A Monoclonal Antibody Against the PSTAIR Sequence of p34cdc2, Catalytic Subunit of Maturation-Promoting Factor and Key Regulator of the Cell Cycle

A homolog of the serine/threonine protein kinase (p34^cdc2), encoded by the cdc2 ⁺ gene of the fission yeast ( Schizosaccharomyces pombe ), is a catalytic subunit of maturation-promoting factor and a key regulator of the cell cycle. We have raised a monoclonal antibody against the most conserved amino acid sequence, the PSTAIR sequence (EGVPSTAIREISLLKE) of p34^cdc2 This antibody recognizes 31–34 kDa proteins by immunoblotting in all species examined so far. The proteins recognized by the anti-PSTAIR antibody are probably either p34^cdc2 itself or proteins highly homologous to p34^cdc2 in the given species, since, in all species studies to date, they are all precipitated with p13^suc1, the fission yeast suc 1⁺ gene product, which binds to p34^cdc2 with high specificity. The anti-PSTAIR immunoprecipitate had no histone H1 kinase activity and did not contain cyclin B, suggesting that the PSTAIR region is masked when p34^cdc2 forms a complex with cyclin B as an active kinase. Immunoblotting with the anti-PSTAIR antibody demonstrated that the fastest-migrating form of p34^cdc2 homologues becomes abundant, when oocytes mature or the cell enters M phase. The possible significance of this observation is discussed in relation to the phosphorylation and activity state of p34^cdc2 The observed broad cross-reactivity of the anti-PSTAIR antibody against p34^cdc2 homologues in various species should permit us to examine the role of p34^cdc2 homologues in the regulation of the cell cycle in a variety of organisms.     

Influence of the G2 cell cycle block abrogator pentoxifylline on the expression and subcellular location of cyclin B1 and p34cdc2 in HeLa cervical carcinoma cells

The progression of cells from G₂ into mitosis is mainly controlled by formation of the cyclin B1/p34^cdc2 complex. The behaviour of this complex in the irradiation-induced G₂ cell cycle delay is still unclear. A prior study demonstrated that the expression of the cyclin B1 protein is reduced by irradiation, and restored to control levels by the methylxanthine drug pentoxifylline, which is a potent G₂ block abrogator. The present study shows that irradiation, and 2 mM pentoxifylline affect the expression of the cyclin-dependent kinase p34^cdc2 in HeLa cells. Irradiation induces p34^cdc2 levels to increase and cyclin B1 levels to decrease. Addition of pentoxifylline at the G₂ maximum reverses these trends. This is also evident from the cyclin B1/p34^cdc2 ratios which decline after irradiation and are rapidly restored to control levels upon addition of pentoxifylline. It is concluded that cyclin B1 and p34^cdc2 protein expression are important events and act in concert to control the irradiation induced G₂ block. Analysis of cyclin B1 expression in whole cells and in isolated nuclei furthermore show that cyclin B1 is translocated from the nucleus into the cytoplasm when the G₂ block is abrogated by pentoxifylline.     

Involvement in Meiotic Prophase of H1 Histone Kinase and p34cdc2 Homologues in Lily (Lilium longiflorum) Microsporocytes

We have taken advantage of the synchrony of meiotic prophase I in Lilium microsporocytes to investigate the presence and involvement in four stages of meiotic prophase I (leptotene, zygotene, pachytene, and diplotene) of the p34^cdc2 H1 histone kinase, a component of MPF and a key participant in division control in other eukaryotes. H1 kinase activity showed a peak pattern during meiotic prophase I with the highest kinase activity at pachytene. A monoclonal antibody directed against a highly conserved region of p34^cdc2 (termed the 'PSTAIR') recognized three major protein forms by immunoblotting. The highest level of the fastest-migrating form was observed at pachytene, coinciding with the highest activity of H1 kinase. Both the proteins recognized by the anti-PSTAIR antibody and H1 histone kinase activity were retained on beads conjugated with p13^suc1, a protein known to physically associate with p34^cdc2. These observations suggest that p34^cdc2 or protein(s) highly homologous to p34^cdc2 is a component of Lilium H1 histone kinase and plays a role in regulating meiotic prophase I.     

