Evidence that Dim1 associates with proteins involved in pre-mRNA splicing,and delineation of residues essential for Dim1 interactions with hnRNP F and Npw38/PQBP-1

The small evolutionarily conserved protein Dim1p/hDim1/Dib1p/DML-1 was init ially defined as a factor essential for progression through the G2/M transi tion, and shown to be required to maintain the steady state level of a comp onent of the fission yeast anaphase promoting complex/cyclosome. More recen tly, Dib1p has been defined as a component of the U4/U6 . U5 tri-snRNP, req uired for pre-mRNA splicing. To investigate the mechanism(s) of Dim1 functi on, reiterative two-hybrid screening was performed to identify interacting proteins. Proteins thus identified were solely those involved in pre-mRNA s plicing or related functions, and one partner induced a striking synthetic phenotype when co-expressed with hDim1 in mammalian cells. Saturating alani ne scanning mutagenesis of Dim1 allowed delineation of amino acids essentia l for its ability to interact with its defined partners: mapping these resi dues on the structural coordinates of hDim1 defined an interactive sector o f the protein. Finally, depletion studies have recently shown that Dim1 fun ction is essential for pre-mRNA splicing in yeast. We find that elimination of DML-1 expression in C. elegans by RNA interference leads to embryonal l ethality during gastrulation, marked by a failure to correctly express earl y zygotic transcripts. These results parallel the arrest phenotypes associa ted with global disruption of zygotic gene expression, suggesting that Dim1 proteins maintain an essential function in gene expression in higher eukar yotes. (C) 2000 Elsevier Science B.V. All rights reserved.