Unique synteny and alternate splicing of the chitin synthases in closely related heliothine moths

Authors: Shirk, Paul; Perera, Omaththage; Shelby, Kent; Furlong, Richard; Lovullo, Eric; Popham, Holly

Submitted to: Gene
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 8/1/2015
Publication Date: 12/10/2015
Citation: Shirk, P.D., Perera, O.P., Shelby, K., Furlong, R.B., LoVullo, E.D., Popham, H.J. 2015. Unique synteny and alternate splicing of the chitin synthases in closely related heliothine moths. Gene. 574(1):121-139.
DOI: https://doi.org/10.1016/j.gene.2015.08.001

Interpretive Summary: Heliothine moths the corn earworm, Helicoverpa zea, the cotton bollworm, Helicoverpa armigera and the tobacco budworm Heliothis virescens are major crop pests. The molting in these moths requires the production of new cuticle at every stage. Interfering with the enzymes that produce the cuticle is a major target for chemical pesticides. Scientists at USDA, Agricultural Research Service, Center for Medical, Agricultural and Veterinary Entomology, Gainesville, Florida, Southern Insect Management Research Unit and Biological Control of Insects Research Laboratory collaborated to determine the genomic organization of these critical genes and to identify how these genes are controlled in the corn earworm, the cotton bollworm and the tobacco budworm. These finding will be used to construct non-chemical methods of control based on disruption of the gene expression through RNA interference biotechnologies.

Technical Abstract: Two chitin synthase genes were characterized in the genomes of two heliothine moths: the corn earworm/cotton bollworm, Helicoverpa zea (Boddie) (Lepidoptera: Noctuidae) and the cotton bollworm, Helicoverpa armigera (Hübner) (Lepidoptera: Noctuidae). In both moths, the coding sequences for the two genes were arranged in tandem with the same orientation on the same strand with ChSB 5’ of ChSA. Each chitin synthase gene had putative consensus regulatory elements 5’ to the individual transcription start sites. The 5’ sequences of HzChSB and HzChSA in H. zea contained putative regulatory elements that may provide the control for the stage and tissue specific expression for these genes. Sequence comparisons showed that each gene was highly conserved with the respective homologues from other species but that the genomic arrangement of the two genes being tandemly juxtaposed was unique in insects. Transcript mapping of HzChSB and HzChSA in H. zea demonstrated that the HzChSB transcript was differentially spliced in various tissues and in the differing developmental stages of the larvae. A transcript comprised of all 23 exons of HzChSB was clearly present in the 1st-3rd instar larvae as well as in the foregut, midgut and hindgut of both 4th and 5th instar larvae. An alternatively spliced transcript of HzChSB that joined exons 12-21 was observed in the 2nd, 3rd, 4th and 5th instar larvae but not the 1st instar larvae. A transcript comprised of all 23 exons of HzChSA was observed in muscle, fat body, hindgut, midgut and foregut of 4th instar larvae. It was also detected in muscle, and hindgut of 5th instar larvae but not in any of the other tissues. The genomic organization of the chitin synthases in these heliothine moths is unique from all other known organisms. Only 4.3 kilobases separated the genes for HzChSB and HzChSA in H. zea. The mechanism that has led to this arrangement is unclear but is most likely a recent recombinational event that placed these two genes together. The identification of the HzChSB-E12b alternate spliced transcript is the first report of alternate splicing for the chitin synthase B group. The importance of this splice form is not clear because the protein produced lacks any enzymatic activity but retains the membrane insertion motifs. As for other insects, these genes provide an important target for potential control through RNAi but also provide a subject for broad scale genomic recombinational events.