| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | ADVANCED SEARCH | TABLE OF CONTENTS |
|
|
|
|||||||
|
|||||||||
1 Department of Biology, Graduate School of Science, Osaka City University, Sugimoto, Sumiyoshi-ku, Osaka 558-8585, Japan
2 Kazusa DNA Research Institute, 2-6-7 Kazusa-kamatari, Kisarazu, Chiba 292-0818, Japan
3 Department of Integrated Biosciences, Graduate School of Frontier Science, University of Tokyo, Kashiwa, Chiba 277-8562, Japan
 |
Abstract |
|---|
 Top Abstract IntroductionResultsDiscussionExperimental proceduresReferences |
|---|
-tubulin complex, was also present in about 40% of the pseudo-SPBs. The forespore membranes initiated from both the SPBs and the pseudo-SPBs. We conclude that Spo20 plays a role in maintaining the structural integrity of the meiotic SPB, besides supplying membrane vesicles for forespore membrane assembly. |
Introduction |
|---|
|
TopAbstractIntroductionResultsDiscussionExperimental proceduresReferences |
|---|
Assembly of the FSM and meiotic nuclear divisions must proceed coordinately for accurate distribution of a complete haploid genome set into spores. A candidate for the key structure linking these two events is the spindle pole body (SPB), which is functionally equivalent to the centrosome in animal cells. During meiosis II, the SPB undergoes a structural modification from a single plaque to a multilayered expanded structure (Hirata & Tanaka 1982; Tanaka & Hirata 1982), and immunofluorescence microscopy using antibodies specific for Sad1 (a component of the SPB) shows that the SPB changes its morphology from a dot to a crescent shape (Hagan & Yanagida 1995; Ikemoto et al. 2000). Formation of the FSM starts on the cytoplasmic face of the modified SPB during meiosis II. A mutation of the spo15+ gene, encoding a constitutive component of the SPB, abolishes both modification of the SPB and formation of the FSM (Ikemoto et al. 2000). These observations suggest that the SPB plays a crucial role in the spatiotemporal coordination of meiosis and sporulation.
Not much is known about the origin of the small vesicles from which the FSM is assembled or the mechanism that controls vesicle fusion to construct the FSM. Our recent studies and those done by other groups have indicated that a general protein secretion machinery may be involved in FSM assembly. Sec12 is responsible for vesicle transport from the endoplasmic reticulum (ER) to the Golgi apparatus by activating the Sar1 GTPase in S. cerevisiae. The S. pombe Sec12 homologue, Spo14, is necessary for proper construction of the FSM (Nakamura-Kubo et al. 2003). Spo20 is structurally and functionally related to the major S. cerevisiae phosphatidylinositol (PI)/phosphatidylcholine (PC)-transfer protein Sec14, which is required for vesicle formation from the Golgi apparatus (Bankaitis et al. 1989, 1990). Fission yeast Spo20 may function in a similar manner to Sec14 of S. cerevisiae. In spo20-KC104 cells, the FSM grows poorly and fails to encapsulate the haploid nuclei. These results indicate that Spo20 regulates formation of the FSM, in addition to its known roles in post-Golgi vesicle trafficking (Nakase et al. 2001).
In budding yeast, several late-acting SEC genes (Novick et al. 1981), including SEC1, SEC4 and SEC8, are required for sporulation (Neiman 1998). The membrane fusion machinery, composed of soluble NSF attachment protein receptor (SNARE) proteins, governs the specificity of docking and fusion between vesicles and target membranes (Rothman & Orci 1992; Sollner et al. 1993; Pelham 1999; McNew et al. 2000). Another sporulation-specific membrane protein of S. pombe, Spo3, which is localized to the FSM, is also essential for construction of the FSM. One of the spo3 alleles, spo3-KC51, is dose-dependently suppressed by psy1+, which encodes a protein similar to mammalian syntaxin-1A, a component of the plasma membrane docking/fusion complex (Nakamura et al. 2001). These findings imply that precursor vesicles for the FSM are provided through a general secretory pathway and are recruited to the growing FSM by means of the SNARE complex.
