Insulin is a key hormone regulating the control of metabolism and the maintenance of normoglycaemia and normolipidaemia. Insulin acts by binding to its cell surface receptor, thus activating the receptor's intrinsic tyrosine kinase activity, resulting in receptor autophosphorylation and phosphorylation of several substrates. Tyrosine phosphorylated residues on the receptor itself and on subsequently bound receptor substrates provide docking sites for downstream signalling molecules, including adapters, protein serine/threonine kinases, phosphoinositide kinases and exchange factors. Collectively, those molecules orchestrate the numerous insulin-mediated physiological responses.A clear picture is emerging of the way in which insulin elicits several intracellular signalling pathways to mediate its physiologic functions. A further challenge, being pursued by several laboratories, is to understand the molecular mechanisms that underlie insulin action at the peripheral level, deregulation of which ultimately leads to hyperglycaemia and Type 2 diabetes.We review how circulating factors such as insulin itself, TNF-α, interleukins, fatty acids and glycation products influence insulin action through insulin signalling molecules themselves or through other pathways ultimately impinging on the insulin-signalling pathway. Understanding how the mechanism by which molecular insulin action is modulated by these factors will potentially provide new targets for pharmacological agents, to enable the control of altered glucose and lipid metabolism and diabetes. [Diabetologia (2004) 47:170-184]

Using the technique of differential display, a maize transcript was identified whose silk tissue expression is induced in the presence of the ear rot pathogen Fusarium graminearum. The 3445 nt transcript includes a 727 nt 5' untranslated leader with the potential for extensive secondary structure and represents the maize gene An2. An2 encodes a copalyl diphosphate synthase (CPS)-like protein with 60% amino acid sequence identity with the maize An1 gene product involved in gibberellin (GA) biosynthesis. Recombinant expression and functional analysis demonstrated that both AN1 and AN2 are ent-copalyl diphosphate (ent-CPP) synthases (ent-CPS). Notably, the presence of an additional ent-CPS gene is consistent with previous reports that maize GA biosynthesis can proceed in the absence of An1. In addition, northern blot analysis showed that An2 transcript levels were strongly up-regulated by Fusarium attack, with an increase in silk, husk and ear tip tissues as early as 6 h after inoculation of silk channels with spore suspensions of various Fusarium sp. Gene expression of a third maize CPS-like gene, Cpsl1, is not affected by Fusarium infection. The Fusarium-inducible nature of An2 is also consistent with a previous report that cell-free extracts from maize seedlings produce ent-CPP derived diterpenes in response to Fusarium infection. However, it is not known whether An2 is involved in defense-related secondary metabolism in addition to GA synthesis.

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In F. graminearum, the transcriptional regulator Tri6 is encoded within the trichothecene gene cluster and regulates genes involved in the biosynthesis of the secondary metabolite deoxynivalenol (DON). The Tri6 protein with its Cys2His2 zinc-finger may also conform to the class of global transcription regulators. This class of global transcriptional regulators mediate various environmental cues and generally responds to the demands of cellular metabolism. To address this issue directly, we sought to find gene targets of Tri6 in F. graminearum grown in optimal nutrient conditions. Chromatin immunoprecipitation followed by Illumina sequencing (ChIP-Seq) revealed that in addition to identifying six genes within the trichothecene gene cluster, Tri1, Tri3, Tri6, Tri7, Tri12 and Tri14, the ChIP-Seq also identified 192 additional targets potentially regulated by Tri6. Functional classification revealed that, among the annotated genes, ∼40% are associated with cellular metabolism and transport and the rest of the target genes fall into the category of signal transduction and gene expression regulation. ChIP-Seq data also revealed Tri6 has the highest affinity toward its own promoter, suggesting that this gene could be subject to self-regulation. Electro mobility shift assays (EMSA) performed on the promoter of Tri6 with purified Tri6 protein identified a minimum binding motif of GTGA repeats as a consensus sequence. Finally, expression profiling of F. graminearum grown under nitrogen-limiting conditions revealed that 49 out of 198 target genes are differentially regulated by Tri6. The identification of potential new targets together with deciphering novel binding sites for Tri6, casts new light into the role of this transcriptional regulator in the overall growth and development of F. graminearum.

