ABSTRACT
In contrast to mRNAs, ribosomal RNAs are generally not considered to be polyadenylated. Only a few recent reports describe non-abundant polyadenylated rRNA-related transcripts that have been detected and characterized in yeast and in human cells. Here we depict the phenomenon of 26S rRNA polyadenylation and degradation that was observed in shoots of Nicotiana tabaccum plants grown in the presence of cadmium. Fragments corresponding to 26S rRNA were identified using suppression subtractive hybridization during screening for genes induced in tobacco plants upon a three-week exposure to 15 microM cadmium chloride. Extracts prepared from the above-ground tissues of cadmium-treated tobacco plants were supposed to contain exclusively polyadenylated mRNAs. Surprisingly, numerous polyadenylated fragments matching parts of 26S rRNA were identified and their presence was confirmed by Northern blot and cDNA amplification techniques. To our knowledge this is the first report on rRNA polyadenylation in plants.
Subject(s)
Cadmium Chloride/toxicity , Nicotiana/drug effects , Nicotiana/metabolism , Polyadenylation , RNA Stability , RNA, Ribosomal/metabolism , Base Sequence , Blotting, Northern , Cadmium Chloride/metabolism , DNA Primers , Genes, Plant/drug effects , Molecular Sequence Data , Polymerase Chain Reaction , RNA, Ribosomal/genetics , Nicotiana/geneticsABSTRACT
During this study His-tagged CcpA protein purified under native conditions to obtain a biologically active protein was used for molecular analysis of CcpA-dependent regulation. Using electrophoretic mobility shift assays it was demonstrated that CcpA of L. lactis can bind DNA in the absence of the HPr-Ser-P corepressor and exhibits DNA-binding affinity for nucleotide sequences lacking cre sites. However, purified HPr-Ser-P protein from Bacillus subtilis was shown to slightly increase the DNA-binding capacity of the CcpA protein. It was also observed that CcpA bound to the cre box forms an apparently more stable complex than that resulting from unspecific binding. Competition gel retardation assay performed on DNA sequences from two PEP:PTS regions demonstrated that the ybhE, bglS, rheB, yebE, ptcB and yecA genes situated in these regions are most probably directly regulated by CcpA.
