Linking plant genes to insect communities: Identifying the genetic bases of plant traits and community composition

Community genetics aims to understand the effects of intraspecific genetic variation on community composition and diversity, thereby connecting community ecology with evolutionary biology. Thus far, research has shown that plant genetics can underlie variation in the composition of associated communities (e.g., insects, lichen and endophytes), and those communities can therefore be considered as extended phenotypes. This work, however, has been conducted primarily at the plant genotype level and has not identified the key underlying genes. To address this gap, we used genome‐wide association mapping with a population of 445 aspen (Populus tremuloides) genets to identify the genes governing variation in plant traits (defence chemistry, bud phenology, leaf morphology, growth) and insect community composition. We found 49 significant SNP associations in 13 Populus genes that are correlated with chemical defence compounds and insect community traits. Most notably, we identified an early nodulin‐like protein that was associated with insect community diversity and the abundance of interacting foundation species (ants and aphids). These findings support the concept that particular plant traits are the mechanistic link between plant genes and the composition of associated insect communities. In putting the “genes” into “genes to ecosystems ecology”, this work enhances understanding of the molecular genetic mechanisms that underlie plant–insect associations and the consequences thereof for the structure of ecological communities.     

Myofibrillar gene expression in differentiating lobster claw muscles

Lobster claw muscles undergo a process of fiber switching during development, where isomorphic muscles containing a mixture of both fast and slow fibers, become specialized into predominantly fast, or exclusively slow, muscles. Although this process has been described using histochemical methods, we lack an understanding of the shifts in gene expression that take place. In this study, we used several complementary techniques to follow changes in the expression of a number of myofibrillar genes in differentiating juvenile lobster claw muscles. RNA probes complementary to fast and slow myosin heavy chain (MHC) mRNA were used to label sections of 7th stage (approximately 3 months old) juvenile claw muscles from different stages of the molt cycle. Recently molted animals (1-5 days postmolt) had muscles with distinct regions of fast and slow gene expression, whereas muscles from later in the molt cycle (7-37 days postmolt) had regions of fast and slow MHC expression that were co-mingled and indistinct. Real-time PCR was used to quantify several myofibrillar genes in 9th and 10th stages (approximately 6 months old) juvenile claws and showed that these genes were expressed at significantly higher levels in the postmolt claws, as compared with the intermolt and premolt claws. Finally, Western blot analyses of muscle fibers from juvenile lobsters approximately 3 to 30 months in age showed a shift in troponin-I (TnI) isoform expression as the fibers differentiated into the adult phenotypes, with expression of the adult fast fiber TnI pattern lagging behind the adult slow fiber TnI pattern. Collectively, these data show that juvenile and adult fibers differ both qualitatively and quantitative in the expression of myofibrillar proteins and it may take as much as 2 years for juvenile fibers to achieve the adult phenotype.     

Physiological Studies on Pea Tendrils : XIV. Effects of Mechanical Perturbation, Light, and 2-Deoxy-d-Glucose on Callose Deposition and Tendril Coiling

When excised tendrils of pea (Pisum sativum L. cv Alaska) are mechanically perturbed there is an immediate and transient increase in callose deposition in the sieve cells. Mechanical perturbation (MP) results in a coiling response in light-grown tendrils and in dark-adapted tendrils, provided, in the latter case, that they receive adequate illumination within a limited period of time after MP. In nonperturbed tendrils the number of callose deposits decreases to some minimum with increasing time in the dark, and their ability to coil in the dark in response to MP diminishes with time in the dark. The transient increase of callose deposition due to MP, however, occurs whether or not tendrils are dark adapted, and whether they receive light or are retained in the dark after MP. This indicates that if callose is directly involved in tendril coiling, then it exerts its effect on the sensory perception of the mechanical stimulus. In the present investigation, there is never tendril coiling without the transient increase in callose, and the time after MP at which the peak of callose deposition occurs precedes the time of the peak amount of coiling.     

The effects of environmental stress on steroid receptor levels and steroid-induced morphogenesis in Achlya ambisexualis

Incubation of Achlya ambisexualis at elevated temperature (heat shock) or in the presence of sodium arsenite resulted in an inhibition of steroid hormone-induced responsiveness. The effect of heat shock was time- and temperature-dependent and more severe than the effect of sodium arsenite. Incubation at 37 degrees C for 30 min completely abolished the steroid-induced response and full recovery was not observed until 6 h after a return to the normal growth temperature of 22 degrees C. Heat shock and arsenite treatment had no effect on the cellular uptake of the steroid hormone, but heat shock resulted in a time- and temperature-dependent loss in the cellular level of steroid receptors. In contrast, arsenite treatment had little effect on the concentration of steroid receptors. However, both heat shock and arsenite treatment produced a long-term (4 h) and transient (1 h) inhibition of total protein synthesis, respectively. The recovery of steroid-induced responsiveness following heat shock was observed after both protein synthesis and steroid hormone receptor levels had returned to normal values.     