In this study, we report a novel mutant allele of the spo20 gene, spo20-H6. This mutant shows an unexpected phenotype: there is an excess of meiotic SPBs, some of which are not located at the spindle poles. We have named these irregular structures ‘pseudo-SPBs’, because they contain regular SPB components but lack spindle assembly activity during meiotic nuclear divisions. These pseudo-SPBs probably split off from the functional SPBs at the spindle pole. Meiotic progression in the spo20-H6 mutant is delayed, but the segregation of sister chromatids is normal. These findings suggest that the Sec14 family of PI/PC-transfer proteins is also required for the structural integrity of meiotic SPBs in fission yeast.
|
Results |
|---|
|
TopAbstractIntroductionResultsDiscussionExperimental proceduresReferences |
|---|
Sporulation-defective mutants of the fission yeast S. pombe have been isolated (Okazaki et al. 2000). Among them, the H6 mutant harbours a single recessive mutation of the spo20 locus, and the mutant allele was designated spo20-H6. In addition to sporulation defect, this mutant was unable to grow at temperatures higher than 33 °C (data not shown). This temperature-sensitive phenotype of spo20-H6 was more severe than that of previously reported spo20-KC104 mutants, which poorly proliferated at 33 °C.
Sequencing of the spo20-H6 allele elucidated that it has a single nucleotide change from C to T, which causes the replacement of threonine 170 with isoleucine. This threonine residue is well conserved in the Sec14 family, and is positioned in the bottom of the hydrophobic pocket formed by the six-stranded ß-sheet (Sha et al. 1998). Therefore, the spo20-H6 mutation is likely to compromise its PI/PC-transfer activity.
The abundance of expressed mutant protein, designated Spo20T170I, did not differ from that of the wild-type protein (data not shown). We then examined the subcellular localization of Spo20T170I during proliferation and meiosis/sporulation by using wild-type cells expressing a Spo20T170I-HA fusion protein. Spo20T170I-HA localized at the cell periphery and septa during vegetative growth; after the induction of meiosis, Spo20T170I first accumulated in the nucleus and then translocated to the FSM after meiosis II (data not shown). These localization patterns are the same as the wild-type Spo20 proteins (Nakase et al. 2001). Thus, we detected neither instability nor mislocalization of Spo20T170I.
Modification of the SPB in spo20-H6
During meiosis II, the SPB is structurally modified from a compact dot to a crescent morphology (Hagan & Yanagida 1995). This crescent morphology (cf. Fig. 1B, panels 4 and 5) may correspond to a SPB structure with multilayered outer plaques in meiotic cells that has been observed by electron microscopy (Hirata & Tanaka 1982; Tanaka & Hirata 1982). Such structural modification of the SPB is likely to be essential for FSM formation (Hirata & Shimoda 1994; Ikemoto et al. 2000). We therefore addressed whether the SPB undergoes this morphological modification during meiosis II in the spo20-H6 mutant by using a GFP-tagged SPB marker protein, Spo15-GFP. The SPB underwent structural modification to the crescent-shaped morphology during meiosis II; however, the frequency of the modified SPB structures was appreciably lower in the spo20-H6 mutant than in wild-type cells (Fig. 1B). This conclusion was corroborated by indirect immunofluorescence microscopy using the antibody against another SPB component Sad1. Electron microscopic observations of spo20-H6 cells showed that the SPB underwent structural modifications to develop meiotic outer plaques, though the outermost layers were less evident (Fig. 1A). It is thus possible that the spo20-H6 mutation affects the process of SPB morphogenesis, although the significance of this effect is still elusive.
|
Visualization of the SPB structure in the spo20-H6 mutant unveiled an unexpected trait of the meiotic apparatus. The SPB acts as the spindle microtubule-organizing centre during meiosis; in wild-type cells, it is located only at both ends of the spindles in meiosis I and meiosis II. spo20-H6 cells expressing Spo15-GFP signals were induced to meiosis, and fixed cells were immunostained with an anti-
-tubulin antibody, TAT-1, to visualize spindles. Surprisingly, in a considerable fraction of spo20-H6 cells, Spo15-GFP were observed at sites other than the tips of the spindle microtubules (Fig. 1B). We refer to these SPB-like structures as ‘pseudo-SPBs’. Pseudo-SPBs were not observed in vegetative cells (Fig. 1C), although they were often found in cells through meiosis I and meiosis II. As shown in Fig. 1B, the pseudo-SPBs were not found in cells with a horsetail nucleus at prophase I, but were detected in cells at metaphase I and anaphase I.
