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Crustacean reproduction has been hypothesized to be under complex endocrinological regulation by peptide hormones. To further improve our understanding of the mechanisms underlying this complex regulation, knowledge is needed regarding the hormones not only of the central nervous system (CNS) such as the X-organ/sinus gland (XOSG), brain, and thoracic ganglia, but also the peripheral gonadal tissues. For example, in vertebrates, some gonadal peptide hormones including activin, inhibin, follistatin, and relaxin are known to be involved in the reproductive physiology. Therefore, it is highly likely that some peptide factors from the ovary are serving as the signals among peripheral tissues and central nervous tissues in crustaceans. In this work, we sought to find gonadal peptide hormones and peptide hormone receptors by analyzing the transcriptome of the ovary of the kuruma prawn Marsupenaeus japonicus. The generated ovarian transcriptome data led to the identification of five possible peptide hormones, including bursicon-α and -β, the crustacean hyperglycemic hormone (CHH)-like peptide, insulin-like peptide (ILP), and neuroparsin-like peptide (NPLP). Dominant gene expressions for the bursicons were observed in the thoracic ganglia and the ovary, in the CNS for the CHH-like peptide, in the heart for NPLP, and in the ovary for ILP. Since the gene expressions of CHH-like peptide and NPLP were affected by a CHH (Penaeus japonicus sinus gland peptide-I) from XOSG, we produced recombinant peptides for CHH-like peptide and NPLP using Escherichia coli expression system to examine their possible peripheral regulation. As a result, we found that the recombinant NPLP increased vitellogenin gene expression in incubated ovarian tissue fragments. Moreover, contigs encoding putative receptors for insulin-like androgenic gland factor, insulin, neuroparsin, and neuropeptide Y/F, as well as several contigs encoding orphan G-protein coupled receptors and receptor-type guanylyl cyclases were also identified in the ovarian transcriptome. These results suggest that reproductive physiology in crustaceans is regulated by various gonadal peptide hormones, akin to vertebrates. [ABSTRACT FROM AUTHOR]

Summary: Root system architecture (RSA) influences the effectiveness of resources acquisition from soils but the genetic networks that control RSA remain largely unclear.We used rhizoboxes, X‐ray computed tomography, grafting, auxin transport measurements and hormone quantification to demonstrate that Arabidopsis and Medicago CEP (C‐TERMINALLY ENCODED PEPTIDE)‐CEP RECEPTOR signalling controls RSA, the gravitropic set‐point angle (GSA) of lateral roots (LRs), auxin levels and auxin transport.We showed that soil‐grown Arabidopsis and Medicago CEP receptor mutants have a narrower RSA, which results from a steeper LR GSA. Grafting showed that CEPR1 in the shoot controls GSA. CEP receptor mutants exhibited an increase in rootward auxin transport and elevated shoot auxin levels. Consistently, the application of auxin to wild‐type shoots induced a steeper GSA and auxin transport inhibitors counteracted the CEP receptor mutant's steep GSA phenotype. Concordantly, CEP peptides increased GSA and inhibited rootward auxin transport in wild‐type but not in CEP receptor mutants.The results indicated that CEP–CEP receptor‐dependent signalling outputs in Arabidopsis and Medicago control overall RSA, LR GSA, shoot auxin levels and rootward auxin transport. We propose that manipulating CEP signalling strength or CEP receptor downstream targets may provide means to alter RSA. [ABSTRACT FROM AUTHOR]

The class B G protein-coupled receptor (GPCR) calcitonin receptor (CTR) is a drug target for osteoporosis and diabetes. N-glycosylation of asparagine 130 in its extracellular domain (ECD) enhances calcitonin hormone affinity with the proximal GlcNAc residue mediating this effect through an unknown mechanism. Here, we present two crystal structures of salmon calcitonin-bound, GlcNAc-bearing CTR ECD at 1.78 and 2.85 Å resolutions and analyze the mechanism of the glycan effect. The N130 GlcNAc does not contact the hormone. Surprisingly, the structures are nearly identical to a structure of hormone-bound, N-glycan-free ECD, which suggested that the GlcNAc might affect CTR dynamics not observed in the static crystallographic snapshots. Hydrogen-deuterium exchange mass spectrometry and molecular dynamics simulations revealed that glycosylation stabilized a β-sheet adjacent to the N130 GlcNAc and the N-terminal α-helix near the peptide-binding site while increasing flexibility of the peptide-binding site turret loop. These changes due to N-glycosylation increased the ligand on-rate and decreased its off-rate. The glycan effect extended to RAMP-CTR amylin receptor complexes and was also conserved in the related CGRP receptor. These results reveal that N-glycosylation can modulate GPCR function by altering receptor dynamics. Image 1 • How N-glycosylation enhances the ligand affinity of the calcitonin receptor is unknown. • Crystal structures of calcitonin bound to N-glycosylated calcitonin receptor ECD. • MD simulations and HDX-MS experiments show that N-glycans modulate ECD dynamics. • N-glycans similarly enhance the ligand affinity of related amylin and CGRP receptors. • N-glycosylation of GPCRs can modulate their function by altering receptor dynamics. [ABSTRACT FROM AUTHOR]

