Studies of a Proteinase-activated Cell Surface Receptor

Research output: ThesisDoctoral Thesis (compilation)

Abstract

The G protein-coupled receptors form one of the largest families of proteins involved in signalling between cells. It contains receptors for molecules as diverse as biogenic amines, glycoproteins and enzymes. The manuscripts included in this thesis together describe the initial characterisation of a previously unknown receptor of this class. A DNA fragment was cloned from a mouse genomic library. The peptide sequence deduced from the large open reading frame was about 30% identical to that of the human thrombin receptor. It also contained a putative cleavage site for trypsin-like serine proteinases. Frog oocytes injected with receptor cRNA responded with calcium mobilisation to stimulation with trypsin or a peptide (SLIGRLETQPPI) corresponding to the sequence immediately downstream of the predicted enzyme cleavage site. Receptors with a mutated cleavage site were resistant to activation by trypsin. The receptor was named the proteinase-activated receptor 2 (PAR-2). Cloning of a 3 kb mouse PAR-2 cDNA revealed a coding 5' end different from that in the genomic fragment. The subsequent isolation of mouse and human genomic DNA fragments corresponding to the cDNA showed that the PAR-2 gene consists of - at least - two exons separated by 10-14 kb intronic DNA. The sequence of the predicted human receptor agonist peptide is SLIGKVD. Northern blot and in situ hybridisation studies showed that the PAR-2 is widely expressed in adult mouse and human and fetal mouse tissues, with a preferential localisation to epithelial cells from all three germ cell layers. Messenger RNA was also detected in some non-epithelial tissues, notably in ossifying bone and in the thymus but nowhere in the CNS, except in the retina. In cultured human endothelial cells, the PAR-2 gene was strongly induced by treatment with inflammatory cytokines. Expression peaked after 48 hours but remained elevated for another two days, suggesting a role of the PAR-2 in the later phases of the acute inflammatory response.

Details

Authors
  • Sverker Nystedt
Organisations
Research areas and keywords

Subject classification (UKÄ) – MANDATORY

  • Basic Medicine

Keywords

  • receptor, proteinase, PAR-2, molecular cloning, in situ hybridisation, Expression, G-protein, Neurology, neuropsychology, neurophysiology, Neurologi, neuropsykologi, neurofysiologi
Original languageEnglish
QualificationDoctor
Awarding Institution
Supervisors/Assistant supervisor
  • [unknown], [unknown], Supervisor, External person
Award date1996 May 21
Publisher
  • Department of Physiology and Neuroscience, Lund University
Print ISBNs91-628-1981-X
Publication statusPublished - 1996
Publication categoryResearch

Bibliographic note

Defence details Date: 1996-05-21 Time: 10:15 Place: Segerfalksalen, Wallenberg Neurocentrum External reviewer(s) Name: Lendahl, Urban Title: [unknown] Affiliation: [unknown] ---