To confirm that the pseudo-SPBs had not yet appeared at prophase I, we visualized SPB structures in the mei4
strain, which arrests at prophase I of meiosis (Horie et al. 1998). As shown in Fig. 1D, mei4 spo20-H6 double mutant cells that were arrested at prophase I displayed a single SPB. Quantitative assay showed that approximately 80% of spo20-H6 cells with meiosis I spindles had pseudo-SPBs, as detected by Spo15-GFP or anti-Cut12 antibodies; Cut12 is another SPB component (Table 1). As shown below, Spo15 and Cut12 were largely co-localized to the pseudo-SPB. The average number of pseudo-SPBs per cell as revealed by co-localization of Cut12/Spo15 or Cut12/Sid4 was determined to be 1.36 in spo20-H6 mutants. Taken together, these results suggest that pseudo-SPBs first appear at metaphase I.
|
The fact that these pseudo-SPBs appeared during meiosis suggested that meiotic nuclear division and sister chromatid segregation might be impaired by the spo20-H6 mutation. We therefore monitored meiotic nuclear divisions in a spo20-H6 homozygous diploid strain. A semisynchronous meiosis was induced by shifting log-phase cultures to sporulation medium (SSL-N) at 25 °C. The number of nucleus per cell was counted and the cells were categorized as mononucleate, binucleate and tetranucleate (Fig. 2A). In the spo20-H6 culture, a peak of binucleate cells was reached at 
10 h, about 2 h after the binucleate peak in the wild-type culture, suggesting the possibility that formation of meiotic spindles are influenced by the spo20-H6 mutation. To examine this possibility, we monitored spindle by visualizing microtubules by anti--tubulin antibody TAT-1 (Fig. 2B). In the mutant spo20-H6, cells entered prophase I (characterized by an elongated horsetail nucleus without spindles) in the similar kinetics to that of wild-type cells, but transition from prophase I to prometaphase/metaphase I (defined by a single compact nucleus with short spindles) was markedly delayed. Progression through metaphase I seemed retarded, while the second division was influenced to a lesser extent (Fig. 2B). Despite of these results, the final percentage of tetranucleate cells was fairly high (data not shown) and the morphology of spindles appeared to be normal (Fig. 1B). Together, the progression of the first meiotic division is moderately affected by the spo20-H6 mutation.
|
100% in the spo20-H6 strain (n = 200) (Fig. 2C), suggesting that there is no marked abnormality in the meiotic segregation of sister chromatids in the spo20-H6 mutant. Previously, we reported that ectopic expression of S. cerevisiae Sec14K66,239A complemented the sporulation defect of spo20-KC104 in S. pombe (Nakase et al. 2001). As Sec14K66,239A lacks PtdIns-transfer activity, sporulation-specific function of Spo20 might not rely on this transfer activity. Then we explored the effect of expression of Sec14K66,239A in spo20-H6 on integrity of the SPB during meiosis. The spo20-H6 mutant bearing the spo15-GFP on the chromosomal spo15 locus transformed with pREP2-based plasmids. In the transformant with vector plasmid pREP2, the percentage of cells forming no pseudo-SPB was only 15.8%. Expression of the wild-type Sec14 from pREP2(Sec14WT) appreciably increased such cells (60.7%). However, the effect of Sec14K66,239A was far below (42.9%) that of wild-type Sec14. These observations suggest that the PtdIns-transfer activity of Spo20 is necessary for the integrity of the SPBs during meiosis, although this activity is dispensable for the forespore membrane formation (Nakase et al. 2001).