Plants use leucine-rich repeat receptor kinases (LRR-RKs) to sense sequence diverse peptide hormones at the cell surface. A 3.0-Å crystal structure of the LRR-RK GSO1/SGN3 regulating Casparian strip formation in the endodermis reveals a large spiral-shaped ectodomain. The domain provides a binding platform for 21 amino acid CIF peptide ligands, which are tyrosine sulfated by the tyrosylprotein sulfotransferase TPST/SGN2. GSO1/SGN3 harbors a binding pocket for sulfotyrosine and makes extended backbone interactions with CIF2. Quantitative biochemical comparisons reveal that GSO1/SGN3-CIF2 represents one of the strongest receptor-ligand pairs known in plants. Multiple missense mutations are required to block CIF2 binding in vitro and GSO1/SGN3 function in vivo. Using structure-guided sequence analysis we uncover previously uncharacterized CIF peptides conserved among higher plants. Quantitative binding assays with known and novel CIFs suggest that the homologous LRR-RKs GSO1/SGN3 and GSO2 have evolved unique peptide binding properties to control different developmental processes. A quantitative biochemical interaction screen, a CIF peptide antagonist and genetic analyses together implicate SERK proteins as essential coreceptor kinases required for GSO1/ SGN3 and GSO2 receptor activation. Ourwork provides amechanistic framework for the recognition of sequence-divergent peptide hormones in plants. [ABSTRACT FROM AUTHOR]

The purpose of this study was to compare changes in muscle insulin-like growth factor-I (IGF-I) content resulting from resistance-exercise training (RET) and creatine supplementation (CR). Male (n = 24) and female (n = 18) participants with minimal resistance-exercise-training experience (≥1 year) who were participating in at least 30 min of structured physical activity (i.e., walking, jogging, cycling) 3-5 ×/wk volunteered for the study. Participants were randomly assigned in blocks (gender) to supplement with creatine (CR: 0.25 g/kg lean-tissue mass for 7 days; 0.06 g/kg lean-tissue mass for 49 days; n = 22, 12 males, 10 female) or isocaloric placebo (PL: n = 20, 12 male, 8 female) and engage in a whole-body RET program for 8 wk. Eighteen participants were classified as vegetarian (lacto-ovo or vegan; CR: 5 male, 5 female; PL: 3 male, 5 female). Muscle biopsies (vastus lateralis) were taken before and after the intervention and analyzed for IGF-I using standard immunohistochemical procedures. Stained muscle cross-sections were examined microscopically and IGF-I content quantified using image-analysis software. Results showed that RET increased intramuscular IGF-I content by 67%, with greater accumulation from CR (+78%) than PL (+54%; p = .06). There were no differences in IGF-I between vegetarians and nonvegetarians. These findings indicate that creatine supplementation during resistance-exercise training increases intramuscular IGF-I concentration in healthy men and women, independent of habitual dietary routine. [ABSTRACT FROM AUTHOR]

