Diterpenoids and triterpenoids from Euphorbia guyoniana

Two new compounds with tigliane and cycloartane skeletons: 4,12-dideoxy(4alpha)phorbol-13-hexadecanoate (1) and 24-methylenecycloartane-3,28-diol (2), respectively, in addition of four known diterpenoids and 13 triterpenoids: 3-benzoyloxy-5,15-diacetoxy-9,14-dioxojatropha-6(17),11-diene (4), ent-abieta-8(14),13(15)-dien-16,12-olide (5), ent-8alpha,14alpha-epoxyabieta-11,13(15)-dien-16,12-olide (6), ent-3-hydroxyatis-16(17)-ene-2,14-dione (7), 3beta-hydroxytaraxer-14-en-28-oic acid (8), beta-sitosteryl-3beta-glucopyranoside-6'-O-palmitate (9), multiflorenyl acetate (10), multiflorenyl palmitate (11), peplusol (12), 24-methylenecycloartanol (3), lanosterol (13), euferol (14), butyrospermol (15), cycloartenol (16), obtusifoliol (17), cycloeucalenol (18) and beta-sitosterol (19), were isolated from the roots of Euphorbia guyoniana. Their structures were established on the basis of physical and spectroscopic analysis, including 1D and 2D homo- and heteronuclear NMR experiments (COSY, HSQC, HMBC and NOESY) and by comparison with the literature data.     

Both caffeoyl Coenzyme A 3-<Emphasis Type="Italic">O</Emphasis>-methyltransferase 1 and caffeic acid <Emphasis Type="Italic">O</Emphasis>-methyltransferase 1 are involved in redundant functions for lignin,flavonoids and sinapoyl malate biosynthesis in Arabidopsis

Two methylation steps are necessary for the biosynthesis of monolignols, the lignin precursors. Caffeic acid O-methyltransferase (COMT) O-methylates at the C5 position of the phenolic ring. COMT is responsible for the biosynthesis of sinapyl alcohol, the precursor of syringyl lignin units. The O-methylation at the C3 position of the phenolic ring involves the Caffeoyl CoA 3-O-methyltransferase (CCoAOMT). The CCoAOMT 1 gene (At4g34050) is believed to encode the enzyme responsible for the first O-methylation in Arabidopsis thaliana. A CCoAOMT1 promoter-GUS fusion and immunolocalization experiments revealed that this gene is strongly and exclusively expressed in the vascular tissues of stems and roots. An Arabidopsis T-DNA null mutant named ccomt 1 was identified and characterised. The mutant stems are slightly smaller than wild-type stems in short-day growth conditions and has collapsed xylem elements. The lignin content of the stem is low and the S/G ratio is high mainly due to fewer G units. These results suggest that this O-methyltransferase is involved in G-unit biosynthesis but does not act alone to perform this step in monolignol biosynthesis. To determine which O-methyltransferase assists CCoAOMT 1, a comt 1 ccomt1 double mutant was generated and studied. The development of comt 1 ccomt1 is arrested at the plantlet stage in our growth conditions. Lignins of these plantlets are mainly composed of p-hydroxyphenyl units. Moreover, the double mutant does not synthesize sinapoyl malate, a soluble phenolic. These results suggest that CCoAOMT 1 and COMT 1 act together to methylate the C3 position of the phenolic ring of monolignols in Arabidopsis. In addition, they are both involved in the formation of sinapoyl malate and isorhamnetin.     

A rhizosphere fungus enhances Arabidopsis thermotolerance through production of an HSP90 inhibitor

The molecular chaperone HEAT SHOCK PROTEIN90 (HSP90) is essential for the maturation of key regulatory proteins in eukaryotes and for the response to temperature stress. Earlier, we have reported that fungi living in association with plants of the Sonoran desert produce small molecule inhibitors of mammalian HSP90. Here, we address whether elaboration of the HSP90 inhibitor monocillin I (MON) by the rhizosphere fungus Paraphaeosphaeria quadriseptata affects plant HSP90 and plant environmental responsiveness. We demonstrate that MON binds Arabidopsis (Arabidopsis thaliana) HSP90 and can inhibit the function of HSP90 in lysates of wheat (Triticum aestivum) germ. MON treatment of Arabidopsis seedlings induced HSP101 and HSP70, conserved components of the stress response. Application of MON, or growth in the presence of MON, allowed Arabidopsis wild type but not AtHSP101 knockout mutant seedlings to survive otherwise lethal temperature stress. Finally, cocultivation of P. quadriseptata with Arabidopsis enhanced plant heat stress tolerance. These data demonstrate that HSP90-inhibitory compounds produced by fungi can influence plant growth and responses to the environment.     

Stone artifacts and hominins in island Southeast Asia: new insights from Flores, eastern Indonesia

This study reexamines the current understanding of Pleistocene stone-artifact assemblages in island Southeast Asia. A differentiation has long been made between assemblages of large-sized "core tools" and assemblages of small-sized "flake tools." "Core tool" assemblages are often argued to be the handiwork of early hominin species such as Homo erectus, while small-sized "flake tool" assemblages have been attributed to Homo sapiens. We argue that this traditional Southeast Asian perspective on stone tools assumes that the artifacts recovered from a site reflect a complete technological sequence. Our analyses of Pleistocene-age artifact assemblages from Flores, Indonesia, demonstrate that large pebble-based cores and small flake-based cores are aspects of one reduction sequence. We propose that the Flores pattern applies across island Southeast Asia: large-sized "core tool" assemblages are in fact a missing element of the small-sized flake-based reduction sequences found in many Pleistocene caves and rock-shelters. We conclude by discussing the implications of this for associating stone-artifact assemblages with hominin species in island Southeast Asia.     