Origin of the pseudo-SPBs
By time-lapse experiments, we explored in detail the origin of the pseudo-SPBs. Strain YN138, which expresses Spo15-GFP, was cultured in sporulation medium (MEA) for 16 h, and then Spo15-GFP was traced by time-lapse fluorescence microscopy. This analysis showed that each observed pseudo-SPB was originating from a normal SPB. The example shown in Fig. 3 clearly indicates that the pseudo-SPB separated from a normal SPB at metaphase I. Repeated observations indicated that most of the pseudo-SPBs appeared between prometaphase I and metaphase I. In wild-type cells, a pseudo-SPB was observed to split off from the normal SPB, but it was not stable (Fig. 3) and was probably reabsorbed by the SPB or degraded. The frequency of the pseudo-SPBs was lower in wild-type than in spo20-H6 cultures (Table 1).
|
We next determined whether the pseudo-SPBs contain major SPB component proteins by examining the localization of five known SPB components, Spo15, Sad1, Cut12, Sid4 and Spo13. Spo15 and Spo13 are known to be indispensable for structural modification of the SPB and for initiating FSM formation (Ikemoto et al. 2000; Y. Nakase, unpublished results), Sad1 and Cut12 are essential for bipolar spindle formation (Hagan & Yanagida 1995), and Sid4 is a major component of the septation initiation network (Chang & Gould 2000). Spo15, Spo13 and Sid4 were expressed as GFP fusion proteins, whereas Sad1 and Cut12 were visualized by indirect immunofluorescence microscopy with the respective antibodies. The spo20-H6 mutant strains were induced to meiosis and the SPB components were visualized in various combinations.
Fluorescence microscopy indicated that Spo15, Sad1, Cut12 and Spo13 were co-localized not only to the SPBs but also to the pseudo-SPBs (Fig. 4). Sid4 was co-localized with Sad1 and Cut12 in roughly half of the cells examined; these co-localization frequencies were significantly lower than those of any other combinations of components. Quantitative assays indicated that the co-localization of Cut12 and Spo15 reached 90% in the pseudo-SPBs, whereas only 65% of Cut12-containing pseudo-SPBs contained Sid4 (Table 2). These results indicate that Spo15, Cut12, Spo13 and Sad1 are standard constituents, whereas Sid4 is not always a component, of the pseudo-SPBs.
|
|
-tubulin complex. To test this possibility, we examined co-localization of Cut12 with a component of the -tubulin complex, Alp4 (Vardy & Toda 2000). As Table 2 and Fig. 4 indicate, the frequency of co-localization was much lower (c. 42%) than that of Cut12 and Spo15 (c. 92%). Ability of pseudo-SPBs to initiate FSMs
We monitored assembly of the FSM by using GFP-tagged Psy1, a syntaxin-like protein (Nakamura et al. 2001). As we previously reported, FSM formation was initiated normally in spo20 mutant cells during meiosis II; however, the FSM did not extend sufficiently and often failed to engulf the nucleus (Fig. 5A-II,III). We therefore investigated the fine structure of spo20-H6 zygotes, which completed meiosis II, by electron microscopy. Before analysis, both wild-type and mutant cells were incubated in sporulation medium (MEA) at 25 °C for 1 day.
|
The FSM originates on the cytoplasmic side of the SPB (Hirata & Tanaka 1982; Tanaka & Hirata 1982). Because spo20-H6 mutant cells produce pseudo-SPBs, we addressed whether these pseudo-SPBs are able to initiate formation of the FSM. Intriguingly, assembly of the FSM clearly started near the pseudo-SPBs in some cells (Fig. 5A-III). The frequency of this latter type of zygote reached 85% of the population (Fig. 5B-III). These results imply that the pseudo-SPBs have the potential to mediate FSM assembly.
