Published in: Molecular and Cellular Biology, Vol. 20, No. 3, pp. 883-891 (February, 2000):

"Trypanosoma brucei Guide RNA Poly(U) Tail Formation Is Stabilized by Cognate mRNA".

Michael T. McManus,¹ Brian K. Adler,² Victoria W. Pollard,³ and Stephen L. Hajduk¹,
Department of Biochemistry and Molecular Genetics¹ and Department of Medicine, School of Medicine,² University of Alabama at Birmingham, Birmingham, Alabama 35294, and Department of Microbiology, School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania 19104-6148³

Abstract:

Guide RNAs (gRNAs) are small RNAs that provide specificity for uridine addition and deletion during mRNA editing in trypanosomes. Terminal uridylyl transferase (TUTase) adds uridines to pre-mRNAs during RNA editing and adds a poly(U) tail to the 3' end of gRNAs. The poly(U) tail may stabilize the association of gRNAs with cognate mRNA during editing. Both TUTase and gRNAs associate with two ribonucleoprotein complexes, I (19S) and II (35S to 40S). Complex II is believed to be the fully assembled active editing complex, since it contains pre-edited mRNA and enzymes thought necessary for editing. Purification of TUTase from mitochondrial extracts resulted in the identification of two chromatographically distinct TUTase activities. Stable single-uridine addition to different substrate RNAs is performed by the 19S complex, despite the presence of a uridine-specific 3' exonuclease within this complex. Multiple uridines are added to substrate RNAs by a 10S particle that may be an unstable subunit of complex I lacking the uridine-specific 3' exonuclease. Multiple uridines could be stably added onto gRNAs by complex I when the cognate mRNA is present. We propose a model in which the purine-rich region of the cognate mRNA protects the uridine tail from a uridine exonuclease activity that is present within the complex. To test this model, we have mutated the purine-rich region of the pre-mRNA to abolish base-pairing interaction with the poly(U) tail of the gRNA. This RNA fails to protect the uridine tail of the gRNA from exoribonucleolytic trimming and is consistent with a role for the purine-rich region of the mRNA in gRNA maturation.

1. Simpson L, "RNA Editing -- An Evolutionary Perspective", in: "The RNA World", Second Edition, (eds. Gesteland RF, Cech TR, and Atkins JF), pp. 585-608, Monograph 37, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY, USA, 1999.