STAT3 Pathways Contribute to ß-HCH Interference with Anticancer Tyrosine Kinase Inhibitors.

Organochlorine pesticides (OCPs) are a class of environmentally persistent and bioaccumulative pollutants. Among these, ß-hexachlorocyclohexane (ß-HCH) is a byproduct of lindane synthesis, one of the most worldwide widespread pesticides. ß-HCH cellular mechanisms inducing chemical carcinogenesis correspond to many of those inducing chemoresistance, in particular, by the activation of signal transducer and activator of transcription 3 (STAT3) signaling pathways. For this purpose, four cell lines, representative of breast, lung, prostate, and hepatocellular cancers, were treated with ß-HCH, specific tyrosine kinase inhibitors (TKIs), and a STAT3 inhibitor. All cell samples were analyzed by a viability assay, immunoblotting analysis, a wound-healing assay, and a colony formation assay. The results show that ß-HCH reduces the efficacy of TKIs. The STAT3 protein, in this context, plays a central role. In fact, by inhibiting its activity, the efficacy of the anticancer drug is restored. Furthermore, this manuscript aimed to draw the attention of the scientific and socio-healthcare community to the issue of prolonged exposure to contaminants and their impact on drug efficacy.

Regulatory role of primary cilia in oral and maxillofacial development and disease.

Primary cilia have versatile functions, such as receiving signals from the extracellular microenvironment, mediating signaling transduction, and transporting ciliary substances, in tissue and organ development and clinical disease pathogenesis. During early development (embryos within 10 weeks) in the oral and maxillofacial region, defects in the structure and function of primary cilia can result in severe craniofacial malformations. For example, mice with mutations in the cilia-related genes Kif3a and IFT88 exhibit midline expansion and cleft lip/palate, which occur due to abnormalities in the fusion of the single frontonasal prominence and maxillary prominences. In the subsequent development of the oral and maxillofacial region, we discussed the regulatory role of primary cilia in the development of the maxilla, mandible, Meckel cartilage, condylar cartilage, lip, tongue, and tooth, among others. Moreover, primary cilia are promising regulators in some oral and maxillofacial diseases, such as tumors and malocclusion. We also summarize the regulatory mechanisms of primary cilia in oral and maxillofacial development and related diseases, including their role in various signaling transduction pathways. For example, aplasia of submandibular glands in the Kif3a mutant mice is associated with a decrease in SHH signaling within the glands. This review summarizes the similarities and specificities of the role of primary cilia in tissue and organ development and disease progression in the oral and maxillofacial region, which is expected to contribute several ideas for the treatment of primary cilia-related diseases.

miR394 modulates brassinosteroid signaling to regulate hypocotyl elongation in Arabidopsis.

The interplay between microRNAs (miRNAs) and phytohormones allows plants to integrate multiple internal and external signals to optimize their survival of different environmental conditions. Here, we report that miR394 and its target gene LEAF CURLING RESPONSIVENESS (LCR), which are transcriptionally responsive to BR, participate in BR signaling to regulate hypocotyl elongation in Arabidopsis thaliana. Phenotypic analysis of various transgenic and mutant lines revealed that miR394 negatively regulates BR signaling during hypocotyl elongation, whereas LCR positively regulates this process. Genetically, miR394 functions upstream of BRASSINOSTEROID INSENSITIVE2 (BIN2), BRASSINAZOLEs RESISTANT1 (BZR1), and BRI1-EMS-SUPPRESSOR1 (BES1), but interacts with BRASSINOSTEROID INSENSITIVE1 (BRI1) and BRI1 SUPRESSOR PROTEIN (BSU1). RNA-sequencing analysis suggested that miR394 inhibits BR signaling through BIN2, as miR394 regulates a significant number of genes in common with BIN2. Additionally, miR394 increases the accumulation of BIN2 but decreases the accumulation of BZR1 and BES1, which are phosphorylated by BIN2. MiR394 also represses the transcription of PACLOBUTRAZOL RESISTANCE1/5/6 and EXPANSIN8, key genes that regulate hypocotyl elongation and are targets of BZR1/BES1. These findings reveal a new role for a miRNA in BR signaling in Arabidopsis.

Chemical proteomics approaches for protein post-translational modification studies.