A neuropeptide (Sco-CHH-L), belonging to the crustacean hyperglycemic hormone (CHH) superfamily and preferentially expressed in the pericardial organs (POs) of the mud crab Scylla olivacea, was functionally and structurally studied. Its expression levels were significantly higher than the alternative splice form (Sco-CHH) in the POs, and increased significantly after the animals were subjected to a hypo-osmotic stress. Sco-CHH-L, but not Sco-CHH, significantly stimulated in vitro the Na+, K+-ATPase activity in the posterior (6th) gills. Furthermore, the solution structure of Sco-CHH-L was resolved using nuclear magnetic resonance spectroscopy, revealing that it has an N-terminal tail, three α-helices (α2, Gly9−Asn28; α3, His34−Gly38; and α5, Glu62−Arg72), and a π-helix (π4, Cys43−Tyr54), and is structurally constrained by a pattern of disulfide bonds (Cys7–Cys43, Cys23–Cys39, and Cys26–Cys52), which is characteristic of the CHH superfamily-peptides. Sco-CHH-L is topologically most similar to the molt-inhibiting hormone from the Kuruma prawn Marsupenaeus japonicus with a backbone root-mean-square-deviation of 3.12 Å. Ten residues of Sco-CHH-L were chosen for alanine-substitution, and the resulting mutants were functionally tested using the gill Na+, K+-ATPase activity assay, showing that the functionally important residues (I2, F3, E45, D69, I71, and G73) are located at either end of the sequence, which are sterically close to each other and presumably constitute the receptor binding sites. Sco-CHH-L was compared with other members of the superfamily, revealing a folding pattern, which is suggested to be common for the crustacean members of the superfamily, with the properties of the residues constituting the presumed receptor binding sites being the major factors dictating the ligand–receptor binding specificity.

Background and Objective: Calcitonin is a small peptide hormone including 32 amino acids and 3.4 KD molecular weight which is produced by the parafollicular cells of the thyroid gland in respond to increasing calcium ions in serum. This peptide is used for adjuvant therapy of osteoporosis, Paget's disease and hypercalcemic shock. In this study, the heterologuse expression of calcitonin was done in Escherichia coli. Methods: In this experimental study, the thioredoxin fusion partner was added to n-terminal of the Salmon calciton in order to increase its stability by the synthetic biology. The recombinant construct was transformed and over expressed into Escherichia coli BL21 (DE3) host cell. Results: SDS-PAGE analysis showed the over expression of recombinant protein after IPTG induction. Conclusion: In this study, the construct including fused Salmon calcitonin gene with thioredoxin was cloned. The SDS-PAGE result showed the stable expression of fused calcitonin.

Various secreted peptides, including peptide hormones, are present in the apoplast, but their biochemical characterization remains a challenge due to their low abundance, difficulty in extraction, and interference from numerous secondary metabolites. Here, we describe a simple and straightforward protocol for the extraction of apoplastic peptides with a high purity. This protocol takes advantage of the fact that apoplastic peptides diffuse and accumulate in the culture medium when Arabidopsis seedlings are subjected to whole-plant submerged culture. The peptides in the culture medium are efficiently recovered by o-chlorophenol extraction followed by acetone precipitation. The recovered peptides can be subjected to nano-liquid chromatography coupled to tandem mass spectrometry (nano-LC-MS/MS) without any additional clean-up. This procedure enables the structural elucidation of mature peptide hormones in the apoplast with the use of Arabidopsis plants that overexpress peptide hormone genes., (© 2024. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.)

Peptide therapeutics is gaining momentum. Advances in the field of peptidomics have enabled researchers to harvest vital information from various organisms and tissue types concerning peptide existence, expression and function. The development of mass spectrometry techniques for high-throughput peptide quantitation has paved the way for the identification and discovery of numerous known and novel peptides. Though much has been achieved, scientists are still facing difficulties when it comes to reducing the search space of the large mass spectrometry-generated peptidomics datasets and focusing on the subset of functionally relevant peptides. Moreover, there is currently no straightforward way to analytically compare the distributions of bioactive peptides in distinct biological samples, which may reveal much useful information when seeking to characterize tissue- or fluid-specific peptidomes. In this chapter, we demonstrate how to identify, rank, and compare predicted bioactive peptides and bioactivity distributions from extensive peptidomics datasets. To aid this task, we utilize MultiPep, a multi-label deep learning approach designed for classifying peptide bioactivities, to identify bioactive peptides. The predicted bioactivities are synergistically combined with protein information from the UniProt database, which assist in navigating through the jungle of putative therapeutic peptides and relevant peptide leads., (© 2024. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.)