Immunological Characterization of cdc2 and wee1 Proteins during the Growth and Differentiation of Dictyostelium discoideum

Two different antibody preparations, raised independently against the conserved EGVPSTAIREISLLKE sequence of the protein kinases encoded by the Schizosaccharomyces pombe cdc2 gene and its species homologs, immunoblotted a Dictyostelium protein of 32 kDa (p32). This polypeptide bound to p13^suc1-agarose beads, suggesting that it represents the Dictyostelium cdc2 and / or cdk2 products. The amounts of p32 detectable in cell free extracts and bound to p13^suc1-agarose were unaltered during the growth of cells synchronized for division. Although there was also essentially no change in the level of p32 during differentiation, the protein from the pseudoplasmodial stage of development did not bind to p13^suc1-agarose, implicating a developmentally regulated modification of the kinase. One of the EGVPSTAIREISLLKE antibodies also recognized a protein of 49 kDa (p49) that increased in amount dramatically during aggregation and then remained at elevated levels throughout the remainder of the differentiation process. This protein was present in low amounts throughout the growth of cells synchronized for division and was not absorbed by p13^suc1-agarose.
A 103 kDa protein (p103) was detected by Western blot analysis using antibodies raised against two different peptides corresponding to sequences in the S. pombe protein kinase p105^wee1, which is a putative upstream negative regulator of p34^cdc2 in fission yeast. The levels of p103 were constant during differentiation and during the growth of cells synchronized for division.     

The cell cycle as a therapeutic target against Trypanosoma brucei: Hesperadin inhibits Aurora kinase-1 and blocks mitotic progression in bloodstream forms

Aurora kinase family members co-ordinate a range of events associated with mitosis and cytokinesis. Anti-cancer therapies are currently being developed against them. Here, we evaluate whether Aurora kinase-1 (TbAUK1) from pathogenic Trypanosoma brucei might be targeted in anti-parasitic therapies as well. Conditional knockdown of TbAUK1 within infected mice demonstrated its essential contribution to infection. An in vitro kinase assay was developed which used recombinant trypanosome histone H3 as a substrate. Tandem mass spectroscopy identified a novel phosphorylation site in the carboxyl-tail of recombinant trypanosome histone H3. Hesperadin, an inhibitor of human Aurora B, prevented the phosphorylation of substrate with IC₅₀ of 40 nM. Growth of cultured bloodstream forms was also sensitive to Hesperadin (IC₅₀ of 50 nM). Hesperadin blocked nuclear division and cytokinesis but not other aspects of the cell cycle. Consequently, growth arrested cells accumulated multiple kinetoplasts, flagella and nucleoli, similar to the effects of RNAi-dependent knockdown of TbAUK1 in cultured bloodstream forms cells. Molecular models predicted high-affinity binding of Hesperadin to both conserved and novel sites in TbAUK1. Collectively, these data demonstrate that cell cycle progression is essential for infections with T. brucei and that parasite Aurora kinases can be targeted with small-molecule inhibitors.     

A cdc2-Like Kinase Associated with Commitment to Division in Paramecium tetraurelia

ABSTRACT. Cell division in higher eukaryotes is mainly controlled by p34 ^cdc2 or related kinases and by other components of these kinase complexes. We present evidence that cdc2 -like kinases also occur in Paramecium. Two polypeptides reacted with an antibody directed against the perfectly conserved PSTAIR region found in cdc2 kinases in other eukaryotes. Only the less abundant peptide bound to p13 ^suc1 from Schizosaccharomyces pombe. Using centrifugal elutriation to select cells on the basis of size, we isolated highly synchronous Paramecium G1 cells. With this procedure, we demonstrated that the p13^suc1-associated cdc2 -like histone H1 kinase was activated before cell division at the point of commitment to division in Paramecium. Further, we show that Paramecium cdc2 -like proteins occurred principally as monomers and that these monomers were active as histone H1 kinases in vitro.     

Interaction of p58(PITSLRE), a G2/M-specific protein kinase,with cyclin D3

The p58(PITSLRE) is a p34(cdc2)-related protein kinase that plays an important role in normal cell cycle progression. Elevated expression of p58(PITSLRE) in eukaryotic cells prevents them from undergoing normal cytokinesis and appears to delay them in late telophase. To investigate the molecular mechanism of p58(PITSLRE) action, we used the yeast two-hybrid system, screened a human fetal liver cDNA library, and identified cyclin D3 as an interacting partner of p58(PITSLRE). In vitro binding assay, in vivo coimmunoprecipitation, and immunofluorescence cell staining further confirmed the association of p58(PITSLRE) with cyclin D3. This binding was observed only in the G(2)/M phase but not in the G(1)/S phase of the cell cycle; meanwhile, no interaction between p110(PITSLRE) and cyclin D3 was observed in all the cell cycle. The overexpression of cyclin D3 in 7721 cells leads to an exclusively accumulation of p58(PITSLRE) in the nuclear region, affecting its cellular distribution. Histone H1 kinase activity of p58(PITSLRE) was greatly enhanced upon interaction with cyclin D3. Furthermore, kinase activity of p58(PITSLRE) was found to increase greatly in the presence of cyclin D3 using a specific substrate, beta-1,4-galactosyltransferase 1. These data provide a new clue to our understanding of the cellular function of p58(PITSLRE) and cyclin D3.     

