NAEGLERIA NUCLEOLAR INTRONS CONTAIN 2 GROUP-I RIBOZYMES WITH DIFFERENT FUNCTIONS IN RNA SPLICING AND PROCESSING

We have characterized the structural organization and catalytic proper ties of the large nucleolar group I introns (NaSSU1) of the different Naegleria species N. jamiesoni, N. andersoni, N. italica, and N. grube ri. NaSSU1 consists of three distinct RNA domains: an open reading fra me encoding a homing-type endonuclease, and a small group I ribozyme ( NaGIR1) inserted into the P6 loop of a second group I ribozyme (NaGIR2 ). The two ribozymes have different functions in RNA splicing and proc essing. NaGIR1 is an unusual self-cleaving group I ribozyme responsibl e for intron processing at two internal sites (IPS1 and IPS2), both cl ose to the 5' end of the open reading frame. This processing is hypoth esized to lead to formation of a messenger RNA for the endonuclease. S tructurally NaGIR2 is a typical group IC1 ribozyme, catalyzing intron excision and exon ligation reactions. NaGIR2 is responsible for circul arization of the excised intron, a reaction that generates full-length RNA circles of wild-type intron. Although it: is only distantly relat ed in primary sequence, NaSSU1 RNA has a predicted organization and fu nction very similar to that of the mobile group I intron DiSSU1 of Did ymium, the only other group I intron known to encode two ribozymes. We propose that these twin-ribozyme introns define a distinct category o f group I Introns with a conserved structural organization and functio n.