The insulin receptor (IR, with its isoforms IR-A and IR-B) and the insulin-like growth factor 1 receptor (IGF-1R) are related tyrosine kinase receptors. Recently, the portfolio of solved hormone-receptor structures has grown extensively thanks to advancements in cryo-electron microscopy. However, the dynamics of how these receptors transition between their inactive and active state are yet to be fully understood. The C-terminal part of the alpha subunit ( α CT) of the receptors is indispensable for the formation of the hormone-binding site. We mutated the α CT residues Arg717 and His710 of IR-A and Arg704 and His697 of IGF-1R. We then measured the saturation binding curves of ligands on the mutated receptors and their ability to become activated. Mutations of Arg704 and His697 to Ala in IGF-1R decreased the binding of IGF-1. Moreover, the number of binding sites for IGF-1 on the His697 IGF-1R mutant was reduced to one-half, demonstrating the presence of two binding sites. Both mutations of Arg717 and His710 to Ala in IR-A inactivated the receptor. We have proved that Arg717 is important for the binding of insulin to its receptor, which suggests that Arg717 is a key residue for the transition to the active conformation.

CEPs (C-TERMINALLY ENCODED PEPTIDEs) inhibit Arabidopsis primary root growth by unknown mechanisms. We investigated how CEP3 levels control primary root growth. CEP3 peptide application decreased cell division, S-phase cell number, root meristematic cell number, and meristem zone (MZ) size in a dose- and CEP RECEPTOR1 -dependent manner. Grafting showed that CEP3-dependent growth inhibition requires root and shoot CEPR1. CEP3 induced mitotic quiescence in MZ cells significantly faster than that induced by nutrient limitation alone. CEP3 also inhibited the restoration of S-phase to mitotically quiescence cells by nutrient resupply without quantitatively reducing TARGET OF RAPAMYCIN (TOR) kinase activity. In contrast, cep3-1 had an increased meristem size and S-phase cell number under nitrogen (N)-limited conditions, but not under N-sufficient conditions. Furthermore, cep3-1 meristematic cells remained in S-phase longer than wild-type cells during a sustained carbon (C) and N limitation. RNA sequencing showed that CEP3 peptide down-regulated genes involved in S-phase entry, cell wall and ribosome biogenesis, DNA replication, and meristem expansion, and up-regulated genes involved in catabolic processes and proteins and peptides that negatively control meristem expansion and root growth. Many of these genes were reciprocally regulated in cep3-1. The results suggest that raising CEP3 induces starvation-related responses that curtail primary root growth under severe nutrient limitation. [ABSTRACT FROM AUTHOR]

Lateral root (LR) proliferation is a major determinant of soil nutrient uptake. How resource allocation controls the extent of LR growth remains unresolved. We used genetic, physiological, transcriptomic, and grafting approaches to define a role for C-TERMINALLY ENCODED PEPTIDE RECEPTOR 1 (CEPR1) in controlling sucrose-dependent LR growth. CEPR1 inhibited LR growth in response to applied sucrose, other metabolizable sugars, and elevated light intensity. Pathways through CEPR1 restricted LR growth by reducing LR meristem size and the length of mature LR cells. RNA-sequencing of wild-type (WT) and cepr1-1 roots with or without sucrose treatment revealed an intersection of CEP–CEPR1 signalling with the sucrose transcriptional response. Sucrose up-regulated several CEP genes, supporting a specific role for CEP–CEPR1 in the response to sucrose. Moreover, genes with basally perturbed expression in cepr1-1 overlap with WT sucrose-responsive genes significantly. We found that exogenous CEP inhibited LR growth via CEPR1 by reducing LR meristem size and mature cell length. This result is consistent with CEP–CEPR1 acting to curtail the extent of sucrose-dependent LR growth. Reciprocal grafting indicates that LR growth inhibition requires CEPR1 in both the roots and shoots. Our results reveal a new role for CEP–CEPR1 signalling in controlling LR growth in response to sucrose. [ABSTRACT FROM AUTHOR]

Orchestration of endocrine factors modulates many biological and physiological events including feeding behaviour. Concomitantly occurring impacts by neuropeptides on food intake and visceral contraction imply endocrine communication among the bioactive neuropeptides, which will cause successful feeding behaviour in insects. A number of physiological studies have demonstrated that neuropeptides and peptide hormones are involved in regulating insect feeding behaviours. In particular, allatoregulatory peptides modulate both food intake and gut contraction. In addition, adipokinetic hormone and insulin-like peptides might be significant factors contributing to energy homeostasis. Harmonization of those factors may generate the innate feeding behaviour. In this review, we address the communication of those endocrine factors which may lead to a putative endocrine network to execute a nutritionally appropriate feeding behaviour in insects. [ABSTRACT FROM AUTHOR]

