Evidence for dna endonuclease activity in nuclear extracts from mosquito cells

We describe a deoxyribonuclease activity from nuclear protein extracts of cultured Aedes albopictus mosquito cells. The nuclease cleaved linear and circular double-stranded DNA, first generating 3′ OH single-stranded nicks followed by second strand cleavage, but had little or no exonucleolytic activity. Detection of this activity was optimal at pH 7.1, in the presence of a divalent cation (Mg²⁺, Ca²⁺, Mn²⁺, Ba²⁺). In the presence of Mg²⁺, Zn²⁺, Hg²⁺ and Cu²⁺ inhibited activity, sulfhydryl reagents and ATP had no effect. At physiological temperatures (18-35°C), linear double-stranded DNA probes were preferentially cleaved near sites containing 3-6 consecutive deoxyadenine/thymine base pairs. Results from salt dependency and drug inhibition studies, combined with inspection of DNA sequence, suggested that DNA structure is among the parameters that determine preferred cleavage sites.