Cytoplasmic inositol hexakisphosphate production is sufficient for mediating the Gle1-mRNA export pathway.
Production of inositol hexakisphosphate (IP6) by Ipk1, the inositol-1,3,4,5,6-pentakisphosphate 2-kinase, is required for Gle1-mediated mRNA export in Saccharomyces cerevisiae cells. To examine the network of interactions that require IP6 production, an analysis of fitness defects was conducted in mutants harboring both an ipk1 null allele and a mutant allele in genes encoding nucleoporins or transport factors. Enhanced lethality was observed with a specific subset of mutants, including nup42, nup116, nup159, dbp5, and gle2, all of which had been previously connected to Gle1 function. Complementation of the nup116Deltaipk1Delta and nup42Deltaipk1Delta double mutants did not require the Phe-Gly repeat domains in the respective nucleoporins, suggesting that IP6 was acting subsequent to heterogeneous nuclear ribonucleoprotein targeting to the nuclear pore complex. With Nup42 and Nup159 localized exclusively to the nuclear pore complex cytoplasmic side, we speculated that IP6 may regulate a cytoplasmic step in mRNA export. To test this prediction, the spatial requirements for the production of IP6 were investigated. Restriction of Ipk1 to the cytoplasm did not block IP6 production. Moreover, coincident sequestering of both Ipk1 and Mss4 (an enzyme required for phosphatidylinositol 4,5-bisphosphate production) to the cytoplasm also did not block IP6 production. Given that the kinase required for inositol 1,3,4,5,6-pentakisphosphate production (Ipk2) is localized in the nucleus, these results indicated that soluble inositides were diffusing between the nucleus and the cytoplasm. Additionally, the cytoplasmic production of IP6 by plasma membrane-anchored Ipk1 rescued a gle1-2 ipk1-4 synthetic lethal mutant. Thus, cytoplasmic IP6 production is sufficient for mediating the Gle1-mRNA export pathway.
Miller
Aimee L
AL
Department of Cell and Developmental Biology, Vanderbilt University Medical Center, Nashville, Tennessee 37232-8240, USA.
Suntharalingam
Mythili
M
Johnson
Sylvia L
SL
Audhya
Anjon
A
Emr
Scott D
SD
Wente
Susan R
SR
D021581:Active Transport, Cell Nucleus; D000483:Alleles; D002352:Carrier Proteins / Q000502:physiology*; D002462:Cell Membrane / Q000378:metabolism; D002467:Cell Nucleus / Q000378:metabolism; D002851:Chromatography, High Pressure Liquid; D020285:Cryoelectron Microscopy; D003593:Cytoplasm / Q000378:metabolism*; D017353:Gene Deletion; D005816:Genetic Complementation Test; D005822:Genetic Vectors; D005838:Genotype; D049452:Green Fluorescent Proteins / Q000737:chemistry / Q000378:metabolism; D008954:Models, Biological; D009154:Mutation; D028861:Nuclear Pore Complex Proteins / Q000737:chemistry; D010641:Phenotype; D010770:Phosphotransferases / Q000378:metabolism; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000378:metabolism; D010833:Phytic Acid / Q000378:metabolism*; D010957:Plasmids / Q000378:metabolism; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D012333:RNA, Messenger / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000378:metabolism / Q000502:physiology*; D013696:Temperature; D013997:Time Factors
D002352:Carrier Proteins; C101408:GLE1 protein, S cerevisiae; D028861:Nuclear Pore Complex Proteins; D012333:RNA, Messenger; D029701:Saccharomyces cerevisiae Proteins; D049452:Green Fluorescent Proteins; D010833:Phytic Acid; D010770:Phosphotransferases; C425775:ARG82 protein, S cerevisiae; C120127:IPK1 protein, S cerevisiae; D017853:Phosphotransferases (Alcohol Group Acceptor); C489795:MSS4 protein, S cerevisiae