Apelin is a peptide hormone that binds to a class A GPCR (the apelin receptor/APJ) to regulate various bodily systems. Upon signal peptide removal, the resulting 55-residue isoform, proapelin/apelin-55, can be further processed to 36-, 17-, or 13-residue isoforms with length-dependent pharmacological properties. Processing was initially proposed to occur intracellularly. However, detection of apelin-55 in extracellular fluids indicates that extracellular processing may also occur. To test for this, apelin-55 was applied exogenously to HEK293A cells overexpressing proprotein convertase subtilisin kexin 3 (PCSK3), the only apelin processing enzyme identified thus far, and to differentiated 3T3-L1 adipocytes, which endogenously express apelin, PCSK3 and other proprotein convertases. Analysis of culture media constituents from each cell type by high performance liquid chromatography-mass spectrometry and western blot demonstrated a time-dependent decrease in apelin-55 levels. This decrease was partially, but not fully, attenuated by PCSK inhibitor treatment in both cell lines. Comparison of the resulting apelin-55-derived peptide profile between the two cell lines demonstrated distinct processing patterns, with apelin-36 production apparent in 3T3-L1 adipocytes vs. detection of the prodomain of a shorter isoform (likely the apelin-13 prodomain, observed after additional proteolytic processing) in PCSK3-transfected HEK293A cells. Extracellular processing of apelin, with distinct cell type dependence, provides an alternative mechanism to regulate isoform-mediated physiological effects of apelin. [ABSTRACT FROM AUTHOR]

Legumes form root nodules to house beneficial nitrogen‐fixing rhizobia bacteria. However, nodulation is resource demanding; hence, legumes evolved a systemic signalling mechanism called autoregulation of nodulation (AON) to control nodule numbers. AON begins with the production of CLE peptides in the root, which are predicted to be glycosylated, transported to the shoot, and perceived. We synthesized variants of nodulation‐suppressing CLE peptides to test their activity using petiole feeding to introduce CLE peptides into the shoot. Hydroxylated, monoarabinosylated, and triarabinosylated variants of soybean GmRIC1a and GmRIC2a were chemically synthesized and fed into recipient Pisum sativum (pea) plants, which were used due to the availability of key AON pathway mutants unavailable in soybean. Triarabinosylated GmRIC1a and GmRIC2a suppressed nodulation of wild‐type pea, whereas no other peptide variant tested had this ability. Suppression also occurred in the supernodulating hydroxyproline O‐arabinosyltransferase mutant, Psnod3, but not in the supernodulating receptor mutants, Pssym29, and to some extent, Pssym28. During our study, bioinformatic resources for pea became available and our analyses identified 40 CLE peptide‐encoding genes, including orthologues of nodulation‐suppressive CLE peptides. Collectively, we demonstrated that soybean nodulation‐suppressive CLE peptides can function interspecifically in the AON pathway of pea and require arabinosylation for their activity. Autoregulation of nodulation is the systemic signalling mechanism used by legumes to regulate the number of root nodules they form in symbiosis with compatible rhizobia bacteria. Several components of the pathway are now known, including rhizobia‐induced CLE peptides that are produced in the root and transported to the shoot; however, further characterization of these peptides is required to better understand their mode of action. Delivering synthetic glycosylated CLE peptides into the shoot via petiole feeding suppressed nodulation in wild‐type pea plants, whereas partially modified peptides failed to suppress. The activity of the CLE peptides was also examined in key autoregulation of nodulation mutants of pea, with findings demonstrating that arabinosylation was required for their activity. [ABSTRACT FROM AUTHOR]

Vesicles in neurons and neuroendocrine cells store neurotransmitters and peptide hormones, which are released by vesicle fusion in response to Ca2+‐evoking stimuli. Synaptotagmin‐1 (Syt1), a Ca2+ sensor, mediates ultrafast exocytosis in neurons and neuroendocrine cells. After vesicle docking, Syt1 has two main groups of binding partners: anionic phospholipids and the SNARE (soluble N‐ethylmaleimide‐sensitive factor attachment protein receptors) complex. The molecular mechanisms by which Syt1 triggers vesicle fusion remain controversial. This Review introduces and summarizes six molecular models of Syt1: (a) Syt1 triggers SNARE unclamping by displacing complexin, (b) Syt1 clamps SNARE zippering, (c) Syt1 causes membrane curvature, (d) membrane bridging by Syt1, (e) Syt1 is a vesicle‐plasma membrane distance regulator, and (f) Syt1 undergoes circular oligomerization. We discuss important conditions to test Syt1 activity in vitro and attempt to illustrate the possible roles of Syt1. [ABSTRACT FROM AUTHOR]

