reproduction increase in postfire environments

Human activities are changing patterns of ecological disturbance globally. In North American deserts, wildfire is increasing in size and frequency due to fuel characteristics of invasive annual grasses. Fire reduces the abundance and cover of native vegetation in desert ecosystems. In this study, we sought to characterize stem growth and reproductive output of a dominant native shrub in the Mojave Desert, creosote bush (Larrea tridentata (DC.) Coville) following wildfires that occurred in 2005. We sampled 55 shrubs along burned and unburned transects 12 years after the fires (2017) and quantified age, stem diameter, stem number, radial and vertical growth rates, and fruit production for each shrub. The shrubs on the burn transects were most likely postfire resprouts based on stem age while stems from unburn transects dated from before the fire. Stem and vertical growth rates for shrubs on burned transects were 2.6 and 1.7 times higher than that observed for shrubs on unburned transects. Fruit production of shrubs along burned transects was 4.7‐fold more than shrubs along paired unburned transects. Growth rates and fruit production of shrubs in burned areas did not differ with increasing distance from the burn perimeter. Positive growth and reproduction responses of creosote following wildfires could be critical for soil stabilization and re‐establishment of native plant communities in this desert system. Additional research is needed to assess if repeat fires that are characteristic of invasive grass‐fire cycles may limit these benefits.     

A Brevibacillus choshinensis system that secretes cytoplasmic proteins

Brevibacillus choshinensis has previously been shown to be a useful strain for the secretion of heterologous proteins via the Sec secretory pathway. This pathway involves the secretion of proteins prior to folding, whereas the alternative TAT (twin-arginine translocation) pathway enables pre-folded proteins to be secreted. We have modified the signal peptide of the Brevibacillus expression vector pNCMO2 to accommodate a Sec avoidance signal as well as the twin arginines required for secretion via the TAT system. Use of this modified signal peptide with the phosphotriesterase OpdA enabled B. choshinensis transformants to express and secrete the enzyme in an active and substantially pure form. The system was also used successfully to secrete two cytoplasmic proteins, the phosphotriesterase HocA from Pseudomonas monteilii and the phenylcarbamate-degrading enzyme, PCD, from Arthrobacter oxydans. The inhibitors carbonyl cyanide m-chlorophenyl hydrazine and sodium azide were used to confirm that secretion was occurring via the TAT secretion pathway. The modified B. choshinensis system we have developed may have general utility in secreting a wide range of heterologous proteins in active and conveniently processed form.     

Modulation of the electrophoretic mobility of the linker for activation of T cells (LAT) by the calcineurin inhibitors CsA and FK506: LAT is a potential substrate for PKC and calcineurin signaling pathways

The linker for activation of T cells (LAT) is essential for T cell activation. Cyclosporin A (CsA) and FK506, inhibitors of T cell proliferation, have been very useful for preventing autoimmune and inflammatory disease and graft rejection. However, both compounds are associated with side effects. We show that TCR ligation in the presence of FK506 or CsA induced rapid modifications in LAT that modulate the electrophoretic mobility of the molecule in SDS-PAGE. Calcineurin, a target for CsA and FK506, dephosphorylated LAT in vitro and restored its electrophoretic mobility. Stimulating T cells with the protein kinase C (PKC) activator PMA induced a shift in the mobility of LAT, whereas inhibitors of PKC blocked the effect of PMA. Thus, manipulating calcineurin or PKC activation alters the electrophoretic mobility of LAT. These results shed light on the molecular actions of CsA and FK506 in T cells and implicate LAT in mediating the drugs' actions.     

Obligate multivalent recognition of cell surface tomoregulin following selection from a multivalent phage antibody library

A therapeutic antibody candidate (AT-19) isolated using multivalent phage display binds native tomoregulin (TR) as a mul-timer not as a monomer. This report raises the importance of screening and selecting phage antibodies on native antigen and reemphasizes the possibility that potentially valuable antibodies are discarded when a monomeric phage display system is used for screening. A detailed live cell panning selection and screening method to isolate multivalently active antibodies is described. AT-19 is a fully human antibody recognizing the cell surface protein TR, a proposed prostate cancer target for therapeutic antibody internalization. AT-19 was isolated from a multivalent single-chain variable fragment (scFv) antibody library rescued with hyperphage. The required multivalency for isolation of AT-19 is supported by fluorescence activated cell sorting data demonstrating binding of the multivalent AT-19 phage particles at high phage concentrations and failure of monovalent particles to bind. Pure monomeric scFv AT-19 does not bind native receptor on cells, whereas dimeric scFv or immunoglobulin G binds with nanomolar affinity. The isolation of AT-19 antibody with obligate bivalent binding activity to native TR is attributed to the use of a multivalent display of scFv on phage and the method for selecting and screening by alternate use of 2 recombinant cell lines.