Nucleic Acids Research, 1995, Vol. 23, No. 24 4963-4970
© 1995
Articles |
Identification of a 67 kDa protein that binds specifically to the pre-rRNA primary processing site in a higher plant
Laboratorie de Physiologie et Biologie Moleèculaire Veègeétale, Universiteé de PErpignan URA CNRS 565, Avenue de Villeneuve, 66860 Perpignan Cedex France
*To whom correspondence should be addressed
Received October 9, 1995. Accepted November 16, 1995.
In radish pre-rRNA primary processing cleavage occurs at a UUUUCGCGC element (motif P) mapped in the 5'-external transcribed spacer (Delcasso-Tremousaygue et al., 1988). Significantly, motif P is part of a cluster of homologous elements including three UUUUCCGG elements (motifs A123) and a single UUUUGCCCC element (motif B). Here we used the EMSA to Identify In radish extracts an RNA-blndlng activity, NF C, that specifically interacts with the pre-rRNA A123BP sequence. Using different RNA probes and competitors we show that NFC recognises a 38 base RNA sequence including the 3'-end of motif A3 and motifs B and P. NF C binds to poly U, but not to poly A, poly C or poly G. Therefore we used poly (U) Sepharose chromatography as a final step to obtain pure NF C fractions. These, analysed by SDS-PAGE, revealed two major polypeptides of 67 and 60 kDa. According to UV cross-linking analysis the 67 kDa polypeptide corresponds to NF C activity, while the 60 kDa species is a proteolysed form of this protein. We also showed that NF C is enriched in nuclear extracts. Based on Its stringent RNA substrate specificity and its nuclear localisation we propose that NF C is involved in pre-rRNA primary processing in plants.