hide
Free keywords:
-
Abstract:
The U4/U6.U5 tri-snRNP is a key component of spliceosomes. By using chemical reagents and
RNases, we performed the first extensive experimental analysis of the structure and accessibility of
U4 and U6 snRNAs in tri-snRNPs. These were purified from HeLa cell nuclear extract and
Saccharomyces cerevisiae cellular extract. U5 accessibility was also investigated. For both species,
data demonstrate the formation of the U4/U6 Y-shaped structure. In the human tri-snRNP and
U4/U6 snRNP, U6 forms the long range interaction, that was previously proposed to be
responsible for dissociation of the deproteinized U4/U6 duplex. In both yeast and human
tri-snRNPs, U5 is more protected than U4 and U6, suggesting that the U5 snRNP-specific protein
complex and other components of the tri-snRNP wrapped the 5' stem-loop of U5. Loop I of U5 is
partially accessible, and chemical modifications of loop I were identical in yeast and human
tri-snRNPs. This reflects a strong conservation of the interactions of proteins with the functional
loop I. Only some parts of the U4/U6 Y-shaped motif (the 5' stem-loop of U4 and helix II) are
protected. Due to difference of protein composition of yeast and human tri-snRNP, the U6 segment
linking the 5' stem-loop to the Y-shaped structure and the U4 central single-stranded segment are
more accessible in the yeast than in the human tri-snRNP, especially, the phylogenetically conserved
ACAGAG sequence of U6. Data are discussed taking into account knowledge on RNA and
protein components of yeast and human snRNPs and their involvement in splicesome assembly.
