Overexpression of the inositol phosphatase SopB in human 293 cells stimulates cellular chloride influx and inhibits nuclear mRNA export.
SopB is an inositol phosphate phosphatase that is a virulence factor in Salmonella species. We have overexpressed SopB cDNA in a tetracycline-dependent system in human embryonic 293 cells, and used this model system to directly analyze the role of SopB in altering inositol metabolite levels in vivo. Addition of tetracycline to these cells resulted in the rapid induction of SopB expression, which was coincident with perturbations in the cellular levels of multiple soluble inositol phosphates. All of the changes induced by SopB expression were reversed within 24 h on removal of tetracycline from media. Specifically, cellular inositol 1,3,4,5,6-pentakisphosphate (InsP(5)) and inositol hexakisphosphate (InsP(6)) levels were depleted within 4 to 6 h after inducing SopB expression. A transient rise in cellular inositol 1,4,5,6-tetrakisphosphate was also observed and was accompanied by increased chloride channel activity. This indicates that SopB alone is sufficient for changes in chloride channel function in cells infected with Salmonella organisms. Depletion of inositol phosphates, including InsP(5) and InsP(6) metabolites, was coincident with the accumulation of polyadenylated RNA in the nucleus. This suggested that a defect in nuclear export had occurred. Moreover, the penetrance of the export defect required localization of SopB to the nucleus. These results provide evidence that inositol phosphate productions may be required for efficient mRNA export in mammalian cells.
Feng
Y
Y
Division of Hematology and Department of Cell Biology and Physiology, Washington University School of Medicine, St. Louis, MO 63110, USA.
Wente
S R
SR
Majerus
P W
PW
D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism*; D018118:Chloride Channels / Q000502:physiology; D002712:Chlorides / Q000378:metabolism*; D006801:Humans; D017403:In Situ Hybridization; D007295:Inositol Phosphates / Q000378:metabolism; D007700:Kinetics; D010744:Phosphoric Monoester Hydrolases / Q000235:genetics / Q000378:metabolism*; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism; D012475:Salmonella / Q000378:metabolism; D014162:Transfection
D001426:Bacterial Proteins; D018118:Chloride Channels; D002712:Chlorides; D007295:Inositol Phosphates; D012333:RNA, Messenger; D011994:Recombinant Proteins; C060972:SopB protein, Bacteria; D010744:Phosphoric Monoester Hydrolases; C021822:myo-inositol-1 (or 4)-monophosphatase; C489224:SopB protein, Salmonella