Amyloids are highly ordered fibrous cross-β protein aggregates that are notorious primarily because of association with a variety of incurable human and animal diseases (termed amyloidoses), including Alzheimer’s disease (AD), Parkinson’s disease (PD), type 2 diabetes (T2D), and prion diseases. Some amyloid-associated diseases, in particular T2D and AD, are widespread and affect hundreds of millions of people all over the world. However, recently it has become evident that many amyloids, termed “functional amyloids,” are involved in various activities that are beneficial to organisms. Functional amyloids were discovered in diverse taxa, ranging from bacteria to mammals. These amyloids are involved in vital biological functions such as long-term memory, storage of peptide hormones and scaffolding melanin polymerization in animals, substrate attachment, and biofilm formation in bacteria and fungi, etc. Thus, amyloids undoubtedly are playing important roles in biological and pathological processes. This review is focused on functional amyloids in mammals and summarizes approaches used for identifying new potentially amyloidogenic proteins and domains.

Nucleation and growth of amyloid fibrils were found to only occur in supersaturated solutions above a critical concentration (c crit ). The biophysical meaning of c crit remained mostly obscure, since typical low values of c crit in the sub-μM range hamper investigations of potential oligomeric states and their structure. Here, we investigate the parathyroid hormone PTH 84 as an example of a functional amyloid fibril forming peptide with a comparably high c crit of 67±21 μM. We describe a complex concentration dependent prenucleation ensemble of oligomers of different sizes and secondary structure compositions and highlight the occurrence of a trimer and tetramer at c crit as possible precursors for primary fibril nucleation. Furthermore, the soluble state found in equilibrium with fibrils adopts to the prenucleation state present at c crit . Our study sheds light onto early events of amyloid formation directly related to the critical concentration and underlines oligomer formation as a key feature of fibril nucleation. Our results contribute to a deeper understanding of the determinants of supersaturated peptide solutions. In the current study we present a biophysical approach to investigate c crit of amyloid fibril formation of PTH 84 in terms of secondary structure, cluster size and residue resolved intermolecular interactions during oligomer formation. Throughout the investigated range of concentrations (1 μM to 500 μM) we found different states of oligomerization with varying ability to contribute to primary fibril nucleation and with a concentration dependent equilibrium. In this context, we identified the previously described c crit of PTH 84 to mark a minimum concentration for the formation of homo-trimers/tetramers. These investigations allowed us to characterize molecular interactions of various oligomeric states that are further converted into elongation competent fibril nuclei during the lag phase of a functional amyloid forming peptide., (© 2023 The Authors. ChemPhysChem published by Wiley-VCH GmbH.)

Peptides are the largest/most diverse class of molecules used by animals for chemical communication, and with their cognate receptors, are key players in modulating physiological/behavioral control systems, including those involved in adaptation to environmental change. Crustaceans have long served as models for investigating peptidergic control of physiology/behavior, and members of Notostraca, an ancient branchiopod order, have recently been proposed as models for investigating the genetic/physiological underpinnings of ecoresponsiveness; nothing is currently known about the genes/proteins underlying peptidergic signaling in any member of this crustacean taxon. Transcriptome mining is a powerful tool for peptidome prediction in crustaceans, and all large-scale discovery of crustacean peptide receptors has been achieved via transcriptomics. Here, in silico transcriptome mining was used to elucidate the peptidergic signaling systems of the tadpole shrimp Triops newberryi , a member of Notostraca. Transcripts encoding putative precursor proteins and/or receptors for 28 peptide families were identified within the T. newberryi dataset. The deduced precursor proteins included those for allatostatin A, allatostatin B, allatostatin C, allatotropin, bursicon, CCHamide, crustacean cardioactive peptide, crustacean hyperglycemic hormone, diuretic hormone 44, ecdysis-triggering hormone, eclosion hormone, elevenin, FMRFamide-like peptide, glycoprotein hormone, GSEFLamide, inotocin, insulin-like peptide, neuroparsin, neuropeptide F, orcokinin/orcomyotropin, proctolin, pyrokinin/periviscerokinin, SIFamide, sulfakinin and tachykinin-related peptide; 117 distinct mature peptides were predicted from the collective set of deduced pre/preprohormones. Transcripts encoding putative receptors for most of the abovementioned peptide groups were also identified from the T. newberryi assembly, as were those for several families for which no precursors were found, i.e. , corazonin, RYamide and short neuropeptide F. This is the first description of a peptidome and peptide receptors from any member of the Notostraca, and as such, provide a foundation for beginning to investigate the roles played by peptidergic signaling systems in T. newberryi and other notostracans, including how they may contribute to modulating organism-environment interactions. [ABSTRACT FROM AUTHOR]

