Identification of novel peptide hormones in the human proteome by hidden Markov model screening

Abstract

Peptide hormones are small, processed, and secreted peptides that signal via membrane receptors and play critical roles in normal and pathological physiology. The search for novel peptide hormones has been hampered by their small size, low or restricted expression, and lack of sequence similarity. To overcome these difficulties, we developed a bioinformatics search tool based on the hidden Markov model formalism that uses several peptide hormone sequence features to estimate the likelihood that a protein contains a processed and secreted peptide of this class. Application of this tool to an alignment of mammalian proteomes ranked 90% of known peptide hormones among the top 300 proteins. An analysis of the top scoring hypothetical and poorly annotated human proteins identified two novel candidate peptide hormones. Biochemical analysis of the two candidates, which we called spexin and augurin, showed that both were localized to secretory granules in a transfected pancreatic cell line and were recovered from the cell supernatant. Spexin was expressed in the submucosal layer of the mouse esophagus and stomach, and a predicted peptide from the spexin precursor induced muscle contraction in a rat stomach explant assay. Augurin was specifically expressed in mouse endocrine tissues, including pituitary and adrenal gland, choroid plexus, and the atrio-ventricular node of the heart. Our findings demonstrate the utility of a bioinformatics approach to identify novel biologically active peptides. Peptide hormones and their receptors are important diagnostic and therapeutic targets, and our results suggest that spexin and augurin are novel peptide hormones likely to be involved in physiological homeostasis.

Global view of the evolution and diversity of metazoan neuropeptide signaling

“…pNPs have a signal peptide (SP) and enter the secretory apparatus where dedicated proteases cleave them at mono-or dibasic cleavage sites (12) and where the maturing peptides are often further modified (13). pNPs are ubiquitous in eumetazoans, and genomics and mass spectrometry revealed the full neuropeptide repertoire of several species (14)(15)(16)(17)(18)(19)(20). Given their wide occurrence in metazoans, and importance in neuronal regulation, a global pNP phylogeny would further our knowledge of nervous system evolution.…”

Augurin stimulates the hypothalamo‐pituitary‐adrenal axis via the release of corticotrophin‐releasing factor in rats

“…ECRG4 protein contains one canonical dibasic prohormone cleavage site corresponding to residues Lys69 and Arg70. Cleavage at this site has been demonstrated in vitro following transfection of a pancreatic cell line with the c2orf40 gene in which the Flag antigen sequence was inserted between Arg70 and Gln71 (Mirabeau et al, 2007). Western blotting of supernatant from Flag augurin-transfected cells revealed two immunoreactive bands consistent with secretion of an Arg70 cleaved 10 kDa peptide (augurin 71-148) and a smaller 8 kDa peptide.…”

“…Western blotting of supernatant from Flag augurin-transfected cells revealed two immunoreactive bands consistent with secretion of an Arg70 cleaved 10 kDa peptide (augurin 71-148) and a smaller 8 kDa peptide. It is suggested the 8 kDa peptide results from a second, unknown, C-terminal non-canonical cleavage site (Mirabeau et al, 2007).…”

“…Augurin is a recently identified secreted peptide of unknown function encoded by the gene c2orf40 (Mirabeau et al, 2007), a potential tumour suppressor gene in human oesophageal epithelium (Su et al, 1998). Down-regulation of c2orf40 expression due to hypermethylation can occur in human oesophageal squamous cell carcinoma and may be an independent prognostic factor for poor survival (Yue et al, 2003).…”

“…ECRG4 protein has characteristic features of a preprohormone; it contains a canonical signal peptide sequence and at least one basic residue, prohormone cleavage, site. Augurin is produced by post-translational processing of ECRG4 protein (Mirabeau et al, 2007).…”