Identification of a plant-specific Zn2+-sensitive ribonuclease activity.
Ribonucleases (RNases) play a variety of cellular and biological roles in all three domains of life. In an attempt to perform RNA immuno-precipitation assays of Arabidopsis proteins, we found an EDTA-dependent RNase activity from Arabidopsis suspension tissue cultures. Further investigations proved that the EDTA-dependent RNase activity was plant specific. Characterization of the RNase activity indicated that it was insensitive to low pH and high concentration of NaCl. In the process of isolating the activity with cation exchange chromatography, we found that the EDTA dependency of the activity was lost. This led us to speculate that some metal ions, which inhibited the RNase activity, may be removed during cation exchange chromatography so that the nuclease activity was released. The EDTA dependency of the activity could be due to the ability of the EDTA chelating those metal ions, mimicking the effect of the cation exchange chromatography. Indeed, Zn(2+) strongly inhibited the activity, and the inhibition could be released by EDTA based on both in-solution and in-gel assays. In-gel assays identified two RNase activity bands. Mass spectrometry assays of those activity bands revealed more than 20 proteins. However, none of them has an apparent known nuclease domain, suggesting that one or more of those proteins might possess a currently uncharacterized nuclease domain. Our results may shed light on RNA metabolism in plants by introducing a novel plant-specific RNase activity.
Xing
Denghui
D
Department of Botany, Miami University, Oxford, OH 45056, USA.
Ni
Shuisong
S
Kennedy
Michael A
MA
Li
Qingshun Quinn
QQ
D017360:Arabidopsis / Q000201:enzymology*; D004492:Edetic Acid / Q000494:pharmacology; D006863:Hydrogen-Ion Concentration / Q000187:drug effects; D012260:Ribonucleases / Q000037:antagonists & inhibitors / Q000302:isolation & purification / Q000378:metabolism*; D012965:Sodium Chloride / Q000494:pharmacology; D013045:Species Specificity; D015032:Zinc / Q000494:pharmacology*
D012965:Sodium Chloride; D004492:Edetic Acid; D012260:Ribonucleases; D015032:Zinc