Abstract: The development of a highly sensitive analytical method for oxytocin could be useful in the diagnosis and treatment of autistic spectrum disorder. We previously developed a colorimetric enzyme immunoassay (EIA) for plasma oxytocin measurement. In this study, we developed a method to measure oxytocin concentrations using a higher sensitivity bioluminescent EIA. Biotinylated oxytocin bridged with five lysine residues was used in a competitive format. The standard curve range for oxytocin was 1.0 to 1000 pg/assay. In addition, there was good correlation between the colorimetric and bioluminescent immunoassays in terms of measured oxytocin concentration (r = 0.9665, n = 48). The bioluminescent EIA for plasma oxytocin was more rapid and provided higher sensitivity than the colorimetric immunoassay, making it suitable for clinical application. [ABSTRACT FROM AUTHOR]

Peptides encoded by small coding genes play an important role in plant development, acting in a similar manner as phytohormones. Few hormone-like peptides, however, have been shown to play a role in abiotic stress tolerance. In the current study, 17 Arabidopsis genes coding for small peptides were found to be up-regulated in response to salinity stress. To identify peptides leading salinity stress tolerance, we generated transgenic Arabidopsis plants overexpressing these small coding genes and assessed survivability and root growth under salinity stress conditions. Results indicated that 4 of the 17 overexpressed genes increased salinity stress tolerance. Further studies focused on AtPROPEP3, which was the most highly up-regulated gene under salinity stress. Treatment of plants with synthetic peptides encoded by AtPROPEP3 revealed that a C-terminal peptide fragment (AtPep3) inhibited the salt-induced bleaching of chlorophyll in seedlings. Conversely, knockdown AtPROPEP3 transgenic plants exhibited a hypersensitive phenotype under salinity stress, which was complemented by the AtPep3 peptide. This functional AtPep3 peptide region overlaps with an AtPep3 elicitor peptide that is related to the immune response of plants. Functional analyses with a receptor mutant of AtPep3 revealed that AtPep3 was recognized by the PEPR1 receptor and that it functions to increase salinity stress tolerance in plants. Collectively, these data indicate that AtPep3 plays a significant role in both salinity stress tolerance and immune response in Arabidopsis. [ABSTRACT FROM AUTHOR]

Summary: Peptide hormones are implicated in many important aspects of plant life and are usually synthesized as precursor proteins. In contrast to animals, data for plant peptide hormone maturation are scarce and the specificity of processing enzyme(s) is largely unknown. Here we tested a hypothesis that processing of prosystemin, a precursor of tomato (Solanum lycopersicum) wound hormone systemin, is performed by phytaspases, aspartate‐specific proteases of the subtilase family. Following the purification of phytaspase from tomato leaves, two tomato phytaspase genes were identified, the cDNAs were cloned and the recombinant enzymes were obtained after transient expression in Nicotiana benthamiana. The newly identified tomato phytaspases hydrolyzed prosystemin at two aspartate residues flanking the systemin sequence. Site‐directed mutagenesis of the phytaspase cleavage sites in prosystemin abrogated not only the phytaspase‐mediated processing of the prohormone in vitro, but also the ability of prosystemin to trigger the systemic wound response in vivo. The data show that the prohormone prosystemin requires processing for signal biogenesis and biological activity. The identification of phytaspases as the proteases involved in prosystemin maturation provides insight into the mechanisms of wound signaling in tomato. Our data also suggest a novel role for cell death‐related proteases in mediating defense signaling in plants. [ABSTRACT FROM AUTHOR]