Cellular localization of apelin and its receptor in the anterior pituitary: evidence for a direct stimulatory action of apelin on ACTH release

Apelin is a bioactive peptide recently identified as the endogenous ligand of the human orphan G protein-coupled receptor APJ. The presence of apelin-immunoreactive nerve fibers, together with the detection of apelin receptor mRNA in the parvocellular part of the paraventricular nucleus and the stimulatory action of apelin on corticotropin-releasing hormone release, indicate that apelin modulates adrenocorticotropin (ACTH) release via an indirect action on the hypothalamus. However, a direct action of apelin in the anterior pituitary cannot be excluded. Here, we provided evidence for the existence of an apelinergic system within the adult male rat pituitary gland. Double immunofluorescence staining indicated that apelin is highly coexpressed in the anterior pituitary, mainly in corticotrophs (96.5 +/- 0.3%) and to a much lower extent in somatotropes (3.2 +/- 0.2%). Using in situ hybridization combined with immunohistochemistry, a high expression of apelin receptor mRNA was also found in corticotrophs, suggesting a local interaction between apelin and ACTH. In an ex vivo perifusion system of anterior pituitaries, apelin 17 (K17F, 10(-6) M) significantly increased basal ACTH release by 41%, whereas apelin 10 (R10F, 10(-6) M), an inactive apelin fragment, was ineffective. In addition, K17F but not R10F induced a dose-dependent increase in K(+)-evoked ACTH release, with maximal increase being observed for a 10(-6) M concentration. Taken together, these data outline the potential role of apelin as an autocrine/paracrine-acting peptide on ACTH release and provide morphological and neuroendocrine basis for further studies that explore the physiological role of apelin in the regulation of anterior pituitary functions.     

Autonomic involvement in the permanent metabolic programming of hyperinsulinemia in the high-carbohydrate rat model

Exposure to a high-carbohydrate (HC) milk formula during the suckling period results in permanent metabolic programming of hyperinsulinemia in HC rats. Previous studies have shown that hyperinsulinemia in HC rats involves a programmed hyperresponsiveness to glucose. However, the immediate onset and persistence of enhanced insulin secretion throughout life suggests a role for numerous factors that control insulin secretion. Present in vivo and in vitro studies have shown a role for altered autonomic activity, including increased parasympathetic and decreased sympathetic activities, in the maintenance of hyperinsulinemia in 100-day-old HC rats. HC rats were shown to be more sensitive to cholinergic-induced potentiation of glucose-stimulated insulin secretion (GSIS) in response to acetylcholine and showed increased sensitivity to blockade of cholinergic-induced insulin secretion by the muscarinic-type 3 receptor-specific antagonist 4-diphenylacetoxy-N-methylpiperidine. In addition, HC rats were less sensitive to adrenergic-induced inhibition of insulin secretion by oxymetazoline, whereas treatment with yohimbine resulted in increased GSIS. Furthermore, HC rats showed greater reductions in plasma insulin levels after vagotomy, as well as an attenuation of yohimbine-induced potentiation of GSIS, suggesting that yohimbine-mediated changes are mediated by parasympathetic activity. Changes in autonomic regulation of GSIS are supported by increased mRNA levels of the parasympathetic signaling molecules muscarinic-type 3 receptor, phospholipase Cbeta1, and protein kinase C-alpha and decreased levels of alpha(2a)-adrenergic receptors in islets from adult HC rats. In conclusion, metabolic programming of hyperinsulinemia throughout adulthood of HC rats involves changes in autonomic activity in response to the HC dietary intervention in the suckling period.     

Human CD5 protects circulating tumor antigen-specific CTL from tumor-mediated activation-induced cell death

We previously characterized several tumor-specific T cell clones from PBL and tumor-infiltrating lymphocytes of a lung cancer patient with identical TCR rearrangements and similar lytic potential, but with different antitumor response. A role of the TCR inhibitory molecule CD5 to impair reactivity of peripheral T cells against the tumor was found to be involved in this process. In this report, we demonstrate that CD5 also controls the susceptibility of specific T cells to activation-induced cell death (AICD) triggered by the tumor. Using a panel of tumor-infiltrating lymphocytes and PBL-derived clones expressing different levels of CD5, our results indicate that T lymphocyte AICD in response to the cognate tumor is inversely proportional to the surface expression level of CD5. They also suggest a direct involvement of CD5 in this process, as revealed by an increase in tumor-mediated T lymphocyte AICD following neutralization of the molecule with specific mAb. Mechanistically, our data indicate that down-regulation of FasL expression and subsequent inhibition of caspase-8 activation are involved in CD5-induced T cell survival. These results provide evidence for a role of CD5 in the fate of peripheral tumor-specific T cells and further suggest its contribution to regulate the extension of CTL response against tumor.     

The activating immunoreceptor NKG2D and its ligands are involved in allograft transplant rejection

Although the linkage between innate and adaptive immunity in transplantation has been recognized, the mechanisms underlying this cooperation remain to be fully elucidated. In this study, we show that early "danger" signals associated with transplantation lead to rapid up-regulation of NKG2D ligands. A second wave of NKG2D ligand up-regulation is mediated by the adaptive immune response to allografts. Treatment with an Ab to NKG2D was highly effective in preventing CD28-independent rejection of cardiac allografts. Notably, NKG2D blockade did not deplete CD8(+) T cells or NK1.1(+) cells nor affect their migration to the allografts. These results establish a functional role of NKG2D and its ligands in the rejection of solid organ transplants.