|
Discussion |
|---|
|
TopAbstractIntroductionResultsDiscussionExperimental proceduresReferences |
|---|
The FSM is first formed on the cytoplasmic face of the SPBs in metaphase II. The FSM extends and eventually encapsulates each of the four nuclei. In the spo20-H6 mutant, the FSM was formed near both SPBs and pseudo-SPBs in meiosis II, but failed to continue its development in both cases. Why is formation of the FSM incomplete in spo20-H6? The S. pombe spo14+ gene encodes a homologue of S. cerevisiae Sec12, another component of the protein secretion pathway. Similar to spo20 mutants, the spo14-B221 mutant exhibits a sporulation defect; that is, the morphology of the SPBs alters to a crescent shape during meiosis II, but FSM development ceases at the halfway point. The fact that Sec mutants are defective in FSM assembly supports the notion that the vesicles needed for FSM formation are supplied through a general vesicle trafficking machinery. Aberrant FSM assembly in spo14 and spo20 mutants is probably caused by insufficient vesicle formation. As the spo20-H6 mutant has pseudo-SPBs in addition to the normal SPBs, these multiple FSM start sites may exaggerate the shortage of vesicles.
Apparently normal bipolar spindles were constructed in the first and second divisions, indicating that the pseudo-SPBs did not act as spindle poles. We have no information on how these pseudo-SPBs affect meiotic nuclear divisions. However, spo20-H6 mutants showed significant delays in the onset and progression of meiosis I. The FSM is formed during meiosis II, but notably, the spo20-H6 mutation affected meiosis II to a lesser extent than meiosis I. Thus, it is possible that the disturbance of meiotic nuclear divisions indirectly affects assembly of the FSM.
Our time-lapse microscopy images showed that the pseudo-SPBs appeared to originate by splitting off from a normal SPB. We therefore examined whether known SPB components were localized in the pseudo-SPBs. Of six SPB components tested, Spo13, Spo15, Sad1 and Cut12 were mostly localized in the pseudo-SPBs. A -tubulin complex protein Alp4 is also present in pseudo-SPBs, although a frequency of the Alp4-containing pseudo-SPBs was significantly low. Because even these Alp4-containing pseudo-SPBs are unable to assemble microtubules, the absence of the -tubulin complex is not directly related to microtubule assembly defect.
On the basis of the present study and our previous reports, we speculate that Spo20 has a role in regulating SPB organization. Spo20 is imported into the nucleus when cells are committed to meiosis. Possibly, the S. pombe PI/PC-transfer protein Spo20 is involved in phospholipid homeostasis of the nuclear envelope during meiosis and sporulation. Fluorescence microscopy revealed that Spo15-GFP is preferentially localized to the SPB, but faint fluorescent signals were also seen on the nuclear envelope. In particular, the time-lapse microscopy images raise the intriguing possibility that certain SPB components such as Spo15 both associate with and dissociate from the SPB (Fig. 3). Such dynamic aspects of SPB components may be influenced by the lipid composition of the nuclear envelope. If this is true, then mutations in spo20 would affect the physical integrity of the SPB by altering the nuclear envelope. As a procedure for preparing nuclear envelopes in a high purity has not established yet, this highly hypothetical notion remains to be investigated.
|
Experimental procedures |
|---|
|
TopAbstractIntroductionResultsDiscussionExperimental proceduresReferences |
|---|
The S. pombe strains used in this study are listed in Table 3. Complete medium YEA was used for growth. Malt extract medium, MEA, and synthetic sporulation media, SSA and SSL-N, were used for mating and sporulation. These media have been described by Egel & Egel-Mitani (1974), Gutz et al. (1974), and Moreno et al. (1990). In time-lapse experiments mannose synthetic medium MSM was used (Okazaki et al. 2000). S. pombe cells were grown at 30 °C and sporulated at 28 °C, except for the spo20 mutants, which were grown and sporulated at 25 °C. For visualization of the cen1 locus, we used the GFP-LacI and LacO repeats integrated near lys1 and his7, respectively (Nabeshima et al. 1998). For staining nucleoplasm, GFP-Fcp1C was expressed in the same cells. This fusion construct contains the C-terminal 77 amino acid residues of Fcp1 (Kimura et al. 2002). This fusion gene was integrated at the leu1 locus.
|
Plasmid pAL(spo13-GFP) was constructed as follows. The spo13 gene was amplified by PCR using 5'-CCCCTCGAG(XhoI)ATTGTAAGTTTATTACGCCG-3' and 5'-CCCGCGGCCGC(NotI)AGTGGTAATTTGATTCGTTAG-3' as forward and reverse primers, respectively. The PCR product was digested with XhoI and NotI, and then ligated into the same restriction sites of pTN143 (Ikemoto et al. 2000), yielding pAL (spo13-GFP).
