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Title: Functions of Ion Transport Peptide and Ion Transport Peptide-Like in the Red Flour Beetle Tribolium castaneum

Author
item BEGUM, KHURSHIDA - Kansas State University
item LI, BIN - Kansas State University
item Beeman, Richard
item PARK, YOONSEONG - Kansas State University

Submitted to: Insect Biochemistry and Molecular Biology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 8/19/2009
Publication Date: 2/15/2010
Citation: Begum, K., Li, B., Beeman, R.W., Park, Y. 2010. Functions of Ion Transport Peptide and Ion Transport Peptide-Like in the Red Flour Beetle Tribolium castaneum. Insect Biochemistry and Molecular Biology. 39: 717-725.

Interpretive Summary: Water balance is a critical function in insects, especially in extremely dry environments such as those associated with post-harvest pest species. Nevertheless, insect water balance physiology remains one of the major, unexploited targets for biopesticide targeting. The red flour beetle, Tribolium castaneum, is a major pest of dry storage and is the best laboratory model for gene discovery in pest insect species. In order to increase our understanding of water balance physiology in this insect, we undertook a functional genomics study of ion transport in relation to fluid retention. We identified three peptide hormones in Tribolium that appear to have additional, critical roles in ovarian development and molting, in addition to their probable functions in fluid balance. This research reveals unexpected diversity and complexity in insect neuroendocrinology, and could lead to new ways to disrupt water balance and other critical physiological functions in pest insects.

Technical Abstract: Ion transport peptide (ITP) and ITP-like (ITPL) are highly conserved neuropeptides in insects and crustaceans. We investigated the alternatively spliced variants of ITP/ITPL in Tribolium castaneum to understand their functions. We identified three alternatively spliced transcripts named itp, itpl-1, and itpl-2. Expression patterns of the splice variants investigated by exon-specific in situ hybridization were somewhat different from those previously reported in other insect species. Most importantly, we found for the first time that itpl-1 transcripts are abundantly expressed in the midgut at the late larval stage, showing an expression pattern similar to that of the crustacean hyperglycemic hormone (CHH) in the crab Carcinus maenas. CHH was shown to function by increasing the body volume through fluid absorption, resulting in breakage of the outer shell at the time of molt, and a similar function in enabling the molt is possible in Tribolium. Exon-specific RNA interference (RNAi) was designed to distinguish between itp and itpl-1, but we were unable to design a dsRNA uniquely targeting or uniquely excluding itpl-2; therefore, RNAi targeting was limited to either itp/itpl-2 or itpl-1/itpl-2. For dsRNA injections in the larval stages, either RNAi led to gradually increasing mortality in the larval and pupal stages, with 100% cumulative mortality at the time of eclosion or shortly afterward. Developmental deficiencies in the adult tarsal segments were observed after RNAi suppressing either itp/itpl-2 or itpl-1/itpl-2. After dsRNA injections at the pupal stage, the most striking observation was a significant reduction in egg numbers (8% of control) and reduced survival of the offspring (5%) in RNAi targeting itpl-1/itpl-2, while a milder degree of the same phenotype was observed in that targeting itp/itpl-2. These results indicate multiple, vital roles for ion transport peptides in Tribolium.