Molecular cloning of a putative Na(+)-K(+)-2Cl- cotransporter from the Malpighian tubules of the tobacco hornworm, Manduca sexta.

Fluid secretion in insects is controlled, in part, by diuretic hormones which bind to receptors on the surface of Malpighian tubule cells, leading to an increase in intracellular cAMP. Recent evidence suggests that Na(+)-K(+)-2Cl- cotransporters may be involved, upon stimulus by diuretic hormones, in transport of ions from the hemolymph to the lumen of the Malpighian tubules. In this report, I describe the cloning of a putative bumetanide sensitive Na(+)-K(+)-2Cl- cotransporter from the Malpighian tubules of the tobacco hornworm, Manduca sexta. This Na(+)-K(+)-2Cl- cotransporter is a Malpighian tubule specific isoform by Northern blot analysis. The protein consists of 1060 amino acids, contains 12 putative membrane spanning regions, and is related to other Na(+)-(K+)-Cl- cotransporters that have been cloned from shark, flounder, rat, rabbit and mouse. This protein is the first Na(+)-K(+)-2Cl- cotransporter that has been identified in an invertebrate.