Poster abstracts

Poster number 134 submitted by Yaqi Wang

A unique non-spliceosomal rnp-4f nuclear intron class in Drosophila melanogaster

Yaqi Wang (Biology, Miami University), Sushmita Ghosh (Biology, Miami University), Amy L. Feiber (Biology, Miami University), Dang-Khoa Nguyen (Biology, Miami University), Jack C. Vaughn (Biology, Miami University)

Abstract:
Introns are transcribed DNA segments which are excised from the primary transcript by splicing together the flanking exons. Major eukaryotic intron classes include the GU/AG and AU/AC spliceosomal introns, self-splicing group I and group II introns, tRNA introns, and the hac1 (yeast)/xbp1 (Drosophila/human) gene intron class which plays a role in the unfolded protein response. Drosophila rnp-4f is a nuclear gene containing nine classical spliceosomal introns which encodes an evolutionarily conserved protein functioning as a chaperone enabling association between U4- and U6-snRNAs during spliceosome assembly. During an extensive screen of polyadenylated EST clones derived from D. melanogaster developmental stages, we unexpectedly found transcripts containing an identically spliced 58-nt segment from some 0-4 h embryo and also pupal clones. Excision of this intron causes a frame-shift which completely changes the encoded C-terminus and results in a long unique 83 amino acid sequence tract. This intron does not conform to any of the major intron classes, and does not arise from a second rnp-4f gene. We have identified several interesting correlations between the Drosophila xbp1 gene intron and that of the novel rnp-4f intron: (1) both produce rare mRNA transcripts, the xbp1 class being highly expressed during stress; (2) intron splicing in both creates a frame-shift which results in a long unique C-terminal amino acid sequence; (3) both are flanked by a short direct repeat (C-U-G-C-N-G-C in xbp1 and C-C-C-A-C-C-N-C in rnp-4f); (4) both excision sites occur 3-nt from a stem-loop at the 5’-splice junction; and (5) the intronic secondary structure in both brings the 5’- and 3’-ends into close proximity for subsequent ligation. Intron excision in hac1/xbp1 mRNAs is via Ire-1 endonuclease (Drosophila CG45830), while the putative rnp-4f endonuclease remains to be discovered. We have made an antibody specific to the long unique C-terminal amino acid sequence resulting from splicing by this new intron class, which should enable locating sites within fly embryos where this event occurs and to eventually determine its functional significance.

Keywords: rnp-4f gene, non-spliceosomal intron, novel new intron class