The neuronal apoptosis inhibitory protein (NAIP) was identified as a candidate gene for the inherited neurodegenerative disorder spinal muscular atrophy. NAIP is the founding member of a human protein family that is characterized by highly conserved N-terminal motifs called baculovirus inhibitor of apoptosis repeats (BIR). Five members of the human family of inhibitor of apoptosis proteins including NAIP have been shown to be antiapoptotic in various systems. To date, a mechanism for the antiapoptotic effect of NAIP has not been elucidated. To investigate NAIP function, we found cytoprotection of NAIP-expressing primary cortical neurons treated to undergo caspase-3-dependent apoptosis. The additional treatment of these neurons with the pancaspase inhibitor boc-aspartyl(OMe)-fluoromethylketone did not result in increased survival. Similar cytoprotective effects were obtained using HeLa cells transiently transfected with a NAIP N-terminal construct and treated to undergo a caspase-3-dependent cell death. To examine whether NAIP inhibits caspases directly, recombinant N-terminal NAIP protein containing BIR domains was overexpressed, purified, and tested for caspase inhibition potential. Our results demonstrate that inhibition of caspases is selective and restricted to the effector group of caspases, with K i values as low as ϳ14 nM for caspase-3 and ϳ45 nM for caspase-7. Additional investigations with NAIP fragments containing either one or two NAIP BIRs revealed that the second BIR and to a lesser extent the third BIR alone are sufficient to mediate full caspase inhibition.

The insulin receptor substrate (IRS) proteins act as important mediators of insulin action. Their regulation serves to augment the speci¢city of the insulin signalling cascade. They can be regulated^both positively and negatively^at the level of phosphorylation, and signalling through these proteins can be further modulated through the actions of SOCS (suppressor of cytokine signalling) proteins. Understanding the mechanisms of IRS regulation will provide further insight into the pathophysiology of insulin resistance and type 2 diabetes. ß

Objective To compare maternal characteristics, prenatal care and newborn outcomes in a cohort of opioid-dependent pregnant women treated with methadone vs. buprenorphine. Methods Retrospective cohort study. 609 pregnant, opioid-dependent women were treated with methadone (n=248) or buprenorphine (n=361) between 2000–2012 at a single institution. Results Mothers treated with buprenorphine were more likely to start medication prior to or earlier in pregnancy, had longer gestation and larger infants. Newborns of buprenorphine- vs. methadone-maintained mothers required treatment for neonatal abstinence significantly less often and for a shorter duration. Conclusions These data suggest pregnancy outcomes following buprenorphine to treat opioid dependence during pregnancy in clinical practice are as good and often better than outcomes with methadone. These results are consistent with efficacy data from randomized clinical trials and further support the use of buprenorphine for the treatment of opioid-dependence during pregnancy.

Fusarium graminearum is a broad host pathogen threatening cereal crops in temperate regions around the world. To better understand how F. graminearum adapts to different hosts, we have performed a comparison of the transcriptome of a single strain of F. graminearum during early infection (up to 4 d post-inoculation) of barley, maize, and wheat using custom oligomer microarrays. Our results showed high similarity between F. graminearum transcriptomes in infected wheat and barley spike tissues. Quantitative RT-PCR was used to validate the gene expression profiles of 24 genes. Host-specific expression of genes was observed in each of the three hosts. This included expression of distinct sets of genes associated with transport and secondary metabolism in each of the three crops, as well as host-specific patterns for particular gene categories such as sugar transporters, integral membrane protein PTH11-like proteins, and chitinases. This study identified 69 F. graminearum genes as preferentially expressed in developing maize kernels relative to wheat and barley spikes. These host-specific differences showcase the genomic flexibility of F. graminearum to adapt to a range of hosts.