David L. Garbers

Guanylyl cyclase is a heat-stable enterotoxin receptor

Plasma membrane forms of guanylyl cyclase have been shown to function as natriuretic peptide receptors. We describe a new clone (GC-C) encoding a guanylyl cyclase receptor for heat-stable enterotoxin. GC-C encodes a protein containing an extracellular amino acid sequence divergent from that of previously cloned guanylyl cyclases; however, the protein retains the intracellular protein kinase-like and cyclase catalytic domains. Expression of GC-C in COS-7 cells results in high guanylyl cyclase activity. In addition, heat-stable enterotoxin from E. coli, but not natriuretic peptides, causes marked elevations of cyclic GMP and is specifically bound by cells transfected with GC-C. The enterotoxin fails to elevate cyclic GMP in nontransfected cells or in cells transfected with the natriuretic peptide/guanylyl cyclase receptors. These results show that a heat-stable enterotoxin receptor responsible for acute diarrhea is a plasma membrane form of guanylyl cyclase.

Further characterization of a speract receptor on sea urchin spermatozoa.

It has been known for several decades that spermatozoa respond to substances associated with or produced by eggs and/or the female reproductive tract. Noted responses of sperm cells include the activation (or repression) of motility and metabolism, and induction of the acrosome reaction. In addition, sperm chemotaxis has been reported in various invertebrates and lower vertebrates (for review, see reference 4). A number of years ago our laboratory initiated research to describe the structure of those molecules that specifically interact with sperm receptors, to identify the receptor molecules, to define the behavioral responses of the spermatozoon, and to then determine the biochemical basis of the signaling system. One aspect of this work has concerned small egg-associated peptides that interact specifically with sea urchin spermatozoa. In 1976, Ohtake published a report demonstrating that the respiration of sea urchin spermatozoa could be reproducibly stimulated by crude mixtures of egg factors under conditions of acidic extracellular pH. Prior to this report, such stimulation had been seen on occasion but, in general, had been difficult to reprod ~ c e . ~ In 1981, Hansbrough and Garbers7 and Suzuki and coworkers,8 using respiration-stimulation as a bioassay, purified the peptide speract (Gly-Phe-AspLeu-Asn-Gly-Gly-Gly-Val-Gly) from the egg-jelly of the sea urchins Strongylocentrotus purpuratus and Hemicentrotus pulcherrimus. Similar peptides have been purified from the eggs of other closely related specie^.^*^ Recently, another peptide, resact, (Cys-Val-Thr-Gly-Ala-Pro-Gly-Cys-Val-Gly-Gly-Gly-Arg-LeuNH2) has been purified from the sea urchin Arbacia punctulata. Speract and resact do not appear to cross-react with spermatozoa of the opposite species. Despite the specificity of the peptides, however, they cause similar biological responses in the homologous spermatozoa. Aside from the stimulation of respiration rates and motility, the responses include enhanced H+ eftlux,* increased cyclic AMP and cyclic GMP concentration^,^ and decreased intracellular K + concentrations.3 Also, Shackman and Chock have recently demonstrated transient elevation of intracellular Ca++ in response to speract.I4 In addition, resact has been shown to be a potent chemoattractant for spermatozoa from the sea urchin Arbacia punctulata.l5

Cyclic GMP and the second messenger hypothesis

A plasma membrane form of guanylate cyclase appears to contain a single transmembrane domain that divides the protein into a highly conserved intracellular domain and a variable extracellular domain. Various extracellular peptides can bind directly to guanylate cyclase to increase the production of the second messenger, cyclic GMP.