Analysis of the human guanylin gene and the processing and cellular localization of the peptide.

Hill, O.; Kühn, Michaela; Zucht, Hans-Dieter; Cetin, Yalcin; Kulaksiz, Hasan; Adermann, Knut; Klöck, Gerd; Rechkemmer, Gerhard; Forssmann, Wolf‐Georg; Mägert, Hans-Jürgen

doi:10.1073/pnas.92.6.2046

Cited by 49 publications

(28 citation statements)

References 39 publications

(58 reference statements)

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“…This is in clear contrast to our results, where maximum receptor expression is detected in the colonic crypts. In the human intestine, a single report suggested that guanylin was expressed in the epithelium of the crypts of Leiberkühn in the secretory mucosa [Hill et al, 1995] and we do detect receptor expression in these cells in the rat.…”

Section: Discussionmentioning

confidence: 88%

Epitope conservation and immunohistochemical localization of the guanylin/stable toxin peptide receptor, guanylyl cyclase C

1997

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The heat-stable enterotoxins (ST) are a family of cysteine-rich low-molecular weight peptides produced by pathogenic bacteria, and are one of the major causes of watery diarrhea all over the world. These toxins mediate their action by binding to an intestinal cell surface receptor that is a membrane-associated guanylyl cyclase (GCC). This receptor also serves as the receptor for the recently characterised endogenous ligand, guanylin. We have expressed various domains of the receptor in Escherichia coli and used purified proteins for the generation of both polyclonal and monoclonal antibodies. While polyclonal antibodies were able to partially inhibit ST binding to the native receptor present in the T84 human colonic cell line, GCC:B10 monoclonal antibody did not interfere with ligand binding. Western blot analysis, using membranes prepared from human colonic T84 cells, detected two bands of size 160 and 140 kDa, representing alternately glycosylated forms of the receptor. Using the recombinant proteins, we could map the epitope of GCC:B10 monoclonal antibody to the intracellular domain of the receptor. We used the antibody to localize the receptor throughout the rat intestine, and in the porcine and bonnet monkey colon. We could detect receptor expression in the villus and the crypts of the duodenum, jejunum, ileum, and caecum, and in the crypts of the colon. Receptor expression was observed in cells that had earlier been shown to express cGMP-dependent kinase, but not the cystic fibrosis transmembrane regulator, a known downstream target of cGMP/G-kinase, which suggests that GCC/ cGMP could regulate additional cellular signal transduction machinery.

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Section: Discussionmentioning

confidence: 88%

Epitope conservation and immunohistochemical localization of the guanylin/stable toxin peptide receptor, guanylyl cyclase C

1997

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“…A human genomic library in phage (Stratagene) was screened as described (2) by using a partial 269-bp, HCC-1-specific cDNA fragment (1) as a probe. Three independent clones were isolated, and the SstI restriction fragments of one of them were subcloned into pBSK ϩ .…”

Section: Methodsmentioning

confidence: 99%

HCC-2, a human chemokine: Gene structure, expression pattern, and biological activity

Pardigol

Forssmann

Zucht

et al. 1998

Proc. Natl. Acad. Sci. U.S.A.