The diversity and dynamics of proteins play essential roles in maintaining the basic constructions and functions of cells. The abundance of functional proteins is regulated by the transcription and translation processes, while the alternative splicing enables the same gene to generate distinct protein isoforms of different lengths. Beyond the transcriptional and translational regulations, post-translational modifications (PTMs) are able to further expand the diversity and functional scope of proteins. PTMs have been shown to make significant changes in the surface charges, structures, activation states, and interactome of proteins. Due to the functional complexity, highly dynamic nature, and low presence percentage, the study of protein PTMs remains challenging. Here we summarize and discuss the major chemical biology tools and chemical proteomics approaches to enrich and investigate the protein PTM of interest.

Transporters and phytohormones analysis reveals differential regulation of ryegrass (Lolium perenne L.) in response to cadmium and arsenic stresses.

Cadmium (Cd) and arsenic (As) are two highly toxic heavy metals and metalloids that coexist in many situations posing severe threats to plants. Our investigation was conducted to explore the different regulatory mechanisms of ryegrass (Lolium perenne L.) responding to individual and combined Cd and As stresses in hydroponics. Results showed that the ryegrass well-growth phenotype was not affected by Cd stress of 10 mg·L-1. However, As of 10 mg·L-1 caused rapid water loss, proline surge, and chlorosis in shoots, suggesting that ryegrass was highly sensitive to As. Transcriptomic analysis revealed that the transcription factor LpIRO2 mediated the upregulation of ZIP1 and YSL6 that played an important role in Cd tolerance. We found that the presence of As caused the overexpression of LpSWT12, a process potentially regulated by bHLH14, to mitigate hyperosmolarity. Indoleacetic acid (IAA) and abscisic acid (ABA) contents and expression of their signaling-related genes were significantly affected by As stress rather than Cd. We predict a regulatory network to illustrate the interaction between transporters, transcription factors, and signaling transduction, and explain the antagonism of Cd and As toxicity. This present work provides a research basis for plant protection from Cd and As pollution.

The role of kinesin family members in hepatobiliary carcinomas: from bench to bedside.

As a major component of the digestive system malignancies, tumors originating from the hepatic and biliary ducts seriously endanger public health. The kinesins (KIFs) are molecular motors that enable the microtubule-dependent intracellular trafficking necessary for mitosis and meiosis. Normally, the stability of KIFs is essential to maintain cell proliferation and genetic homeostasis. However, aberrant KIFs activity may destroy this dynamic stability, leading to uncontrolled cell division and tumor initiation. In this work, we have made an integral summarization of the specific roles of KIFs in hepatocellular and biliary duct carcinogenesis, referring to aberrant signal transduction and the potential for prognostic evaluation. Additionally, current clinical applications of KIFs-targeted inhibitors have also been discussed, including their efficacy advantages, relationship with drug sensitivity or resistance, the feasibility of combination chemotherapy or other targeted agents, as well as the corresponding clinical trials. In conclusion, the abnormally activated KIFs participate in the regulation of tumor progression via a diverse range of mechanisms and are closely associated with tumor prognosis. Meanwhile, KIFs-aimed inhibitors also carry out a promising tumor-targeted therapeutic strategy that deserves to be further investigated in hepatobiliary carcinoma (HBC).

Arabidopsis ERD15 regulated by BBX24 plays a positive role in UV-B signaling.

Ultraviolet-B (UV-B, 280-315 nm) is a minor component of solar radiation, but it has a major regulatory impact on plant growth and development. Solar UV-B regulates numerous aspects of plant metabolism, morphology and physiology through altering the expression of hundreds of genes. EARLY RESPONSIVE TO DEHYDRATION 15 (ERD15) is a drought-induced rapid response gene, formerly known as a negative regulator of the abscisic acid (ABA) signaling pathway. It is unclear whether ERD15 is involved in UV-B-induced photomorphogenesis. Previously, we reported that the BBX24 transcriptional factor negatively regulated UV-B signaling. In the present study, we identified that ERD15 is involved in UV-B photomorphogenesis as a positive regulator at phenotypic, physiological and molecular levels. Our results indicated that ERD15 expression is suppressed by UV-B, inhibited the elongation of Arabidopsis hypocotyls in a UV-B-dependent manner, promoted the expression of related UV-B signaling genes and increased the total antioxidant capacity of Arabidopsis under UV-B. Genetic hybridization results show that ERD15 acts downstream of BBX24, and BBX24 protein mediated the expression of ERD15 by binding to its promoter. Thus, ERD15 is a novel positive regulator of the UV-B signaling pathway, which is downstream of BBX24 and regulated by BBX24 protein to participate in UV-B photomorphogenesis.