Outward-rectifying K+ channel activities regulate cell elongation and cell division of tobacco BY-2 cells

Potassium ions (K⁺) are required for plant growth and development, including cell division and cell elongation/expansion, which are mediated by the K⁺ transport system. In this study, we investigated the role of K⁺ in cell division using tobacco BY-2 protoplast cultures. Gene expression analysis revealed induction of the Shaker -like outward K⁺ channel gene, NTORK1 , under cell-division conditions, whereas the inward K⁺ channel genes NKT1 and NtKC1 were induced under both cell-elongation and cell-division conditions. Repression of NTORK1 gene expression by expression of its antisense construct repressed cell division but accelerated cell elongation even under conditions promoting cell division. A decrease in the K⁺ content of cells and cellular osmotic pressure in dividing cells suggested that an increase in cell osmotic pressure by K⁺ uptake is not required for cell division. In contrast, K⁺ depletion, which reduced cell-division activity, decreased cytoplasmic pH as monitored using a fluorescent pH indicator, SNARF-1. Application of K⁺ or the cytoplasmic alkalizing reagent (NH₄)₂SO₄ increased cytoplasmic pH and suppressed the reduction in cell-division activity. These results suggest that the K⁺ taken up into cells is used to regulate cytoplasmic pH during cell division.     

Sea urchin fertilization stimulates CaM kinase-II (multifunctional [type II] Ca2+/CaM kinase) activity and association with p34cdc2

Upon fertilization, the sea urchin egg synthesizes proteins which impart a Ca²⁺ dependence to M-phase onset. A potential target of this Ca²⁺ dependence may be CaM kinase-II (the multifunctional [type II] Ca²⁺/calmodulin [CaM]-dependent protein kinase) which is necessary for nuclear envelope breakdown in fertilized sea urchin eggs. This study was intended to determine whether sea urchin CaMK-II is activated after fertilization and whether it interacts with other known M-phase regulators, such as p34^cdc2. We report that total CaMK-II activity, measured by solution assays, increases after fertilization, peaking just prior to cleavage. Interestingly, total CaMK-II activity continues to fluctuate, peaking again prior to second and third cleavage. Gel assays also reveal enhanced levels of the 56 and 62 kDa potential CaMK-II phosphoproteins after fertilization. Finally, CaMK-II activity and only the 62 kDa phosphoprotein physically associate with p34^cdc2, but again only after fertilization. These changes in CaMK-II activity and p34^cdc2-association after fertilization may ensure that Ca²⁺ signals are targeted to the M-phase machinery at the appropriate developmental times.     

Two distinct classes of mitotic cyclin homologues, Cyc1 and Cyc2, are involved in cell cycle regulation in the ciliate Paramecium tetraurelia

The eukaryotic cell cycle is regulated by the sequential activation of different CDK/cyclin complexes. Two distinct classes of mitotic cyclin homologues, CYC1 and CYC2, have been identified and cloned for the first time in the ciliate Paramecium. Cyc1 is 324 amino acids long with a predicted molecular mass of 38 kDa, whereas Cyc2 is 336 amino acids long with a predicted molecular mass of 40 kDa. They display 42-51% sequence identity to other eukaryotic mitotic cyclins within the 'cyclin box' region. The conserved 'cyclin box' and 'destruction box' elements can be identified within each of the sequences. Genomic Southern blot analysis indicated that the CYC1 gene has two isoforms, with 92.3% and 85.9% identify at the amino acid level and at the nucleotide level, respectively. Both Cyc1 and Cyc2 proteins showed characteristic patterns of accumulation and destruction during the vegetative cell cycle, with Cyc1 peaking at the point of commitment to division (PCD), and Cyc2 reaching the maximal level late in the cell cycle. Immunoprecipitation experiments with antibodies specific to Cyc1 and Cyc2 indicated that Cyc1 and Cyc2 associate with distinct CDK homologues. Both immunoprecipitates exhibited histone H1 kinase activity that oscillated in the cell cycle in parallel with the respective amount of cyclins present. Histone H1 kinase activity associated with Cyc1 reached a peak at PCD while Cyc2 showed maximal activity when about 75% cells have completed cytokinesis. We propose that Cyc1 may be involved in commitment to division, in association with the CDK that binds to p13suc1, Cdk3, and that the Cyc2/Cdk2 complex may regulate cytokinesis. PCR-amplification revealed similar sequences in Tetrahymena, Sterkiella, Colpoda and Blepharisma, suggesting the conservation of the cyclin genes within ciliates. Although cell cycle regulation in ciliates differs in some respects from that of other eukaryotes, the cyclin motifs have clearly been conserved during evolution.     