Small peptides mediate cell–cell communication to coordinate a variety of plant developmental processes. Signaling peptides specifically bind to the extracellular domains of receptors that belong to the receptor-like kinase family, and the peptide–receptor interaction activates a range of biochemical and physiological processes. The plant root is crucial for the anchorage of plants in soil as well as for the uptake of water and nutrients. Over recent years great progress has been made in the identification of receptors, structural analysis of peptide–receptor pairs, and characterization of their signaling pathways during plant root development. We review here recent advances in the elucidation of the functions and molecular mechanisms of signaling peptides, the peptide–receptor pairs that activate signal initiation, and their signaling pathways during root development. [ABSTRACT FROM AUTHOR]

Abstract: We are developing a rapid, time‐resolved method using laser‐activated cross‐linking to capture protein‐peptide interactions as a means to interrogate the interaction of serum proteins as delivery systems for peptides and other molecules. A model system was established to investigate the interactions between bovine serum albumin (BSA) and 2 peptides, the tridecapeptide budding‐yeast mating pheromone (α‐factor) and the decapeptide human gonadotropin‐releasing hormone (GnRH). Cross‐linking of α‐factor, using a biotinylated, photoactivatable p‐benzoyl‐L‐phenylalanine (Bpa)–modified analog, was energy‐dependent and achieved within seconds of laser irradiation. Protein blotting with an avidin probe was used to detect biotinylated species in the BSA‐peptide complex. The cross‐linked complex was trypsinized and then interrogated with nano‐LC–MS/MS to identify the peptide cross‐links. Cross‐linking was greatly facilitated by Bpa in the peptide, but some cross‐linking occurred at higher laser powers and high concentrations of a non‐Bpa–modified α‐factor. This was supported by experiments using GnRH, a peptide with sequence homology to α‐factor, which was likewise found to be cross‐linked to BSA by laser irradiation. Analysis of peptides in the mass spectra showed that the binding site for both α‐factor and GnRH was in the BSA pocket defined previously as the site for fatty acid binding. This model system validates the use of laser‐activation to facilitate cross‐linking of Bpa‐containing molecules to proteins. The rapid cross‐linking procedure and high performance of MS/MS to identify cross‐links provides a method to interrogate protein‐peptide interactions in a living cell in a time‐resolved manner. [ABSTRACT FROM AUTHOR]

In silico transcriptome mining is one of the most effective methods for neuropeptide discovery in crustaceans, particularly for species that are small, rare or from geographically inaccessible habitats that make obtaining the large pools of tissue needed for other peptide discovery platforms impractical. Via this approach, large peptidomes have recently been described for members of many of the higher crustacean taxa, one notable exception being the Isopoda; no peptidome has been predicted for any member of this malacostracan order. Using a publicly accessible transcriptome for the isopod Proasellus cavaticus , a subcentimeter subterranean ground water dweller, the first in silico -predicted peptidome for a member of the Isopoda is presented here. BLAST searches employing known arthropod neuropeptide pre/preprohormone queries identified 49 transcripts as encoding putative homologs within the P. cavaticus transcriptome. The proteins deduced from these transcripts allowed for the prediction of 171 distinct mature neuropeptides. The P. cavaticus peptidome includes members of the adipokinetic hormone-corazonin-like peptide, allatostatin A, allatostatin B, allatostatin C, allatotropin, bursicon α, bursicon β, CCHamide, crustacean cardioactive peptide, crustacean hyperglycemic hormone/molt-inhibiting hormone, diuretic hormone 31, eclosion hormone, elevenin, FMRFamide-like peptide, glycoprotein hormone α2, leucokinin, myosuppressin, neuroparsin, neuropeptide F, pigment dispersing hormone, pyrokinin, red pigment concentrating hormone, RYamide, short neuropeptide F, sulfakinin, tachykinin-related peptide and trissin families, as well as many linker/precursor-related sequences that may or may not represent additional bioactive molecules. Interestingly, many of the predicted P. cavaticus neuropeptides possess structures identical (or nearly so) to those previously described from members of several other malacostracan orders, i.e. , the Decapoda, Amphipoda and Euphausiacea, a finding that suggests broad phylogenetic conservation of bioactive peptide structures, and possibly functions, may exist within the Malacostraca. [ABSTRACT FROM AUTHOR]