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1.  Cloning and characterization of cDNAs encoding putative CTCFs in the mosquitoes, Aedes aegypti and Anopheles gambiae 
One of the many ascribed functions of CCCTC-binding factor (CTCF) in vertebrates is insulation of genes via enhancer-blocking. Insulation allows genes to be shielded from "cross-talk" with neighboring regulatory elements. As such, endogenous insulator sequences would be valuable elements to enable stable transgene expression. Recently, CTCF joined Su(Hw), Zw5, BEAF32 and GAGA factor as a protein associated with insulator activity in the fruitfly, Drosophila melanogaster. To date, no known insulators have been described in mosquitoes.
We have identified and characterized putative CTCF homologs in the medically-important mosquitoes, Aedes aegypti and Anopheles gambiae. These genes encode polypeptides with eleven C2H2 zinc fingers that show significant similarity to those of vertebrate CTCFs, despite at least 500 million years of divergence. The mosquito CTCFs are constitutively expressed and are upregulated in early embryos and in the ovaries of blood-fed females. We have uncovered significant bioinformatics evidence that CTCF is widespread, at least among Drosophila species. Finally, we show that the An. gambiae CTCF binds two known insulator sequences.
Mosquito CTCFs are likely orthologous to the widely-characterized vertebrate CTCFs and potentially also serve an insulating function. As such, CTCF may provide a powerful tool for improving transgene expression in these mosquitoes through the identification of endogenous binding sites.
PMCID: PMC1174870  PMID: 15985163
2.  High-level gene expression in Aedes albopictus cells using a baculovirus Hr3 enhancer and IE1 transactivator 
Aedes aegypti is the key vector of both the Yellow Fever and Dengue Fever viruses throughout many parts of the world. Low and variable transgene expression levels due to position effect and position effect variegation are problematic to efforts to create transgenic laboratory strains refractory to these viruses. Transformation efficiencies are also less than optimal, likely due to failure to detect expression from all integrated transgenes and potentially due to limited expression of the transposase required for transgene integration.
Expression plasmids utilizing three heterologous promoters and three heterologous enhancers, in all possible combinations, were tested. The Hr3/IE1 enhancer-transactivator in combination with each of the constitutive heterologous promoters tested increased reporter gene expression significantly in transiently transfected Aedes albopictus C7-10 cells.
The addition of the Hr3 enhancer to expression cassettes and concomitant expression of the IE1 transactivator gene product is a potential method for increasing the level of transgene expression in insect systems. This mechanism could also potentially be used to increase the level of transiently-expressed transposase in order to increase the number of integration events in transposon-mediated transformation experiments.
PMCID: PMC487899  PMID: 15251037

Results 1-2 (2)