Structure-based design of non-hypertrophic apelin receptor modulator.

Apelin is a key hormone in cardiovascular homeostasis that activates the apelin receptor (APLNR), which is regarded as a promising therapeutic target for cardiovascular disease. However, adverse effects through the ß-arrestin pathway limit its pharmacological use. Here, we report cryoelectron microscopy (cryo-EM) structures of APLNR-Gi1 complexes bound to three agonists with divergent signaling profiles. Combined with functional assays, we have identified "twin hotspots" in APLNR as key determinants for signaling bias, guiding the rational design of two exclusive G-protein-biased agonists WN353 and WN561. Cryo-EM structures of WN353- and WN561-stimulated APLNR-G protein complexes further confirm that the designed ligands adopt the desired poses. Pathophysiological experiments have provided evidence that WN561 demonstrates superior therapeutic effects against cardiac hypertrophy and reduced adverse effects compared with the established APLNR agonists. In summary, our designed APLNR modulator may facilitate the development of next-generation cardiovascular medications.

A Hierarchical Mechanotransduction System: From Macro to Micro.

Mechanotransduction is a strictly regulated process whereby mechanical stimuli, including mechanical forces and properties, are sensed and translated into biochemical signals. Increasing data demonstrate that mechanotransduction is crucial for regulating macroscopic and microscopic dynamics and functionalities. However, the actions and mechanisms of mechanotransduction across multiple hierarchies, from molecules, subcellular structures, cells, tissues/organs, to the whole-body level, have not been yet comprehensively documented. Herein, the biological roles and operational mechanisms of mechanotransduction from macro to micro are revisited, with a focus on the orchestrations across diverse hierarchies. The implications, applications, and challenges of mechanotransduction in human diseases are also summarized and discussed. Together, this knowledge from a hierarchical perspective has the potential to refresh insights into mechanotransduction regulation and disease pathogenesis and therapy, and ultimately revolutionize the prevention, diagnosis, and treatment of human diseases.

Comparative Physiological and Transcriptome Analysis of Crossostephium chinense Reveals Its Molecular Mechanisms of Salt Tolerance.

Crossostephium chinense is a wild species with strong salt tolerance that has great potential to improve the salt tolerance of cultivated chrysanthemums. Conversely, the unique salt-tolerant molecular mechanisms of Cr. chinense are still unclear. This study performed a comparative physiological and transcriptome analysis of Cr. chinense, Chrysanthemum lavandulifolium, and three hybrids to investigate the salt-tolerant molecular mechanisms of Cr. chinense. The physiological results showed that Cr. chinense maintained higher superoxide dismutase (SOD) activity, alleviating oxidative damage to the membrane. KEGG enrichment analysis showed that plant hormone signaling transduction and the MAPK signaling pathway were mostly enriched in Cr. chinense and hybrids under salt stress. Further weighted gene co-expression network analysis (WGCNA) of DEGs suggested that abscisic acid (ABA) signaling transduction may play a significant role in the salt-tolerant mechanisms of Cr. chinense and hybrids. The tissue-specific expression patterns of the candidate genes related to ABA signaling transduction and the MAPK signaling pathway indicate that genes related to ABA signaling transduction demonstrated significant expression levels under salt stress. This study offers important insights into exploring the underlying salt-tolerant mechanisms of Cr. chinense mediated by ABA signaling transduction and broadens our understanding of the breeding strategies for developing salt-tolerant cultivars utilizing salt-tolerant chrysanthemum germplasms.

Snapshot of the cannabinoid receptor 1-arrestin complex unravels the biased signaling mechanism.