The conserved C-terminal tail of FtsZ is required for the septal localization and division inhibitory activity of MinCC/MinD

The Escherichia coli Min system contributes to spatial regulation of cytokinesis by preventing assembly of the Z ring away from midcell. MinC is a cell division inhibitor whose activity is spatially regulated by MinD and MinE. MinC has two functional domains of similar size, both of which have division inhibitory activity in the proper context. However, the molecular mechanism of the inhibitory action of either domain is not very clear. Here, we report that the septal localization and division inhibitory activity of MinC^C/MinD requires the conserved C-terminal tail of FtsZ. This tail also mediates interaction with two essential division proteins, ZipA and FtsA, to link FtsZ polymers to the membrane. Overproduction of MinC^C/MinD displaces FtsA from the Z ring and eventually disrupts the Z ring, probably because it also displaces ZipA. These results support a model for the division inhibitory action of MinC/MinD. MinC/MinD binds to ZipA and FtsA decorated FtsZ polymers located at the membrane through the MinC^C/MinD–FtsZ interaction. This binding displaces FtsA and/or ZipA, and more importantly, positions MinC^N near the FtsZ polymers making it a more effective inhibitor.     

Functional Conservation of Calreticulin in Euglena gracilis

Calreticulin is the major high capacity, low affinity Ca²⁺ binding protein localized within the endoplasmic reticulum. It functions as a reservoir for triggered release of Ca²⁺ by the endoplasmic reticulum and is thus integral to eukaryotic signal transduction pathways involving Ca²⁺ as a second messenger. The early branching photosynthetic protist Euglena gracilis is shown to possess calreticulin as its major high capacity Ca²⁺ binding protein. The protein was purified, microsequenced and cloned. Like its homologues from higher eukaryotes, calreticulin from Euglena possesses a short signal peptide for endoplasmic reticulum import and the C-terminal retention signal KDEL, indicating that these components of the eukaryotic protein routing apparatus were functional in their present form prior to divergence of the euglenozoan lineage. A gene phytogeny for calreticulin and calnexin sequences in the context of eukaryotic homologues indicates i) that these Ca²⁺ binding endoplasmic reticulum proteins descend from a gene duplication that occurred in the earliest stages of eukaryotic evolution and furthermore iii that Euglenozoa express the calreticulin protein of the kinetoplastid (trypanosomes and their relatives) lineage, rather than that of the eukaryotic chlorophyte which gave rise to Euglena's plastids. Evidence for conservation of endoplasmic reticulum routing and Ca²⁺ binding function of calreticulin from Euglena traces the functional history of Ca²⁺ second messenger signal transduction pathways deep into eukaryotic evolution.     

Ca2+-Mediated Activation of c-Jun N-Terminal Kinase and Nuclear Factor κB by NMDA in Cortical Cell Cultures

Abstract: We examined the possibility that c-Jun N-terminal kinase (JNK) and nuclear factor κB (NF-κB) might be involved in intracellular signaling cascades that mediate NMDA-initiated neuronal events. Exposure of cortical neurons to 100 µ M NMDA induced activation of JNK within 1 min. Activity of JNK was further increased over the next 5 min and then declined by 30 min. Similarly, ionomycin, a selective Ca²⁺ ionophore, induced activation of JNK. The NMDA-induced activation of JNK was abrogated in the absence of extracellular Ca²⁺, suggesting that Ca²⁺ entry is necessary and sufficient for the JNK activation. Immunohistochemistry with anti-NF-κB antibody demonstrated nuclear translocation of NF-κB within 5 min following NMDA treatment. NMDA treatment also enhanced the DNA binding activity of nuclear NF-κB in a Ca²⁺-dependent manner. Treatment with 3 m M aspirin blocked the NMDA-induced activation of JNK and NF-κB. Neuronal death following a brief exposure to 100 µ M NMDA was Ca²⁺ dependent and attenuated by addition of aspirin or sodium salicylate. The present study suggests that Ca²⁺ influx is required for NMDA-induced activation of JNK and NF-κB as well as NMDA neurotoxicity. This study also implies that aspirin may exert its neuroprotective action against NMDA through blocking the NMDA-induced activation of NF-κB and JNK.     

