The newt ribozyme is part of a riboprotein complex.
Strand-specific transcripts of a satellite DNA of the newts, Notophthalmus and Triturus, are present in cells in monomeric and multimeric sizes. These transcripts undergo self-catalyzed, site-specific cleavage in vitro: the reaction requires Mg2+ and is mediated by a "hammerhead" domain. Transcription of the newt ribozyme appears to be performed by RNA polymerase II under the control of a proximal sequence element and a distal sequence element. In vitro, the newt ribozyme can cleave in trans an RNA substrate, suggesting that in vivo it might be involved in RNA processing events, perhaps as a riboprotein complex. Here we show that the newt ribozyme is in fact present as a riboprotein particle of about 12 S in the oocytes of Triturus. In addition, reconstitution experiments and gel-shift analyses show that a complex is assembled in vitro on the monomeric ribozyme molecules. UV cross-linking studies identify a few polypeptide species, ranging from 31 to 65 kDa, associated to the newt ribozyme with different affinities. Finally, we find that an appropriate oligoribonucleotide substrate is specifically cleaved by the riboproteic activity in S-100 ovary extracts.
Luzi
E
E
Laboratorio di Biologia Cellulare e dello Sviluppo, Università degli Studi di Pisa, Via Carducci 13, I-56010 Ghezzano, Pisa, Italy. LUZI@mail.mpiem.gwdg.de
Eckstein
F
F
Barsacchi
G
G
D000818:Animals; D004247:DNA / Q000235:genetics / Q000378:metabolism; D019185:Notophthalmus; D016337:RNA, Catalytic / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D012261:Ribonucleoproteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D014158:Transcription, Genetic; D014319:Triturus
D016337:RNA, Catalytic; D012261:Ribonucleoproteins; D004247:DNA