Nuclear localization of poly(A)+ mRNA following siRNA reduction of expression of the mammalian RNA helicases UAP56 and URH49.
UAP56 is a eukaryotic RNA helicase that is important for mRNA splicing and nuclear export. Although most eukaryotes have a single protein corresponding to UAP56, we have shown previously that in human and mouse cells there is a second protein, URH49, which is 90% identical to UAP56. Both proteins interact with the mRNA export factor Aly and both are able to rescue the loss of Sub2p (the yeast homolog of UAP56), suggesting that both proteins have similar functions. However, the two helicases have different expression profiles in different tissues and in growth-stimulated cells, which raises the possibility that they might be involved in the splicing and export of non-identical populations of mRNA. In the present study, we have used RNA interference to further explore the functions of these two helicases. Reducing the expression of either URH49 or UAP56 in HeLa cells had little effect on cell proliferation or expression of a co-transfected gene. However, analysis of poly(A)+ RNA localization by fluorescent in situ hybridization revealed a speckled pattern of RNA accumulation throughout the nucleus. Reducing the expression of both helicases resulted in a major reduction in reporter gene expression as well as cell death within 72 h. We also observed a more prominent speckled pattern of nuclear poly(A)+ RNA accumulation as well as reduced accumulation in the cytoplasmic compartment. These observations suggest that both helicases have essential but largely overlapping functions in the processing and export of mammalian mRNAs.
Kapadia
Fehmida
F
The Ohio State Biochemistry Program, The Ohio State University, Columbus, OH 43210, USA.
Pryor
Anne
A
Chang
Tien-Hsien
TH
Johnson
Lee F
LF
D021581:Active Transport, Cell Nucleus; D002467:Cell Nucleus / Q000378:metabolism*; D002470:Cell Survival; D053487:DEAD-box RNA Helicases / Q000235:genetics / Q000378:metabolism*; D017930:Genes, Reporter; D006367:HeLa Cells; D006801:Humans; D017404:In Situ Hybridization, Fluorescence; D034622:RNA Interference; D012326:RNA Splicing; D012333:RNA, Messenger / Q000032:analysis / Q000378:metabolism*; D034741:RNA, Small Interfering / Q000378:metabolism; D014162:Transfection
D012333:RNA, Messenger; D034741:RNA, Small Interfering; C498428:DDX39A protein, human; C487886:DDX39B protein, human; D053487:DEAD-box RNA Helicases