Non-classic characteristics define prominent DNase activities from the intestine and other tissues of Haemonchus contortus.
The anthelmintic fenbendazole (FBZ) induces nuclear DNA fragmentation (DF) in intestinal cells of Haemonchus contortus. The DNA fragments had 3'-OH, which suggests involvement of a neutral DNase. To identify candidate DNase(s) involved, DNase activity in H. contortus intestine and other worm fractions was characterized relative to classic DNases I (neutral) and II (acidic). Seven distinct DNase activities were identified and had Mrs of 34, 36, 37 or 38.5 kDa on zymographic analysis. The different activities were distinguished according to pH requirement, sensitivity to 10 mM EDTA and worm compartment. Activities of intestinal DNases at 34, 36 and 38.5 kDa were sensitive to EDTA at pH 5.0 and 7.0. Sensitivity to EDTA at pH 5.0 was unexpected compared to classic acidic DNase II activity, suggesting unusual properties of these DNases. In whole worms, however, the activities at 36 and 38.5 kDa were relatively insensitive to EDTA, indicating predominance of DNases that are distinct from the intestine. The activity at 37 kDa in excretory/secretory products had an acidic pH requirement and was insensitive to EDTA, resembling classic acidic DNase activity. Under conditions of pH 5.0 and 7.0, intestinal DNases produced 3'-ends that could be labeled by terminal deoxynucleotidyl transferase, indicating presence of 3'-OH. The labeling of 3'-ends at pH 5.0, again, was unexpected for acidic DNase activity. These results and several other activities suggest that multiple H. contortus DNases have characteristics distinct from the classic mammalian DNases I and II. Treatment of H. contortus with FBZ did not induce any detectable DNase activities distinct from normal intestine, although relative activities of intestinal DNases appear to have been altered by this treatment.
Kwak
Dongmi
D
Department of Veterinary Microbiology and Pathology, Washington State University, Pullman, WA 99164-7040, USA.
Jasmer
Douglas P
DP
D000818:Animals; D000969:Antinematodal Agents / Q000494:pharmacology; D001312:Aurintricarboxylic Acid / Q000494:pharmacology; D002614:Chelating Agents / Q000494:pharmacology; D003851:Deoxyribonucleases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D004492:Edetic Acid / Q000494:pharmacology; D004591:Electrophoresis, Polyacrylamide Gel; D005273:Fenbendazole / Q000494:pharmacology; D006189:Haemonchus / Q000187:drug effects / Q000201:enzymology*; D006863:Hydrogen-Ion Concentration; D007422:Intestines / Q000187:drug effects / Q000201:enzymology; D010744:Phosphoric Monoester Hydrolases / Q000378:metabolism; D010957:Plasmids / Q000378:metabolism; D012756:Sheep; D014638:Vanadates / Q000494:pharmacology
D000969:Antinematodal Agents; D002614:Chelating Agents; D014638:Vanadates; D001312:Aurintricarboxylic Acid; D005273:Fenbendazole; D004492:Edetic Acid; D003851:Deoxyribonucleases; D010744:Phosphoric Monoester Hydrolases