Newton, R. P. | bioRxiv Science

bioRxiv2024

Detecting the bodys reproductive hormonal brake against tissue overgrowth: micrin/SgII-70

A new humoral factor has been detected, within a project aiming to disclose the bodys reproductive hormonal brake against tissue overgrowth, micrin ( my-crin). It is hypothesized that micrin braking, gonadal and hypothalamic, is lifted at puberty and wanes with age, bringing on prostatic enlargement and cancers. Factor purification has involved fractionation of ovine and bovine ovarian follicular fluid and blood plasma and serum, with evaluation via rat bioassays in vivo and in vitro. Analysis averse, the molecular effector provided a chemical conundrum. Evidence from mass spectrometry (MS) has been problematic, with spectra from MALDI-TOF, the only productive MS modality, confused by a target polypeptide exhibiting artefacts during processing, storage and MALDI set-up in terms of C-terminal truncation and water losses, together with N-terminal grand fragmentations and dimerizations (factor fragments doubled up). Evidence from chemical sequencing of amino acid (aa) residues was likewise baffling, but consistent with a spiralised depolymerisation within the Edman reaction chamber of a unitary polypeptide. Data decryption has overcome molecular intractability, supported by the results of tryptic digestion and epitope mapping using immunohistochemistry (IHC). The detected inhibitory factor is sleuthed to relate to secretogranin II (SgII), the neurosecretory prohormone, in the form of a secreted acidic 70-aa polypeptide derivative called here SgII-70 ( sig two seventy). The product of peptide splicing, micrin/SgII-70 is potentially an amphipathic molecular ampersand (&), with ends entwined (via salt bridging), the knotty totality compromising Edman and MALDI analyses and molecular modelling whilst conferring protease and heat resistance. The two ends of the factor are conceptualised as binding at the same time to a dimeric cellular receptor, to stoichiometric effect, providing fewer smaller cells within a counting mechanism of tissue-mass regulation. Hexapeptide mimetics simulating both ends of the hormone together have been demonstrated in different species and settings, in the cause of antiorganotrophism (tissue reduction) and reproductive modulation. Therapeutic exploitation beckons for mimetics of the bodys hormonal brake, micrin/SgII-70, in tissue overgrowth conditions such as endometriosis, PCOS, BPH and cancer, and in infertility.

immunology↗

bioRxiv2021

Hexapeptides from mammalian inhibitory hormone hunt activate and inactivate nematode reproduction

BackgroundBiopurification has been used to disclose an evolutionarily conserved inhibitory reproductive hormone involved in tissue mass determination. A (rat) bioassay-guided physicochemical fractionation using ovine materials yielded via Edman degradation a 14-residue amino acid (aa) sequence. As a 14mer synthetic peptide (EPL001) this displayed antiproliferative and reproduction-modulating activity, while representing only a part of the native polypeptide. Even more unexpectedly, a scrambled-sequence control peptide (EPL030) did likewise. MethodsReproduction has been investigated in the nematode Steinernema siamkayai, using a fermentation system supplemented with different concentrations of exogenous hexapeptides. Peptide structure-activity relationships have also been studied using prostate cancer and other mammalian cells in vitro, with peptides in solution or immobilized, and via the use of mammalian assays in vivo and through molecular modelling. ResultsReproduction increased (x3) in the entomopathogenic nematode Steinernema siamkayai after exposure to one synthetic peptide (IEPVFT), while fecundity was reduced (x0.5) after exposure to another (KLKMNG), both effects being dose-dependent. These hexamers are opposite ends of the synthetic peptide KLKMNGKNIEPVFT (EPL030). Bioactivity is unexpected as EPL030 is a control compound, based on a scrambled sequence of the test peptide MKPLTGKVKEFNNI (EPL001). EPL030 and EPL001 are both bioinformatically obscure, having no convincing matches to aa sequences in the protein databases. EPL001 has antiproliferative effects on human prostate cancer cells and rat bone marrow cells in vitro. Intracerebroventricular infusion of EPL001 in sheep was associated with elevated growth hormone in peripheral blood and reduced prolactin. The highly dissimilar EPL001 and EPL030 nonetheless have the foregoing biological effects in common in mammalian systems, while being divergently pro- and anti-fecundity respectively in the nematode Caenorhabditis elegans. Peptides up to a 20mer have also been shown to inhibit the proliferation of human cancer and other mammalian cells in vitro, with reproductive upregulation demonstrated previously in fish and frogs, as well as nematodes. EPL001 encodes the sheep neuroendocrine prohormone secretogranin II (sSgII), as deduced on the basis of immunoprecipitation using an anti-EPL001 antibody, with bespoke bioinformatics. Six sSgII residues are key to EPL001s bioactivity : MKPLTGKVKEFNNI. A stereospecific bimodular tri-residue signature is described involving simultaneous accessibility for binding of the side chains of two specific trios of amino acids, MKP & VFN. An evolutionarily conserved receptor is conceptualised having dimeric binding sites, each with ligand-matching bimodular stereocentres. The bioactivity of the 14mer control peptide EPL030 and its hexapeptide progeny is due to the fortuitous assembly of subsets of the novel hormonal motif, MKPVFN, a default reproductive and tissue-building OFF signal. NOTEPlease see the end of the paper for links to ten files of supplementary information and for an independent peer review. KEY POINTSO_LISynthetic hexapeptides have upregulated and downregulated nematode reproduction, following similar work in other nematode species and in fish and frogs. C_LIO_LIPeptides up to a 20mer have also been shown to inhibit the proliferation of human cancer and other mammalian cells in vitro and influence circulating levels of ovine pituitary hormones in vivo via intracerebroventricular peptide infusion. C_LIO_LIThe amino acid sequence of the 14mer master peptide arose from a (rat) bioassay- guided fractionation using ovine materials, whose aim was to disclose an evolutionarily conserved inhibitory reproductive hormone involved in tissue mass determination. C_LIO_LIThe bioactive peptides mimic the non-contiguous six-residue receptor-binding active face of a polypeptide hormone, which in mammals is a derivative of the secretory vesicle prohormone secretogranin II (SgII), taking the form of a novel, structually complex 70mer dubbed SgII-70. C_LIO_LIThe ability has been gained to reinforce or block a default reproductive and tissue- building OFF signal, using peptide mimetics. C_LI

physiology↗