Immunofluorescence microscopy
For cell fixation, we used glutaraldehyde and paraformaldehyde according to the procedure of Hagan & Hyams (1988). The SPB was visualized by indirect immunofluorescence microscopy with the following primary antibodies: rabbit anti-Sad1 antibody and rabbit anti-Cut12 antibody (gifts from O. Niwa, Kazusa DNA Research Institute and I. Hagan, University of Manchester, respectively). The secondary antibody was Alexa 546-conjugated goat anti-rabbit IgG (Molecular Probes, Eugene, OR, USA), used at a 1 : 1000 dilution. Because anti-Cut12 antibody is sensitive to glutaraldehyde, the cells were mixed with an equal volume of prewarmed SSL-N media plus 2.4 M sorbitol for 5 min prior to fixation in 4% paraformaldehyde. For microtubule staining, mouse anti--tubulin antibody TAT-1 (Woods et al. 1989) was used, followed by Cy3-conjugated anti-mouse IgG (Sigma Chemical Co., St. Louis, MO, USA) at a 1 : 1000 dilution. The nuclear chromatin region was stained with 4', 6-diamidino-2-phenylindole (DAPI) at 1 µg/mL. Stained cells were observed under a fluorescence microscope (model BX50 and model IX71; Olympus, Tokyo) equipped with a charge-coupled device (CCD) camera (Cool-SNAP; Roper Science, San Diego, CA and ORCA-ER; Hamamatsu Photonics, Hamamatsu, Japan).
Time-lapse observation
Microscopic observation was performed with the Delta Vision microscope system (Applied Precision, Issaquah, WA, USA), which allows the multicolor and three-dimensional acquisition of digitized images. For time-lapse observation, living cells scraped from an agar plate were suspended in 1 µg/mL Hoechst 33342 and incubated for 5 min. Cells were collected by centrifugation, suspended in MSM medium containing 1 µg/mL Hoechst 33342, and placed on to a thin film of 2% agarose gel. The film was sandwiched between a pair of coverslips and placed on the stage. Each set of images for GFP and Hoechst 33342 was obtained with a 2-s exposure of 490 nm (neutral density filter 10%) and a 0.2-s exposure of 380-nm excitations, respectively, at 25–26 °C. Several Z-axis sections at 0.3-µm intervals were combined by using the quick projection program. The colours in the merged images for each wavelength in this report are all artificial.
Electron microscopy
Cells were mounted on the copper grids to form a thin layer and plunged into liquid propane cooled with liquid N2. Frozen cells were transferred to 2% OsO4 in anhydrous acetone, kept at –80 °C for 48 h in a solid CO2-acetone bath, and then transferred to –35 °C for 2 h, 4 °C for 2 h, and room temperature for 2 h. After washing with anhydrous acetone three times, samples were infiltrated with increasing concentrations of Spurr's resin in anhydrous acetone and finally with 100% Spurr's resin. These samples were then polymerized at 50 °C for 5 h and 60 °C for 50 h. Thin sections were cut on a Reichest Ultracut S, and then stained with uranyl acetate and lead citrate. Sections were viewed on a JEOL 2010 electron microscope at 100 kV.
