Ghrelin axis genes, peptides and receptors : recent findings and future challenges

The ghrelin axis consists of the gene products of the ghrelin gene (GHRL), and their receptors, including the classical ghrelin receptor GHSR. While it is well-known that the ghrelin gene encodes the 28 amino acid ghrelin peptide hormone, it is now also clear that the locus encodes a range of other bioactive molecules, including novel peptides and non-coding RNAs. For many of these molecules, the physiological functions and cognate receptor(s) remain to be determined. Emerging research techniques, including proteogenomics, are likely to reveal further ghrelin axis-derived molecules. Studies of the role of ghrelin axis genes, peptides and receptors, therefore, promises to be a fruitful area of basic and clinical research in years to come

Revised genomic structure of the human ghrelin gene and identification of novel exons, alternative splice variants and natural antisense transcripts

Background Ghrelin is a multifunctional peptide hormone expressed in a range of normal tissues and pathologies. It has been reported that the human ghrelin gene consists of five exons which span 5 kb of genomic DNA on chromosome 3 and includes a 20 bp non-coding first exon (20 bp exon 0). The availability of bioinformatic tools enabling comparative analysis and the finalisation of the human genome prompted us to re-examine the genomic structure of the ghrelin locus. Results We have demonstrated the presence of an additional novel exon (exon -1) and 5' extensions to exon 0 and 1 using comparative in silico analysis and have demonstrated their existence experimentally using RT-PCR and 5' RACE. A revised exon-intron structure demonstrates that the human ghrelin gene spans 7.2 kb and consists of six rather than five exons. Several ghrelin gene-derived splice forms were detected in a range of human tissues and cell lines. We have demonstrated ghrelin gene-derived mRNA transcripts that do not code for ghrelin, but instead may encode the C-terminal region of full-length preproghrelin (C-ghrelin, which contains the coding region for obestatin) and a transcript encoding obestatin-only. Splice variants that differed in their 5' untranslated regions were also found, suggesting a role of these regions in the post-transcriptional regulation of preproghrelin translation. Finally, several natural antisense transcripts, termed ghrelinOS (ghrelin opposite strand) transcripts, were demonstrated via orientation-specific RT-PCR, 5' RACE and in silico analysis of ESTs and cloned amplicons. Conclusion The sense and antisense alternative transcripts demonstrated in this study may function as non-coding regulatory RNA, or code for novel protein isoforms. This is the first demonstration of putative obestatin and C-ghrelin specific transcripts and these findings suggest that these ghrelin gene-derived peptides may also be produced independently of preproghrelin. This study reveals several novel aspects of the ghrelin gene and suggests that the ghrelin locus is far more complex than previously recognised

Complex organisation and structure of the ghrelin antisense strand gene GHRLOS, a candidate non-coding RNA gene

<p>Abstract</p> <p>Background</p> <p>The peptide hormone ghrelin has many important physiological and pathophysiological roles, including the stimulation of growth hormone (GH) release, appetite regulation, gut motility and proliferation of cancer cells. We previously identified a gene on the opposite strand of the ghrelin gene, ghrelinOS (<it>GHRLOS</it>), which spans the promoter and untranslated regions of the ghrelin gene (<it>GHRL</it>). Here we further characterise <it>GHRLOS</it>.</p> <p>Results</p> <p>We have described <it>GHRLOS </it>mRNA isoforms that extend over 1.4 kb of the promoter region and 106 nucleotides of exon 4 of the ghrelin gene, <it>GHRL</it>. These <it>GHRLOS </it>transcripts initiate 4.8 kb downstream of the terminal exon 4 of <it>GHRL </it>and are present in the 3' untranslated exon of the adjacent gene <it>TATDN2 </it>(TatD DNase domain containing 2). Interestingly, we have also identified a putative non-coding <it>TATDN2-GHRLOS </it>chimaeric transcript, indicating that <it>GHRLOS </it>RNA biogenesis is extremely complex. Moreover, we have discovered that the 3' region of <it>GHRLOS </it>is also antisense, in a tail-to-tail fashion to a novel terminal exon of the neighbouring <it>SEC13 </it>gene, which is important in protein transport. Sequence analyses revealed that <it>GHRLOS </it>is riddled with stop codons, and that there is little nucleotide and amino-acid sequence conservation of the <it>GHRLOS </it>gene between vertebrates. The gene spans 44 kb on 3p25.3, is extensively spliced and harbours multiple variable exons. We have also investigated the expression of <it>GHRLOS </it>and found evidence of differential tissue expression. It is highly expressed in tissues which are emerging as major sites of non-coding RNA expression (the thymus, brain, and testis), as well as in the ovary and uterus. In contrast, very low levels were found in the stomach where sense, <it>GHRL </it>derived RNAs are highly expressed.</p> <p>Conclusion</p> <p><it>GHRLOS </it>RNA transcripts display several distinctive features of non-coding (ncRNA) genes, including 5' capping, polyadenylation, extensive splicing and short open reading frames. The gene is also non-conserved, with differential and tissue-restricted expression. The overlapping genomic arrangement of <it>GHRLOS </it>with the ghrelin gene indicates that it is likely to have interesting regulatory and functional roles in the ghrelin axis.</p