Cannabis activates the cannabinoid receptor 1 (CB1), which elicits analgesic and emotion regulation benefits, along with adverse effects, via Gi and ß-arrestin signaling pathways. However, the lack of understanding of the mechanism of ß-arrestin-1 (ßarr1) coupling and signaling bias has hindered drug development targeting CB1. Here, we present the high-resolution cryo-electron microscopy structure of CB1-ßarr1 complex bound to the synthetic cannabinoid MDMB-Fubinaca (FUB), revealing notable differences in the transducer pocket and ligand-binding site compared with the Gi protein complex. ßarr1 occupies a wider transducer pocket promoting substantial outward movement of the TM6 and distinctive twin toggle switch rearrangements, whereas FUB adopts a different pose, inserting more deeply than the Gi-coupled state, suggesting the allosteric correlation between the orthosteric binding pocket and the partner protein site. Taken together, our findings unravel the molecular mechanism of signaling bias toward CB1, facilitating the development of CB1 agonists.

Development of High-Loading Trastuzumab PLGA Nanoparticles: A Powerful Tool Against HER2 Positive Breast Cancer Cells.

Background: Trastuzumab, a therapeutic monoclonal antibody directed against HER2, is routinely used to treat HER2-positive breast cancer with a good response rate. However, concerns have arisen in the clinical practice due to adverse side effects. One way to overcome these limitations is to encapsulate trastuzumab in nanoparticles to improve cytotoxic activity, increase intracellular drug concentrations, escape the immune system and avoid systemic degradation of the drug in vivo. Methods: A double emulsion method was used to encapsulate trastuzumab into poly(lactic-co-glycolic) nanoparticles, effective for their biocompatibility and biodegradability. These nanocarriers, hereafter referred to as TZPs, were characterised in terms of size, homogeneity, zeta potential and tested for their stability and drug release kinetics. Finally, the TZPs cytotoxicity was assessed in vitro on the HER2 positive SKBR3 breast cancer cell line and compared to free trastuzumab. Results: The TZPs were stable, homogeneous in size, with a reduced zeta potential. They showed higher encapsulation efficiency and drug loading, a prolonged trastuzumab release kinetics that retained its physicochemical properties and functionality. TZPs showed a stronger cytotoxicity and increased apoptosis than similar doses of free trastuzumab in the cell line analysed. Confocal microscopy and flow cytometry assessed TZPs and trastuzumab cellular uptake while Western blot evaluated downstream signalling, overall HER2 content and shedding. Conclusion: TZPs exert more robust effects than free trastuzumab via a dual mode of action: TZPs are taken up by cells through an endocytosis mechanism and release the drug intracellularly for longer time. Additionally, the TZPs that remain in the extracellular space release trastuzumab which binds to the cognate receptor and impairs downstream signalling. This is the sole modality used by free trastuzumab. Remarkably, half dose of TZPs is as efficacious as the highest dose of free drug supporting their possible use for drug delivery in vivo.

The deubiquitinase USP10 mediates crosstalk between the LKB1/AMPK axis and Wnt/ß-catenin signaling in cancer.

The liver kinase B1 (LKB1)/AMP-activated protein kinase (AMPK) axis pivotally controls cell metabolism and suppresses abnormal growth in various cancers. Wnt/ß-catenin is a frequently dysregulated signaling pathway that drives oncogenesis. Here, we discovered a crosstalk mechanism between the LKB1/AMPK axis and Wnt/ß-catenin signaling. Activated AMPK phosphorylates the deubiquitinase USP10 to potentiate the deubiquitination and stabilization of the key scaffold protein Axin1. This phosphorylation also strengthens the binding between USP10 and ß-catenin and supports the phase transition of ß-catenin. Both processes suppress Wnt/ß-catenin amplitude in parallel and inhibit colorectal cancer growth in a clinically relevant manner. Collectively, we established a crosstalk route by which LKB1/AMPK regulates Wnt/ß-catenin signaling in cancer. USP10 acts as the hub in this process, thus enabling LKB1/AMPK to suppress tumor growth via regulation of both metabolism and cell proliferation.

PDZ and LIM Domain-Encoding Genes: Their Role in Cancer Development.

PDZ-LIM family proteins (PDLIMs) are a kind of scaffolding proteins that contain PDZ and LIM interaction domains. As protein-protein interacting molecules, PDZ and LIM domains function as scaffolds to bind to a variety of proteins. The PDLIMs are composed of evolutionarily conserved proteins found throughout different species. They can participate in cell signal transduction by mediating the interaction of signal molecules. They are involved in many important physiological processes, such as cell differentiation, proliferation, migration, and the maintenance of cellular structural integrity. Studies have shown that dysregulation of the PDLIMs leads to tumor formation and development. In this paper, we review and integrate the current knowledge on PDLIMs. The structure and function of the PDZ and LIM structural domains and the role of the PDLIMs in tumor development are described.