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Cloning and sequencing of the upstream region of the gene of the CC chemokine HCC-1 led to the discovery of an adjacent gene coding for a CC chemokine that was named ''HCC-2.'' The two genes are separated by 12-kbp and reside in a head-to-tail orientation on chromosome 17. At variance with the genes for HCC-1 and other human CC chemokines, which have a three-exon-two-intron structure, the HCC-2 gene consists of four exons and three introns. Expression of HCC-2 and HCC-1 as studied by Northern analysis revealed, in addition to the regular, monocistronic mRNAs, a common, bicistronic transcript. In contrast to HCC-1, which is expressed constitutively in numerous human tissues, HCC-2 is expressed only in the gut and the liver. HCC-2 shares significant sequence homology with CK␤8 and the murine chemokines C10, CCF18͞MRP-2, and macrophage inf lammatory protein 1␥, which all contain six instead of four conserved cysteines. The two additional cysteines of HCC-2 form a third disulfide bond, which anchors the COOHterminal domain to the core of the molecule. Highly purified recombinant HCC-2 was tested on neutrophils, eosinophils, monocytes, and lymphocytes and was found to exhibit marked functional similarities to macrophage inf lammatory protein 1␣. It is a potent chemoattractant and inducer of enzyme release in monocytes and a moderately active attractant for eosinophils. Desensitization studies indicate that HCC-2 acts mainly via CC chemokine receptor CCR1.We recently have isolated and characterized a new human CC chemokine, HCC-1 (1), which is structurally similar to macrophage inflammatory protein (MIP)-1␣ (46% amino acid identity) and occurs in high concentrations in human plasma like MIP-1␥ in murine blood. Northern blot analysis revealed that, in contrast to other chemokines, HCC-1 is expressed constitutively at high levels in numerous human tissues. We further showed that HCC-1 interacts with receptors that also recognize MIP-1␣ and RANTES and stimulates the proliferation of CD34 ϩ myeloid progenitor cells in vitro. In the present paper, we report the discovery and characterization of a CC chemokine, HCC-2, that arises from a gene that is arranged in tandem with the gene of HCC-1 on chromosome 17. Mono-as well as bicistronic transcripts of the two genes were detected. Recombinant HCC-2 was expressed and shown to act mainly on monocytes and eosinophils in a similar manner as MIP-1␣. MATERIALS AND METHODSCell Culture. Cell lines (T84, HUH-7) were purchased from the American Type Culture Collection and maintained in DMEM supplemented with 10% fetal calf serum.Oligonucleotides. The oligo-deoxyribonucleotides used were synthesized chemically (Perkin-Elmer) and are listed in Table 1.Gene Cloning and Characterization. A human genomic library in phage (Stratagene) was screened as described (2) by using a partial 269-bp, HCC-1-specific cDNA fragment (1) as a probe. Three independent clones were isolated, and the SstI restriction fragments of one of them were subcloned into pBSK ϩ . Both strands of the HCC-1 and HC...

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“…Northern blot and immunohistochemical analysis have revealed that guanylin occurs in the mucosa of the mammalian intestine (see reviews [4] and [5]). The peptide is probably released from entero-endocrine cells [6,7] into the intestinal lumen to activate the GC-C and consequently the chloride transport in enterocytes by means of a paracrine interaction [5]. However, several data indicate that the guanylin/GC-C system is not exclusively confined to the intestine.…”

Section: Introductionmentioning

confidence: 99%

GCAP‐II: Isolation and characterization of the circulating form of human uroguanylin

Hess¹,

Kühn²,

Schulz‐Knappe³

et al. 1995

FEBS Letters

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The systematic isolation of circulating regulatory peptides which generate cGMP as second messenger resulted in the identification of a novel member of the gnanylin family. In the present study we describe the purification and amino acid sequence of a new guanylate cyclase C activating peptide (GCAP-II). GCAP-II contains 24 amino acids in the following sequence: FKTLRTIANDDCELCVNVACTGCL. Its molecular mass is 2597.7 Da. The 16 C-terminal amino acids are identical to uroguanylin from human urine. Native and synthetic GCAP-II activate GC-C, the specific gnanylate cyclase receptor, of cultured human colon carcinoma (T84) cells. GCAP-II stimulates chloride secretion in isolated human intestinal mucosa mediated by intracellular cGMP increase. GCAP-II specific antibodies were used to localize the peptide by immunohistochemistry in entero-endocrine cells of the colonic mucosa. Key words':Guanylate cyclase activating peptide II; Uroguanylin; Guanylin; cGMP; Intestinal chloride secretion; Entero-endocrine cell intestinal tissues, such as kidney, liver, reproductive tract, adrenals, airway epithelia, and pancreas [4,5]. In addition, a larger molecular form of this peptide, namely guanylin-22-115, circulates as a bioactive peptide in human blood, suggesting that this peptide regulates the function of different target organs by an endocrine interaction [8].Uroguanylin, a second ligand of the GC-C receptor, has been recently purified from human and opossum urine [9,10]. The structural homology and the similar biological activity of guanylin and uroguanylin suggest that they are members of a peptide family, the main function of which is the activation of GC-C in the intestine and in other tissues. With regard to putative endocrine interactions of this system we initiated a systematic search for further endogenous activators of GC-C circulating in human blood. Here we report the isolation, the biochemical and functional characterization, and the immunohistochemical localization of a new GCAP related to uroguanylin.

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Analysis of the human guanylin gene and the processing and cellular localization of the peptide.

Cited by 49 publications

References 39 publications

Epitope conservation and immunohistochemical localization of the guanylin/stable toxin peptide receptor, guanylyl cyclase C

Epitope conservation and immunohistochemical localization of the guanylin/stable toxin peptide receptor, guanylyl cyclase C

HCC-2, a human chemokine: Gene structure, expression pattern, and biological activity

GCAP‐II: Isolation and characterization of the circulating form of human uroguanylin

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