Molecular cloning and preliminary function study of iron responsive element binding protein 1 gene from cypermethrin-resistant Culex pipiens pallens

<p>Abstract</p> <p>Background</p> <p>Insecticide resistance jeopardizes the control of mosquito populations and mosquito-borne disease control, which creates a major public health concern. Two-dimensional electrophoresis identified one protein segment with high sequence homology to part of <it>Aedes aegypti </it>iron-responsive element binding protein (IRE-BP).</p> <p>Method</p> <p>RT-PCR and RACE (rapid amplification of cDNA end) were used to clone a cDNA encoding full length IRE-BP 1. Real-time quantitative RT-PCR was used to evaluate the transcriptional level changes in the Cr-IRE strain <it>Aedes aegypti </it>compared to the susceptible strain of <it>Cx. pipiens pallens</it>. The expression profile of the gene was established in the mosquito life cycle. Methyl tritiated thymidine (³H-TdR) was used to observe the cypermethrin resistance changes in C6/36 cells containing the stably transfected IRE-BP 1 gene of <it>Cx. pipiens pallens</it>.</p> <p>Results</p> <p>The complete sequence of iron responsive element binding protein 1 (IRE-BP 1) has been cloned from the cypermethrin-resistant strain of <it>Culex pipiens pallens </it>(Cr-IRE strain). Quantitative RT-PCR analysis indicated that the IRE-BP 1 transcription level was 6.7 times higher in the Cr-IRE strain than in the susceptible strain of 4th instar larvae. The IRE-BP 1 expression was also found to be consistently higher throughout the life cycle of the Cr-IRE strain. A protein of predicted size 109.4 kDa has been detected by Western blotting in IRE-BP 1-transfected mosquito C6/36 cells. These IRE-BP 1-transfected cells also showed enhanced cypermethrin resistance compared to null-transfected or plasmid vector-transfected cells as determined by ³H-TdR incorporation.</p> <p>Conclusion</p> <p>IRE-BP 1 is expressed at higher levels in the Cr-IRE strain, and may confer some insecticide resistance in <it>Cx. pipiens pallens</it>.</p