The role of intron structures in trans-splicing and cap 4 formation for the Leishmania spliced leader RNA.
A 39-nucleotide leader is trans-spliced onto all trypanosome nuclear mRNAs. The precursor spliced leader RNA was tested for trans-splicing function in vivo by mutating the intron. We report that in Leishmania tarentolae spliced leader RNA 5' modification is influenced by the primary sequence of stem-loop II, the Sm-binding site, and the secondary structure of stem-loop III. The sequence of stem-loop II was found to be important for cap 4 formation and splicing. As in Ascaris, mutagenesis of the bulge nucleotide in stem-loop II was detrimental to trans-splicing. Because restoration of the L. tarentolae stem-loop II structure was not sufficient to restore splicing, this result contrasts the findings in the kinetoplastid Leptomonas, where mutations that restored stem-loop II structure supported splicing. Methylation of the cap 4 structure and splicing was also dependent on both the Sm-binding site and the structure of stem-loop III and was inhibited by incomplete 3' end processing. The critical nature of the L. tarentolae Sm-binding site is consistent with its essential role in the Ascaris spliced leader RNA, whereas in Leptomonas mutation of the Sm-binding site and deletion of stem-loop III did not affect trans-splicing. A pathway for Leishmania spliced leader RNA processing and maturation is proposed.
Sturm
N R
NR
Department of Microbiology and Immunology, UCLA School of Medicine, Los Angeles, California 90095-1747, USA.
Campbell
D A
DA
D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D007438:Introns / Q000235:genetics; D007891:Leishmania / Q000235:genetics*; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009690:Nucleic Acid Conformation; D012326:RNA Splicing; D020038:RNA, Spliced Leader / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D013329:Structure-Activity Relationship
D020038:RNA, Spliced Leader