Phosphorylation status of the NR2B subunit of NMDA receptor regulates its interaction with calcium/calmodulin-dependent protein kinase II

Ca²⁺ influx through NMDA-type glutamate receptor at excitatory synapses causes activation of post-synaptic Ca²⁺/calmodulin-dependent protein kinase type II (CaMKII) and its translocation to the NR2B subunit of NMDA receptor. The major binding site for CaMKII on NR2B undergoes phosphorylation at Ser1303, in vivo . Even though some regulatory effects of this phosphorylation are known, the mode of dephosphorylation of NR2B-Ser1303 is still unclear. We show that phosphorylation status at Ser1303 enables NR2B to distinguish between the Ca²⁺/calmodulin activated form and the autonomously active Thr286-autophosphorylated form of CaMKII. Green fluorescent protein–α-CaMKII co-expressed with NR2B sequence in human embryonic kidney 293 cells was used to study intracellular binding between the two proteins. Binding in vitro was studied by glutathione- S -transferase pull-down assay. Thr286-autophosphorylated α-CaMKII or the autophosphorylation mimicking mutant, T286D-α-CaMKII, binds NR2B sequence independent of Ca²⁺/calmodulin unlike native wild-type α-CaMKII. We show enhancement of this binding by Ca²⁺/calmodulin. Phosphorylation or a phosphorylation mimicking mutation on NR2B (NR2B-S1303D) abolishes the Ca²⁺/calmodulin-independent binding whereas it allows the Ca²⁺/calmodulin-dependent binding of α-CaMKII in vitro . Similarly, the autonomously active mutants, T286D-α-CaMKII and F293E/N294D-α-CaMKII, exhibited Ca²⁺-independent binding to non-phosphorylatable mutant of NR2B under intracellular conditions. We also show for the first time that phosphatases in the brain such as protein phosphatase 1 and protein phosphatase 2A dephosphorylate phospho-Ser1303 on NR2B.     

Sphingosine, W-7, and Trifluoperazine Inhibit the Elevation in Cytosolic Calcium Induced by High K+Depolarization in Synaptosomes

Abstract: A possible role for protein kinases in the regulation of free cytosolic Ca²⁺ levels in nerve endings was investigated by testing the effect of several kinase inhibitors on the increase in cytosolic Ca²⁺ (monitored with the Ca²⁺-sensitive dye fura-2) induced by depolarization with 15 or 30 mM K⁺. The ability of various drugs to inhibit the cytosolic Ca²⁺ response appeared to correlate with their reported mechanism of action in inhibiting protein kinases. W-7 and trifluoperazine, drugs reported to inhibit calmodulin-dependent events, were effective inhibitors of the increase in cytosolic Ca²⁺ induced by high K⁺ depolarization, as was sphingosine, a drug that inhibits protein kinase C by binding to the regulatory site, but which also inhibits calcium/calmodulin kinase. On the other hand, drugs that inhibit protein kinases by binding to the catalytic site, such as H-7 (1 m/W ), staurosporine (1μ M ), and K252_a(1μ M ), were ineffective. Activation of protein kinase C, which is blocked by each of these drugs, does not appear to be essential to the maintenance of elevated cytosolic Ca²⁺ in depolarized synaptosomes. All of the drugs, including sphingosine, that functionally inhibit the depolarization-induced elevation in cytosolic Ca²⁺ have in common the ability to bind to calmodulin. Because the drugs that inhibit protein kinases by competing with ATP binding at the active catalytic site did not block the response in this system, we suggest that a calmodulin or a calmodulin-like binding site participates in the regulation of Ca²⁺ increases after depolarization.     

