Jeroen Poels

Insulin-related peptides and their conserved signal transduction pathway.

The insulin superfamily is an ancient category of small, structurally related proteins, such as insulin, insulin-like growth factors (IGF) and relaxin. Insulin-like signaling molecules have also been described in different invertebrates, including nematodes, mollusks, and insects. They initiate an evolutionary conserved signal transduction mechanism by binding to a heterotetrameric, membrane-spanning receptor tyrosine kinase. Recent physiological and genetic studies have revealed that, in different metazoans, the insulin signaling pathway plays a pivotal role in the regulation of a variety of interrelated, fundamental processes, such as metabolism, growth, reproduction and aging.

Discrete mechanisms of mTOR and cell cycle regulation by AMPK agonists independent of AMPK

Significance Cancer cells reprogram their metabolism for optimal growth and survival. AMPK-activated protein kinase (AMPK) is a key energy sensor that controls many metabolic pathways including metabolic reprogramming. However, its role in cancer is poorly understood. Some studies claim that it has a tumor suppressor role while others show its protumor role. Two AMPK-activating compounds (including metformin, now in many clinical trials) are widely used to suppress cancer cell proliferation. We found that AMPK is abundantly expressed in high-grade gliomas and, in contrast to popular belief, these two AMPK activators suppressed glioma cell proliferation through unique AMPK-independent mechanisms. The multifunctional AMPK-activated protein kinase (AMPK) is an evolutionarily conserved energy sensor that plays an important role in cell proliferation, growth, and survival. It remains unclear whether AMPK functions as a tumor suppressor or a contextual oncogene. This is because although on one hand active AMPK inhibits mammalian target of rapamycin (mTOR) and lipogenesis—two crucial arms of cancer growth—AMPK also ensures viability by metabolic reprogramming in cancer cells. AMPK activation by two indirect AMPK agonists AICAR and metformin (now in over 50 clinical trials on cancer) has been correlated with reduced cancer cell proliferation and viability. Surprisingly, we found that compared with normal tissue, AMPK is constitutively activated in both human and mouse gliomas. Therefore, we questioned whether the antiproliferative actions of AICAR and metformin are AMPK independent. Both AMPK agonists inhibited proliferation, but through unique AMPK-independent mechanisms and both reduced tumor growth in vivo independent of AMPK. Importantly, A769662, a direct AMPK activator, had no effect on proliferation, uncoupling high AMPK activity from inhibition of proliferation. Metformin directly inhibited mTOR by enhancing PRAS40’s association with RAPTOR, whereas AICAR blocked the cell cycle through proteasomal degradation of the G2M phosphatase cdc25c. Together, our results suggest that although AICAR and metformin are potent AMPK-independent antiproliferative agents, physiological AMPK activation in glioma may be a response mechanism to metabolic stress and anticancer agents.

Drosophila molting neurohormone bursicon is a heterodimer and the natural agonist of the orphan receptor DLGR2.

Bursicon is a neurohumoral agent responsible for tanning and hardening of the cuticle and expansion of the wings during the final phase of insect metamorphosis. Although the hormonal activity was described more than 40 years ago, the molecular nature of bursicon has remained elusive. We identify here Drosophila bioactive bursicon as a heterodimer made of two cystine knot polypeptides. This conclusion was reached in part from the unexpected observation that in the genome of the honey bee, the orthologs of the two Drosophila proteins are predicted to be fused in a single open reading frame. The heterodimeric Drosophila protein displays bursicon bioactivity in freshly eclosed neck‐ligated flies and is the natural agonist of the orphan G protein‐coupled receptor DLGR2.

Expanding roles for AMP-activated protein kinase in neuronal survival and autophagy.

AMP‐activated protein kinase (AMPK) is an evolutionarily conserved cellular switch that activates catabolic pathways and turns off anabolic processes. In this way, AMPK activation can restore the perturbation of cellular energy levels. In physiological situations, AMPK senses energy deficiency (in the form of an increased AMP/ATP ratio), but it is also activated by metabolic insults, such as glucose or oxygen deprivation. Metformin, one of the most widely prescribed anti‐diabetic drugs, exerts its actions by AMPK activation. However, while the functions of AMPK as a metabolic regulator are fairly well understood, its actions in neuronal cells only recently gained attention. This review will discuss newly emerged functions of AMPK in neuroprotection and neurodegeneration. Additionally, recent views on the role of AMPK in autophagy, an important catabolic process that is also involved in neurodegeneration and cancer, will be highlighted.

Neuroendocrinological and Molecular Aspects of Insect Reproduction

This review summarizes recent advances and novel concepts in the area of insect reproductive neuroendocrinology. The role of ‘classic’ hormones, such as ecdysteroids and juvenoids, to control reproduction is well documented in a large variety of insect species. In adult gonads, ecdysteroids appear to induce a cascade of transcription factors, many of which also occur during the larval molting response. Recent molecular and functional data have created opportunities to study an additional level of regulation, that of neuropeptides, growth factors and their respective receptors. As a result, many homologs of factors playing a role in vertebrate reproductive physiology have been discovered in insects. This review highlights several neuropeptides controlling the biosynthesis and release of the ‘classic’ insect hormones, as well as various peptides and biogenic amines that regulate behavioural aspects of the reproduction process. In addition, hormone metabolizing enzymes and second messenger pathways are discussed with respect to their role in reproductive tissues. Finally, we speculate on future prospects for insect neuroendocrinological research as a consequence of the recent ‘Genomics Revolution’.

