UBC Theses and Dissertations
An analysis of ion transport peptide and related transcripts in desert and migratory locusts Macins, Andris
Maintenance of a homeostatic internal environment is necessary for all organisms to function. or the desert locust, a stable osmotic and ionic internal environment is especially critical when faced with widely fluctuating water and ion availability. The locust hindgut is the primary site of ion and fluid reabsorption. This reabsorptive process is regulated to carefully balance the internal environment. Although the transport mechanisms have been described thoroughly, the control of these processes is not well understood. Ion Transport Peptide (ITP) isolated from Schistocerca gregaria corpora cardiaca (CC) stimulates Na+ , K+ , CI- and water reabsorption in the locust hindgut. Both synthetic and expressed forms of ITP mimic the activity of S. gregaria CC in a biological assay. The ITP peptide and the related transcript ITP-L are the first insect members identified of a large crustacean hormone family. The purpose of this study is to examine two questions regarding ITP: 1. Is ITP restricted to the brain and CC as suggested by bioassay results? 2. Do other insects have ITP or ITP-like peptides as suggested by bioassay results? Using reverse transcription polymerase chain reaction (RT-PCR) I found ITP transcripts in the brain, corpora cardiaca and unexpectedly in the rectum. Using the same method, I found ITP is expressed during embryogenesis as early as day 9, and localized to the heads of day 12 eggs. ITP-L RNA is observed in all tissues tested and the temporal distribution of ITP-L is equally ubiquitous, as transcripts are demonstrated within in utero eggs through to day 14. Western blot analysis shows an ITP peptide of about 7.6 kD in CC homogenates. Possible post-translational differences are suggested at the C-terminal end of the ITP peptide between S. gregaria and ITP of synthetic origin and ITP from Locusta migratoria CC. RT-PCR analysis demonstrates alternate 5' sequences for both ITP and ITP-L and that there are possibly two more ITP-like transcripts in the brain. Finally, I report the sequence of L. migratoria ITP and ITP-L cloned from brain total RNA which shows high (>90%) identity relative to S. gregaria.