I-PpoI, the Endonuclease Encoded by the Group I Intron PpLSU3, Is Expressed from an RNA Polymerase I Transcript

ABSTRACT

PpLSU3, a mobile group I intron in the rRNA genes of Physarum polycephalum, also can home into yeast chromosomal ribosomal DNA (rDNA) (D. E. Muscarella and V. M. Vogt, Mol. Cell. Biol. 13:1023–1033, 1993). By integrating PpLSU3 into the rDNA copies of a yeast strain temperature sensitive for RNA polymerase I, we have shown that the I-PpoI homing endonuclease encoded by PpLSU3 is expressed from an RNA polymerase I transcript. We have also developed a method to integrate mutant forms of PpLSU3 as well as the Tetrahymena intron TtLSU1 into rDNA, by expressing I-PpoI in trans. Analysis of I-PpoI expression levels in these mutants, along with subcellular fractionation of intron RNA, strongly suggests that the full-length excised intron RNA, but not RNAs that are further cleaved, serves as or gives rise to the mRNA.

ACKNOWLEDGMENTS

We are grateful to Masayasu Nomura for providing strain NOY401 and plasmid pNOY103, Sarah Woodson for providing plasmids pSW012, pI3TZ, and pI3ΔORFTZ, and Robert Lowery (Promega Corporation) for providing the purified I-PpoI protein. We thank Steinar Johansen and Wayne Decatur for helpful discussions during the course of this work. We also thank Wayne Decatur for critical reading of the manuscript.

This work is supported by grant GM-51860 from the USPHS.