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 initially defined as a factor essential for progression through the G2/M transition, and shown to be required to maintain the steady state level of a component of the fission yeast anaphase promoting complex/cyclosome. More recently, Dib1p has been defined as a component of the U4/U6.U5 tri-snRNP, required for pre-mRNA splicing. To investigate the mechanism(s) of Dim1 function, reiterative two-hybrid screening was performed to identify interacting proteins. Proteins thus identified were solely those involved in pre-mRNA splicing or related functions, and one partner induced a striking synthetic phenotype when co-expressed with hDim1 in mammalian cells. Saturating alanine scanning mutagenesis of Dim1 allowed delineation of amino acids essential for its ability to interact with its defined partners: mapping these residues on the structural coordinates of hDim1 defined an interactive sector of the protein. Finally, depletion studies have recently shown that Dim1 function 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 lethality during gastrulation, marked by a failure to correctly express early zygotic transcripts. These results parallel the arrest phenotypes associated with global disruption of zygotic gene expression, suggesting that Dim1 proteins maintain an essential function in gene expression in higher eukaryotes.
Zhang
Y
Y
Division of Basic Science, Fox Chase Cancer Center, Philadelphia, PA 19111, USA.
Lindblom
T
T
Chang
A
A
Sudol
M
M
Sluder
A E
AE
Golemis
E A
EA
D000818:Animals; D019556:COS Cells; D017173:Caenorhabditis elegans / Q000196:embryology / Q000235:genetics; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism*; D018797:Cell Cycle Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D002467:Cell Nucleus / Q000378:metabolism; D004268:DNA-Binding Proteins; D005656:Fungal Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D018507:Gene Expression Regulation, Developmental; D049452:Green Fluorescent Proteins; D006367:HeLa Cells; D034541:Heterogeneous-Nuclear Ribonucleoprotein Group F-H; D034441:Heterogeneous-Nuclear Ribonucleoproteins; D006801:Humans; D008164:Luminescent Proteins / Q000235:genetics / Q000378:metabolism; D008958:Models, Molecular; D009154:Mutation; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding; D012322:RNA Precursors / Q000235:genetics*; D012326:RNA Splicing; D012330:RNA, Double-Stranded / Q000502:physiology; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D012261:Ribonucleoproteins / Q000235:genetics / Q000378:metabolism*; D029702:Schizosaccharomyces pombe Proteins; D014158:Transcription, Genetic; D020798:Two-Hybrid System Techniques; D015053:Zygote / Q000254:growth & development / Q000378:metabolism
D002352:Carrier Proteins; D018797:Cell Cycle Proteins; D004268:DNA-Binding Proteins; D005656:Fungal Proteins; D034541:Heterogeneous-Nuclear Ribonucleoprotein Group F-H; D034441:Heterogeneous-Nuclear Ribonucleoproteins; D008164:Luminescent Proteins; D009687:Nuclear Proteins; C119246:PQBP1 protein, human; D012322:RNA Precursors; D012330:RNA, Double-Stranded; D011993:Recombinant Fusion Proteins; D012261:Ribonucleoproteins; D029702:Schizosaccharomyces pombe Proteins; C118369:dim1 protein, S pombe; D049452:Green Fluorescent Proteins