Functional characterization of transcriptional regulatory elements of three highly expressed constitutive genes in the jewel wasp, Nasonia vitripennis

Authors: Shirk, Paul; Furlong, Richard; DOLAN, AMANDA - University Of Rochester; WERREN, JOHN - University Of Rochester

Submitted to: Insect Molecular Biology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 6/15/2017
Publication Date: 7/28/2017
Citation: Shirk, P.D., Furlong, R.B., Dolan, A., Werren, J.H. 2017. Functional characterization of transcriptional regulatory elements of three highly expressed constitutive genes in the jewel wasp, Nasonia vitripennis. Insect Molecular Biology. 26(6):743-750. https://doi.org/10.1111/imb.12333.
DOI: https://doi.org/10.1111/imb.12333

Interpretive Summary: The jewel wasp is a parasitoid used for biocontrol of flesh and that filth flies that has also become a major biological and genetics model for wasps and bees. The biotechnology necessary to assess gene regulation is a critical component of developing strategies for alternative methods of enhancing the biology of these beneficial insects. The use of native gene regulation elements is critical for the advancement of genetic transformation systems for this wasp. Scientists at the USDA, Agricultural Research Service, Center for Medical, Agricultural and Veterinary Entomology, Gainesville, Florida, and the University of Rochester have identified three native gene promoter elements from the jewel wasp that can be used as to support marker genes used in genetic transformation systems in this wasp. The availability of these vectors provides a valuable resource to test transfer systems in wasps and bees for the development of alternative methods to enhance the biology of these beneficial insects.

Technical Abstract: The jewel wasp, Nasonia vitripennis Ashmead (Hymenoptera: Pteromalidae) serves as an easily reared parasitoid that is providing an ever increasing malleable model for examining the biology and genetics of Hymenoptera. In order to conduct genetic transformations, it is preferable to have native highly-active constitutive genetic elements to drive expression of fluorescent reporters. Utilizing the genomic and transcriptome resources, the 5’ upstream promoter sequences from three highly expressed genes were identified and cloned. The criteria for selection included the presence of an adjacent gene 5’ of the translation initiation site. Two of the three genes lacked 5’ DNA methylation while the other is methylated. The three promoters, hsp70, activator of 90 kDa heat shock protein ATPase protein 1 (hsp90A), lipid storage droplet surface-binding protein 1 (lsdp), were linked with EGFP and cloned into both a pDP9e somatic transformation vector and a piggyBac germline transformation vector. Expression patterns based on the visible identification of GFP produced under the control of each promoter were compared with the patterns of DsRed fluorescence produced from the expression cassette of the transformation vector. The promoter activities were assessed and functional activity confirmed in transformed cultured Sf9 cells and in transformed Drosophila melanogaster. Functional activity of the promoters was also confirmed in N. vitripennis embryos demonstrating that all three promoters had functional transcriptional regulatory activity in three different insect orders. The identification and functional characterization of these three promoters will provide critical and necessary resources for further genetic analyses of Nasonia, Hymenoptera and other insects.