The roles of Rrp5p in the synthesis of yeast 18S and 5.8S rRNA can be functionally and physically separated

The yeast nucleolar protein Rrp5p is the only known trans-acting factor tha t is essential for the synthesis of both 18S rRNA and the major, short form of 5.8S (5.8S(s)) rRNA, which were thought to be produced in two independe nt sets of pre-rRNA processing reactions. To identify domains within Rrp5p required for either processing pathway, we have analyzed a set of eight del etion mutants that together cover the entire RRP5 sequence, Surprisingly, o nly one of the deletions is lethal, indicating that regions encompassing ab out 80% of the protein can be removed individually without disrupting its e ssential biological function. Biochemical analysis clearly demonstrated the presence of two distinct functional domains, Removal of each of three cont iguous segments from the N-terminal half specifically inhibits the formatio n of 5.8S(s) rRNA, whereas deleting part of the C-terminaI region of the pr otein only blocks the production of 18S rRNA. The latter phenotype is also caused by a temperature-sensitive mutation within the same C-terminal regio n, The two functional regions identified by the mutational analysis appear to be correlated with the structural domains detected by computer analysis. They can even be physically separated, as demonstrated by the fact that fu ll Rrp5p activity can be supplied by two contiguous protein fragments expre ssed in trans.