A clathrin-related protein FaRRP1/SCD2 integrates ABA trafficking and signaling to regulate strawberry fruit ripening.

Abscisic acid (ABA) is a critical regulator for nonclimacteric fruit ripening such as in the model plant of strawberry (Fragaria × ananassa). Although FaRRP1 is proposed to participate in clathrin-mediated endocytosis of ABA, its action molecular mechanisms in ABA signaling are not fully understood. Here, using our isolated FaRRP1 (ripening-regulation protein) and candidate ABA receptor FaPYL2 and FaABAR from strawberry fruit, a series of silico and molecular interaction analyses demonstrate that they all bind to ABA, and FaRRP1 binds both FaPYL2 and FaABAR; by contrast, the binding affinity of FaRRP1 to FaPYL2 is relatively higher. Interestingly, the binding of FaRRP1 to FaPYL2 and FaABAR affects the perception affinity to ABA. Furthermore, exogenous ABA application and FaRRP1 transgenic analyses confirm that FaRRP1 participates in clathrin-mediated endocytosis and vesicle transport. Importantly, FaRRP1, FaPYL2, and FaABAR all trigger the initiation of strawberry fruit ripening at physiological and molecular levels. In conclusion, FaRRP1 not only binds to ABA but also affects the binding affinity of FaPYL2 and FaABAR to ABA, thus promoting strawberry fruit ripening. Our findings provide novel insights into the role of FaRRP1 in ABA trafficking and signaling, at least in strawberry, a model plant for nonclimacteric fruit ripening.

A big picture of the mitochondria-mediated signals: From mitochondria to organism.

Mitochondria, well-known for years as the powerhouse and biosynthetic center of the cell, are dynamic signaling organelles beyond their energy production and biosynthesis functions. The metabolic functions of mitochondria, playing an important role in various biological events both in physiological and stress conditions, transform them into important cellular stress sensors. Mitochondria constantly communicate with the rest of the cell and even from other cells to the organism, transmitting stress signals including oxidative and reductive stress or adaptive signals such as mitohormesis. Mitochondrial signal transduction has a vital function in regulating integrity of human genome, organelles, cells, and ultimately organism.

Dexmedetomidine Alleviates Abdominal Aortic Aneurysm by Activating Autophagy Via AMPK/mTOR Pathway.

BACKGROUND: Abdominal aortic aneurysms (AAA) are a critical global health issue with increasing prevalence. Dexmedetomidine (DEX) is a highly selective α2-adrenoceptor agonist that has previously been shown to play a protective role in AAA. Nevertheless, the mechanisms underlying its protection effect remain not fully understood. METHODS: A rat AAA model was established via intra-aortic porcine pancreatic elastase perfusion with or without DEX administration. The abdominal aortic diameters of rats were measured. Hematoxylin-eosin and Elastica van Gieson staining were conducted for histopathological observation. TUNEL and immunofluorescence staining were utilized to detect cell apoptosis and α-SMA/LC3 expression in the abdominal aortas. Protein levels were determined using western blotting. RESULTS: DEX administration repressed the dilation of aortas, alleviated pathological damage and cell apoptosis, and suppressed phenotype switching of vascular smooth muscle cells (VSMCs). Moreover, DEX activated autophagy and regulated the AMP-activated protein kinase/mammalian target of the rapamycin (AMPK/mTOR) signaling pathway in AAA rats. Administration of the AMPK inhibitor attenuated the DEX-mediated ameliorative effects on AAA in rats. CONCLUSION: DEX ameliorates AAA in rat models by activating autophagy via the AMPK/mTOR pathway.

Integrated proteomics and phosphoproteomics profiling reveals the cardioprotective mechanism of bioactive compounds derived from Salvia miltiorrhiza Burge.

