3 ISOFORMS OF A HEPATOCYTE NUCLEAR FACTOR-4 TRANSCRIPTION FACTOR WITHTISSUE-SPECIFIC AND STAGE-SPECIFIC EXPRESSION IN THE ADULT MOSQUITO

We cloned three isoforms of hepatocyte nuclear factor-4 (HNF-4) from t he mosquito Aedes aegypti, designated AaHNF-4a, AaHNF-4b, and AaHNF-4c . AaHNF-4a and AaHNF-4b are typical members of the HNF-4 subfamily of nuclear receptors with high amino acid conservation. They differ in N- terminal regions and exhibit distinct developmental profiles in the fe male mosquito fat body, a metabolic tissue functionally analogous to t he vertebrate liver. The AaHNF-4b mRNA is predominant during the previ tellogenic and vitellogenic phases, while the AaHNF-4a mRNA is predomi nant during the termination phase of vitellogenesis, coinciding with t he onset of lipogenesis. The third isoform, AaHNF-4c, lacks part of th e A/B and the entire C (DNA-binding) domains. The AaHNF-4c transcript found in the fat body during the termination of vitellogenesis may ser ve as a transcriptional inhibitor. Both AaHNF-4a and AaHNF-4b bind to the cognate DNA recognition site in electrophoretic mobility shift ass ay. Dimerization of AaHNF-4c with other mosquito HNF-4 isoforms or wit h mammalian HNF-4 prevents binding to the HNF-4 response element. In t ransfected human 293T cells, AaHNF-4c significantly reduced the transa ctivating effect of the human HNF-4 alpha 1 on the apolipoprotein CIII promoter. Electrophoretic mobility shift assay confirmed the presence of HNF-4 binding sites upstream of A. aegypti vg and vcp, two yolk pr otein genes expressed in the female mosquito fat body during vitelloge nesis. Therefore, HNF-4, an important regulator of liver-specific gene s, plays a critical role in the insect fat body.