Evidence For the Presence of A Cdc2-Like Protein Kinase In the Dinoflagellate Crypthecodinium Cohnii

ABSTRACT. The unusual nature of mitosis and ancestral organization of the dinoflagellate nucleus prompted the question of whether the cdc 2-like histone H1 kinase, a presumed ubiquitous cell cycle regulator in eukaryotes, is present in these primitive organisms. Western blotting of Crypthecodinium cohnii protein extracts using antibody against the Pro-Ser-Thr-Ala-Ile-Arg-Glu (=PSTAIRE) amino acid sequence motif, conserved in all cdc 2 homologues known, revealed one prominent band corresponding to a protein with an apparent relative molecular weight ≈ 34,000, identical in mobility to that from HeLa cells and Physarum polycephalum , higher and lower eukaryotic controls, respectively. Incubation of C. cohnii cell lysates with p13 ^{suc 1}-sepharose beads, which preferentially, though not exclusively, bind p34 ^{cdc 2}, resulted in precipitation of a 34-kDa protein which was reactive with anti-PSTAIRE antibody, selectively competed for by the PSTAIRE peptide and able to phosphorylate histone H1 in vitro. We conclude that the dinoflagellate C. cohnii contains a protein very similar to the cdc 2 gene product from fission yeast and its homologues in all eukaryotes studied thus far.     

Homo- and heterodimerization of ROCO kinases: LRRK2 kinase inhibition by the LRRK2 ROCO fragment

Mutations in the gene encoding leucine-rich repeat kinase 2 (LRRK2) are the most common cause of autosomal-dominant familial and late-onset sporadic Parkinson's disease (PD). LRRK2 is a large multi-domain protein featuring a GTP-binding C-terminal of Ras of complex proteins (ROC) (ROCO) domain combination unique for the ROCO protein family, directly followed by a kinase domain. Dimerization is a well-established phenomenon among protein kinases. Here, we confirm LRRK2 self-interaction, and provide evidence for general homo- and heterodimerization potential among the ROCO kinase family (LRRK2, LRRK1, and death-associated protein kinase 1). The ROCO domain was critically, though not exclusively involved in dimerization, as a LRRK2 deletion mutant lacking the ROCO domain retained dimeric properties. GTP binding did not appear to influence ROCO^LRRK2 self-interaction. Interestingly, ROCO^LRRK2 fragments exerted an inhibitory effect on both wild-type and the elevated G2019S LRRK2 autophosphorylation activity. Insertion of PD mutations into ROCO^LRRK2 reduced self-interaction and led to a reduction of LRRK2 kinase inhibition. Collectively, these results suggest a functional link between ROCO interactions and kinase activity of wild-type and mutant LRRK2. Importantly, our finding of ROCO^LRRK2 fragment-mediated LRRK2 kinase inhibition offers a novel lead for drug design and thus might have important implications for new therapeutic avenues in PD.     

Calmodulin and calmodulin-mediated processes in plants

Abstract. The Ca²⁺ -binding protein calmodulin is found in all plants investigated so far. The comparison of the biochemical and functional properties reveals that it is structurally conserved and functionally preserved throughout the plant and animal kingdom. Among the plant enzymes so far known to be dependent on the Ca²⁺ -calmodulin complex are NAD kinase(s), Ca²⁺ -transport ATPase, quinate: NAD⁺ oxidoreductase, soluble and membrane bound protein kinases, and H⁺ -transport ATPase. Calmodulin may play also an important role in the regulation of other cellular reactions, such as hormone-mediated processes, secretion of enzymes, and contractile mechanisms. On the basis of the NAD kinase and its regulation by light and Ca²⁺ -calmodulin, it is suggested that changes in the cellular, free Ca²⁺ concentration following stimulation may alter the metabolism of a plant cell. According to this suggestion free Ca²⁺ may act as a second messenger in plants much as it does in animal cells.     

Identification of novel suppressors for Mog1 implies its involvement in RNA metabolism, lipid metabolism and signal transduction

Mog1 is conserved from yeast to mammal, but its function is obscure. We isolated yeast genes that rescued a temperature-sensitive death of S. cerevisiae Scmog1Δ, and of S. pombe Spmog1^ts. Scmog1Δ was rescued by Opi3p, a phospholipid N-methyltransferase, in addition to S. cerevisiae Ran-homologue Gsp1p, and a RanGDP binding protein Ntf2p. On the other hand, Spmog1^ts was rescued by Cid13 that is a poly (A) polymerase specific for suc22⁺ mRNA encoding a subunit of ribonucleotide reductase, Ssp1 that is a protein kinase involved in stress response pathway, and Crp79 that is required for mRNA export, in addition to Spi1, S. pombe Ran-homologue, and Nxt2, S. pombe homologue of Ntf2p. Consistent with the identification of those suppressors, lack of ScMog1p dislocates Opi3p from the nuclear membrane and all of Spmog1^ts showed the nuclear accumulation of mRNA. Furthermore, SpMog1 was co-precipitated with Nxt2 and Cid13.