BACKGROUND: Natural products are an important source for discovering novel drugs due to their various pharmacological activities. Salvia miltiorrhiza Burge (Danshen) has been shown to have promising therapeutic potential in the management of heart diseases, making it a candidate for cardiovascular drug discovery. Currently, there is limited quantitative analysis of the phosphorylation levels of Danshen-derived natural products on a proteome-wide, which may bias the study of their mechanisms of action. PURPOSE: This study aimed to evaluate the global signaling perturbation induced by Danshen-derived bioactive compounds and their potential relationship with myocardial ischemia/reperfusion (IR) injury therapy. STUDY DESIGN: We employed quantitative proteome and phosphoproteome analysis to identify dysregulated signaling in IR injury hearts from mice. We compared changes induced by Danshen-derived compounds based on IR-associated phospho-events, using an integrative approach that maps relative abundance of proteins and phosphorylation sites. METHODS: Isobaric chemical tandem mass tags (TMT) labeled multiplexing strategy was used to generate unbiased quantitative proteomics and phosphoproteomics data. Highly accurate and precise TMT quantitation was performed using the Orbitrap Fusion Tribrid Mass Spectrometer with synchronous precursor selection MS3 detection mode. Mass spectrometric raw files were analyzed with MaxQuant (2.0.1.0) and statistical and bioinformatics analysis was conducted with Perseus (1.6.15). RESULTS: We quantified 3661 proteins and over 11,000 phosphosites in impaired heart tissue of the IR mice model, expanding our knowledge of signaling pathways and other biological processes disrupted in IR injury. Next, 1548 and 5545 differently expressed proteins and phosphosites were identified by quantifying the proteome and phosphoproteome of H9c2 cells treated by five Danshen bioactive compounds respectively. Results revealed the vast differences in abilities of five Danshen-derived bioactive compounds to regulate phosphorylation modifications in cardiomyocytes, with dihydrotanshinone I (DHT) showing potential for protecting against IR injury by modulating the AMPK/mTOR signaling pathway. CONCLUSIONS: This study provides a new strategy for analyzing drug/natural product-regulated phosphorylation modification levels on a proteome-wide scale, leading to a better understanding of cell signaling pathways and downstream phenotypic responses.

Detecting and measuring of GPCR signaling - comparison of human induced pluripotent stem cells and immortal cell lines.

G protein-coupled receptors (GPCRs) are a large family of transmembrane proteins that play a major role in many physiological processes, and thus GPCR-targeted drug development has been widely promoted. Although research findings generated in immortal cell lines have contributed to the advancement of the GPCR field, the homogenous genetic backgrounds, and the overexpression of GPCRs in these cell lines make it difficult to correlate the results with clinical patients. Human induced pluripotent stem cells (hiPSCs) have the potential to overcome these limitations, because they contain patient specific genetic information and can differentiate into numerous cell types. To detect GPCRs in hiPSCs, highly selective labeling and sensitive imaging techniques are required. This review summarizes existing resonance energy transfer and protein complementation assay technologies, as well as existing and new labeling methods. The difficulties of extending existing detection methods to hiPSCs are discussed, as well as the potential of hiPSCs to expand GPCR research towards personalized medicine.

Identification of GnRH-like peptide and its potential signaling pathway involved in the oocyte meiotic maturation in the Chinese mitten crab, Eriocheir sinensis.

Gonadotropin-releasing hormone (GnRH) plays a pivotal role in reproductive regulation in vertebrates. However, GnRH was rarely isolated and its function remains poorly characterized in invertebrates. The existence of GnRH in ecdysozoa has been controversial for a long. Here, we isolated and identified two GnRH-like peptides from brain tissues in Eriocheir sinensis. Immunolocalization showed that the presence of EsGnRH-like peptide in brain, ovary and hepatopancreas. Synthetic EsGnRH-like peptides can induce germinal vesicle breakdown (GVBD) of oocyte. Similar to vertebrates, ovarian transcriptomic analysis revealed a GnRH signaling pathway in the crab, in which most genes exhibited dramatically high expression at GVBD. RNAi knockdown of EsGnRHR suppressed the expression of most genes in the pathway. Co-transfection of the expression plasmid for EsGnRHR with reporter plasmid bearing CRE-luc or SRE-luc response element into 293T cells showed that EsGnRHR transduces its signal via cAMP and Ca2+ signaling transduction pathways. In vitro incubation of the crab oocyte with EsGnRH-like peptide confirmed the cAMP-PKA cascade and Ca2+ mobilization signaling cascade but lack of a PKC cascade. Our data present the first direct evidence of the existence of GnRH-like peptides in the crab and demonstrated its conserved role in the oocyte meiotic maturation as a primitive neurohormone.