Crosslinking of N-acetyl-phenylalanyl [s4U]tRNAPhe to protein S10 in the ribosomal P site

In order to identify ribosomal components involved in the peptidyl-tRNA binding site on the ribosome, tRNAPhe molecules were prepared in which cytidine residues had been chemically converted into 4-thiouridine (S4U). This nucleoside is photoactive at 335 nm and able to form covalent bonds with nearby nucleophilic groups. The thiolated AcPhe-tRNAPhe was bound to the ribosomal P site in the presence of poly(U) as verified by puromycin reactivity. Direct irradiation of the AcPhe-[s4U]tRNAPhe poly(U) 70-S ribosome complex induced crosslinking of the tRNA molecule exclusively to 30-S subunits. Analysis of the covalent complex revealed that AcPhe-[s4U]tRNAPhe was specifically crosslinked to protein S10.     

Human transferrin. Expression and iron modulation of chimeric genes in transgenic mice

Transferrin (TF) is a plasma protein that transports and is regulated by iron. The aim of this study was to characterize human TF gene sequences that respond in vivo to cellular signals affecting expression in various tissues and to iron administration. Chimeric genes were constructed containing 152, 622, and 1152 base pairs (bp) of the human TF5'-flanking region with the coding region of a reporter gene, CAT (chloramphenicol acetyltransferase), and introduced into the germ line of mice. Transgenes containing TF 5'-flanking sequences to -152 bp were expressed poorly in all tissues examined. In contrast, transgenes containing TF sequences to -622 or -1152 bp were expressed at high levels in brain and liver, greater than or equal to 1000-fold higher than tissues such as heart and testes. Liver and brain are major sites of endogenous TF mRNA synthesis, but liver mRNA levels are 10-fold higher than brain. A significant diminution of CAT enzymatic activity in liver accompanied iron administration in both TF(0.67) and TF(1.2)CAT transgenic mice, mimicking the decrease of transferrin in humans following iron overload. Levels of endogenous plasma transferrin also decreased in iron-treated transgenic mice. Transgenic mouse lines carrying human TF chimeric genes will be useful models for analyzing the regulation of human transferrin by iron and for determining the molecular basis of transferrin regulation throughout mammalian development into the aging process.     

Metabolism of 15-hydroxyeicosatetraenoic acid by Caco-2 cells

Monolayers of Caco-2 cells, a human enterocyte cell line, were incubated with [1-14C]15-hydroxyeicosatetraenoic acid (15-HETE), a lipid mediator of inflammation, and [1-14C]arachidonic acid. Both fatty acids were taken up readily and metabolized by Caco-2 cells. [1-14C]Arachidonic acid was directly esterified in cellular phospholipids and, to a lesser extent, in triglycerides. When [1-14C]15-hydroxyeicosatetraenoic acid was incubated with Caco-2 cells, about 10% was directly esterified into cellular lipids but most (55%) was beta-oxidized to ketone bodies, CO2, and acetate, with very little accumulation of shorter carbon chain products of partial beta-oxidation. The radiolabeled acetate generated from beta-oxidation of [1-14C]15-hydroxyeicosatetraenoic acid was incorporated into the synthesis of new fatty acids, primarily [14C]palmitate, which in turn was esterified into cellular phospholipids, with lesser amounts in triglycerides. Caco-2 cells were also incubated with [5,6,8,9,11,12,14,15-3H]15-hydroxyeicosatetraenoic acid; most of the radiolabel was recovered either in ketone bodies or in [3H]palmitate esterified in phospholipids and triglycerides, demonstrating that most of the [3H]15-hydroxyeicosatetraenoic acid underwent several cycles of beta-oxidation. The binding of both 15-hydroxyeicosatetraenoic acid and arachidonic acid to hepatic fatty acid binding protein, the only fatty acid binding protein in Caco-2 cells, was measured. The Kd (6.0 microM) for 15-HETE was three-fold higher than that for arachidonate (2.1 microM).     

Radiocarbon age of carbonate in fruits of Lithospermum from the early Bronze Age settlement of Hirbet ez-Zeraqōn (Jordan)

Uncarbonized fruits of the Boraginaceae occur widely in cultural layers of archaeological sites in the Mediterranean and the Near East. To date, interpreting their origin remains problematic. It is difficult for archaeobotanists to tell whether such fruits were deposited as part of the cultural layer or were introduced post-depositionally. In an attempt to answer this question for the early Bronze Age site of Hirbet ez-Zeraqon (Jordan), we used direct ¹⁴C dating of biogenic carbonate from calcareous fruits of Lithospermum. The radiocarbon ages of seven fossil samples of the fruits suggest that they originate from plants that grew during or around the time of occupation.