Association of human DEAD box protein DDX1 with a cleavage stimulation factor involved in 3'-end processing of pre-MRNA.
DEAD box proteins are putative RNA helicases that function in all aspects of RNA metabolism, including translation, ribosome biogenesis, and pre-mRNA splicing. Because many processes involving RNA metabolism are spatially organized within the cell, we examined the subcellular distribution of a human DEAD box protein, DDX1, to identify possible biological functions. Immunofluorescence labeling of DDX1 demonstrated that in addition to widespread punctate nucleoplasmic labeling, DDX1 is found in discrete nuclear foci approximately 0.5 microm in diameter. Costaining with anti-Sm and anti-promyelocytic leukemia (PML) antibodies indicates that DDX1 foci are frequently located next to Cajal (coiled) bodies and less frequently, to PML bodies. Most importantly, costaining with anti-CstF-64 antibody indicates that DDX1 foci colocalize with cleavage bodies. By microscopic fluorescence resonance energy transfer, we show that labeled DDX1 resides within a Förster distance of 10 nm of labeled CstF-64 protein in both the nucleoplasm and within cleavage bodies. Coimmunoprecipitation analysis indicates that a proportion of CstF-64 protein resides in the same complex as DDX1. These studies are the first to identify a DEAD box protein associating with factors involved in 3'-end cleavage and polyadenylation of pre-mRNAs.
Bléoo
S
S
Department of Oncology, University of Alberta, Cross Cancer Institute, Edmonton, Alberta T6G 1Z2 Canada.
Sun
X
X
Hendzel
M J
MJ
Rowe
J M
JM
Packer
M
M
Godbout
R
R
D000818:Animals; D002453:Cell Cycle / Q000502:physiology; D002467:Cell Nucleus / Q000378:metabolism*; D002478:Cells, Cultured; D053487:DEAD-box RNA Helicases; D005347:Fibroblasts; D006367:HeLa Cells; D006801:Humans; D051379:Mice; D018613:Microscopy, Confocal; D011233:Precipitin Tests; D020365:RNA Helicases / Q000378:metabolism* / Q000648:ultrastructure; D012322:RNA Precursors / Q000378:metabolism* / Q000648:ultrastructure; D012323:RNA Processing, Post-Transcriptional / Q000502:physiology*; D012333:RNA, Messenger / Q000378:metabolism* / Q000648:ultrastructure; D016601:RNA-Binding Proteins / Q000378:metabolism* / Q000648:ultrastructure; D013347:Subcellular Fractions / Q000378:metabolism; D014407:Tumor Cells, Cultured; D039221:mRNA Cleavage and Polyadenylation Factors
D012322:RNA Precursors; D012333:RNA, Messenger; D016601:RNA-Binding Proteins; D039221:mRNA Cleavage and Polyadenylation Factors; C086252:DDX1 protein, human; D053487:DEAD-box RNA Helicases; D020365:RNA Helicases