Biogenesis, cellular localization, and functional activation of the heat-stable enterotoxin receptor (guanylyl cyclase C).
Enterotoxigenic Escherichia coli elaborate a peptide called heat-stable enterotoxin (ST), which binds to and activates the intestinal ST receptor (STaR). STaR, also known as guanylyl cyclase C (GC-C), is a member of the transmembrane guanylyl cyclase receptor family. The mRNA for STaR encodes an approximately 120 kDa protein with the N-terminal ligand binding domain on the cell surface. Ligand affinity cross-linking studies have previously demonstrated several species of potential ST binding proteins, ranging in size from approximately 50 to 160 kDa. Although these smaller forms of STaR (50-80 kDa) have been proposed to act in vivo as toxin binding proteins, their biogenesis and localization have not previously been examined. Using pulse labeling in vivo and synchronized translation in vitro, we demonstrate that these smaller forms represent incomplete translational products and are not formed through limited proteolysis of the full-length receptor, as had previously been believed. We determined, using fluorescence confocal microscopy and surface labeling, that only approximately 25% of cellular receptors are expressed at the surface, while the remaining population is retained within the endoplasmic reticulum. Only full-length receptor is found at the surface of the cell, indicating this to be the biologically active form of STaR responsible for interacting with the heat-stable enterotoxin and other luminal intestinal peptides. The large intracellular receptor population, and potential for function before translocation to the cell surface, may impact on how pharmacologic modulators of this clinically important receptor are designed.
Rudner
X L
XL
Department of Medicine, Duke University Medical Center, Durham, North Carolina 27710, USA.
Nicchitta
C
C
Almenoff
J S
JS
D000595:Amino Acid Sequence; D000818:Animals; D001427:Bacterial Toxins / Q000378:metabolism*; D002460:Cell Line; D004768:Enterotoxins / Q000378:metabolism*; D004789:Enzyme Activation; D029968:Escherichia coli Proteins; D006162:Guanylate Cyclase / Q000096:biosynthesis* / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D008969:Molecular Sequence Data; D014176:Protein Biosynthesis; D011499:Protein Processing, Post-Translational; D000074261:Receptors, Enterotoxin; D050705:Receptors, Guanylate Cyclase-Coupled; D018000:Receptors, Peptide / Q000096:biosynthesis* / Q000235:genetics / Q000378:metabolism*; D013347:Subcellular Fractions / Q000201:enzymology
D001427:Bacterial Toxins; D004768:Enterotoxins; D029968:Escherichia coli Proteins; D018000:Receptors, Peptide; C034151:heat stable toxin (E coli); D006162:Guanylate Cyclase; D000074261:Receptors, Enterotoxin; D050705:Receptors, Guanylate Cyclase-Coupled