The proximal first exon architecture of the murine ghrelin gene is highly similar to its human orthologue

BACKGROUND: The murine ghrelin gene (Ghrl), originally sequenced from stomach tissue, contains five exons and a single transcription start site in a short, 19 bp first exon (exon 0). We recently isolated several novel first exons of the human ghrelin gene and found evidence of a complex transcriptional repertoire. In this report, we examined the 5' exons of the murine ghrelin orthologue in a range of tissues using 5' RACE. -----FINDINGS: 5' RACE revealed two transcription start sites (TSSs) in exon 0 and four TSSs in intron 0, which correspond to 5' extensions of exon 1. Using quantitative, real-time RT-PCR (qRT-PCR), we demonstrated that extended exon 1 containing Ghrl transcripts are largely confined to the spleen, adrenal gland, stomach, and skin. -----CONCLUSION: We demonstrate that multiple transcription start sites are present in exon 0 and an extended exon 1 of the murine ghrelin gene, similar to the proximal first exon organisation of its human orthologue. The identification of several transcription start sites in intron 0 of mouse ghrelin (resulting in an extension of exon 1) raises the possibility that developmental-, cell- and tissue-specific Ghrl mRNA species are created by employing alternative promoters and further studies of the murine ghrelin gene are warranted

The long non-coding RNA GHSROS reprograms prostate cancer cell lines toward a more aggressive phenotype

It is now appreciated that long non-coding RNAs (lncRNAs) are important players in orchestrating cancer progression. In this study we characterized GHSROS, a human lncRNA gene on the opposite DNA strand (antisense) to the ghrelin receptor gene, in prostate cancer. The lncRNA was upregulated by prostate tumors from different clinical datasets. Transcriptome data revealed that GHSROS alters the expression of cancer-associated genes. Functional analyses in vitro showed that GHSROS mediates tumor growth, migration and survival, and resistance to the cytotoxic drug docetaxel. Increased cellular proliferation of GHSROS-overexpressing PC3, DU145, and LNCaP prostate cancer cell lines in vitro was recapitulated in a subcutaneous xenograft model. Conversely, in vitro antisense oligonucleotide inhibition of the lncRNA reciprocally regulated cell growth and migration, and gene expression. Notably, GHSROS modulates the expression of PPP2R2C, the loss of which may drive androgen receptor pathway-independent prostate tumor progression in a subset of prostate cancers. Collectively, our findings suggest that GHSROS can reprogram prostate cancer cells toward a more aggressive phenotype and that this lncRNA may represent a potential therapeutic target

A systemic secondary vessel system is present in the teleost fish Tandarius tandanus and absent in the elasmobranchs Carcharhinus melanopterus and Rhinobatos typus and in the dipnoan Neoceratodus forsteri

A system of secondary vessels emerging from the primary vessels as numerous coiled capillaries has been described in numerous teleost and holost fishes. The systemic secondary vessels of the teleost Tandanus tandanus are typical of this system and are described in this study. The existence of a secondary vessel system has been postulated in the elasmobranch group. No secondary vessel origins, as seen in the teleosts, are present in the elasmobranchs Rhinobatos typus and Carcharhinus melanopterus. Vessels with a similar distribution to secondary arteries are observed but these are venous rather than arterial in nature and do not connect with the primary arteries. Like the secondary veins in teleosts, the cutaneous veins in R. typus contain blood with a low haematocrit. There is no morphological evidence for a secondary vessel system in the dipnoan Neoceratodus forsteri.</p