Characterization of a Receptor for Insect Tachykinin‐Like Peptide Agonists by Functional Expression in a Stable Drosophila Schneider 2 Cell Line

Abstract: STKR is an insect G protein‐coupled receptor, cloned from the stable fly Stomoxys calcitrans. It displays sequence similarity to vertebrate tachykinin [or neurokinin (NK)] receptors. Functional expression of the cloned STKR cDNA was obtained in cultured Drosophila melanogaster Schneider 2 (S2) cells. Insect tachykinin‐like peptides or “insectatachykinins,” such as Locusta tachykinin (Lom‐TK) III, produced dose‐dependent calcium responses in stably transfected S2‐STKR cells. Vertebrate tachykinins (or neurokinins) did not evoke any effect at concentrations up to 10‐5M, but an antagonist of mammalian neurokinin receptors, spantide II, inhibited the Lom‐TK III‐induced calcium response. Further analysis showed that the agonist‐induced intracellular release of calcium ions was not affected by pretreatment of the cells with pertussis toxin. The calcium rise was blocked by the phospholipase C inhibitor U73122. In addition, Lom‐TK III was shown to have a stimulatory effect on the accumulation of both inositol 1,4,5‐trisphosphate and cyclic AMP. These are the same second messengers that are induced in mammalian neurokinin‐dependent signaling processes.

Tachykinin-like Peptides and Their Receptors: A Review

Abstract: Tachykinin‐like peptides have been identified in many vertebrate and invertebrate species. On the basis of the data reviewed in this paper, these peptides can be classified into two distinct subfamilies, which are recognized by their respective sequence characteristics. All known vertebrate tachykinins and a few invertebrate ones share a common C‐terminal sequence motif, ‐FXGLMa. The insect tachykinins, which have a common ‐GFX1GX2Ra C‐terminus, display about 30% of sequence homology with the first group.

Tachykinin-related peptides and their receptors in invertebrates: A current view

Members of the tachykinin peptide family have been well conserved during evolution and are mainly expressed in the central nervous system and in the intestine of both vertebrates and invertebrates. In these animals, they act as multifunctional messengers that exert their biological effects by specifically interacting with a subfamily of structurally related G protein-coupled receptors. Despite the identification of multiple tachykinin-related peptides (TKRPs) in species belonging to the insects, crustaceans, mollusks and echiuroid worms, only five invertebrate receptors harboring profound sequence similarities to mammalian receptors for tachykinins have been functionally characterized to date. Three of these have been cloned from dipteran insect species, i.e. NKD (neurokinin receptor from Drosophila), DTKR (Drosophila tachykinin receptor) and STKR (tachykinin-related peptide receptor from the stable fly, Stomoxys calcitrans). In addition, two receptors from non-insect species, present in echiuroid worms and mollusks, respectively have been identified as well. In this brief review, we will survey some recent findings and insights into the signaling properties of invertebrate tachykinin-related peptides via their respective receptors. In this context, we will also point out the necessity to take into account differences in signaling mechanisms induced by distinct TKRP isoforms in insects.

Myoinhibiting peptides are the ancestral ligands of the promiscuous Drosophila sex peptide receptor

Male insects change behaviors of female partners by co-transferring accessory gland proteins (Acps) like sex peptide (SP), with their sperm. The Drosophila sex peptide receptor (SPR) is a G protein-coupled receptor expressed in the female’s nervous system and genital tract. While most Acps show a fast rate of evolution, SPRs are highly conserved in insects. We report activation of SPRs by evolutionary conserved myoinhibiting peptides (MIPs). Structural determinants in SP and MIPs responsible for this dual receptor activation are characterized. Drosophila SPR is also expressed in embryonic and larval stages and in the adult male nervous system, whereas SP expression is restricted to the male reproductive system. MIP transcripts occur in male and female central nervous system, possibly acting as endogenous SPR ligands. Evolutionary consequences of the promiscuous nature of SPRs are discussed. MIPs likely function as ancestral ligands of SPRs and could place evolutionary constraints on the MIP/SPR class.

Characterization and distribution of NKD, a receptor for Drosophila tachykinin-related peptide 6.

Neuropeptides related to vertebrate tachykinins have been identified in Drosophila and are referred to as drosotachykinins, or DTKs. Two Drosophila G protein-coupled receptors, designated NKD (neurokinin receptor from Drosophila; CG6515) and DTKR (Drosophila tachykinin receptor; CG7887), display sequence similarities to mammalian tachykinin receptors. Whereas DTKR was shown to be activated by DTKs [Birse RT, Johnson EC, Taghert PH, Nässel DR. Widely distributed Drosophila G-protein-coupled receptor (CG7887) is activated by endogenous tachykinin-related peptides. J Neurobiol 2006;66:33-46; Poels J, Verlinden H, Fichna J, Van Loy T, Franssens V, Studzian K, et al. Functional comparison of two evolutionary conserved insect neurokinin-like receptors. Peptides 2007;28:103-8] and was localized by immunocytochemistry in Drosophila central nervous system (CNS), agonist-dependent activation and distribution of NKD have not yet been investigated in depth. In the present study, we have challenged NKD-expressing mammalian and insect cells with a library of Drosophila neuropeptides and discovered DTK-6 as a specific agonist that can induce a calcium response in these cells. In addition, we have produced antisera to sequences from NKD protein to analyze receptor distribution. We found that NKD is less abundantly distributed in the central nervous system than DTKR, and only NKD was found in the intestine. In fact, the two receptors are distributed in mutually exclusive patterns in the CNS. The combined distribution of the receptors in brain neuropils corresponds well with the distribution of DTKs. Most interestingly, NKD appears to be activated only by DTK-6, known to possess an Ala-substitution in an otherwise conserved C-terminal core motif. Our findings suggest that NKD and DTKR provide substrates for two functionally and spatially separated peptide signaling systems.