|
Acknowledgements |
|---|
-tubulin antibody, TAT-1; I.M. Hagan of the Paterson Institute for Cancer Research for affinity-purified antibodies against Cut12; M. Yanagida of Kyoto University for S. pombe strains; K. Gould for sid4-GFP strain; T. Toda of Cancer Research UK for alp4-GFP strain. This study was supported by a Sasakawa Scientific research grant (to Y.N.); the Japan Securities Scholarship Foundation to (Y.N.); a Grant-in-Aid for Scientific Research ‘B’ and Priority Area ‘Genome Biology’ from the Ministry of Education, Culture, Sports, Science, and Technology of Japan (to C.S.); and a Grant-in-Aid for Scientific Research ‘Cell Cycle Control’ and ‘Life of Proteins’ from the Ministry of Education, Culture, Sports, Science and Technology of Japan (to T.N.). |
Footnotes |
|---|
* Correspondence: E-mail: shimoda{at}sci.osaka-cu.ac.jp
|
References |
|---|
|
TopAbstractIntroductionResultsDiscussionExperimental proceduresReferences |
|---|
Bankaitis, V.A., Malehorn, D.E., Emr, S.D. & Greene, R. (1989) The S. cerevisiae SEC14 gene encodes a cytosolic factor that is required for transport of secretory proteins from the yeast Golgi complex. J. Cell Biol.
108, 1271–1281.
Chang, L. & Gould, K.L. (2000) Sid4p is required to localize components of the septation initiation pathway to the spindle pole body in fission yeast. Proc. Natl. Acad. Sci. USA
97, 5249–5254.
Ding, D.Q., Tomita, Y., Yamamoto, A., Chikashige, Y., Haraguchi, T. & Hiraoka, Y. (2000) Large-scale screening of intracellular protein localization in living fission yeast cells by the use of a GFP-fusion genomic DNA library. Genes Cells 5, 169–190.[Abstract]
Egel, R. & Egel-Mitani, M. (1974) Premeiotic DNA synthesis in fission yeast. Exp. Cell Res. 88, 127–134.[CrossRef][Medline]
Gutz, H., Heslot, H., Leupold, U. & Loprieno, N. (1974) S. pombe. In: Handbook of Genetics, Vol 1 (ed. R.C. King) pp. 395–446. New York: Plenum Press.
Hagan, I.M. & Hyams, J.S. (1988) The use of cell division cycle mutants to investigate the control of microtubule distribution in the fission yeast S. pombe. J. Cell Sci.
89, 343–357.
Hagan, I. & Yanagida, M. (1995) The product of the spindle formation gene sad1+ associates with the fission yeast spindle pole body and is essential for viability. J. Cell Biol.
129, 1033–1047.
Hirata, A. & Shimoda, C. (1994) Structural modification of spindle pole bodies during meiosis II is essential for normal formation of ascospores in S. pombe: ultrastructural analysis of spo mutants. Yeast 10, 173–183.[CrossRef][Medline]
Hirata, A. & Tanaka, K. (1982) Nuclear behavior during conjugation and meiosis in the fission yeast S. pombe. J. General Appl. Microbiol. 28, 263–274.
Horie, S., Watanabe, Y., Tanaka, K., et al. (1998) The S. pombe mei4+ gene encodes a meiosis-specific transcriptional factor containing a forkhead DNA-binding domain. Mol. Cell. Biol.
18, 2118–2129.
Ikemoto, S., Nakamura, T., Kubo, M. & Shimoda, C. (2000) S. pombe sporulation-specific coiled-coil protein Spo15p is localized to the spindle pole body and essential for its modification. J. Cell Sci. 113, 545–554.[Abstract]
Kimura, M., Suzuk, H. & Ishihama, A. (2002) Formation of a carboxy-terminal domain phosphatase (Fcp1)/TFIIF/RNA polymerase II (pol II) complex in Schizosaccharomyces pombe involves direct interaction between Fcp1 and the Rpb4 subunit of pol II. Mol. Cell. Biol.
22, 1577–1588.
McNew, J.A., Parlati, F., Fukuda, R., et al. (2000) Compartmental specificity of cellular membrane fusion encoded in SNARE proteins. Nature 407, 153–159.[CrossRef][Medline]
Moreno, S., Klar, A. & Nurse, P. (1990) Molecular genetic analysis of fission yeast S. pombe. Methods Enzymol. 194, 793–823.
Nabeshima, K., Nakagawa, T., Straight, A.F., et al. (1998) Dynamics of centromeres during metaphase-anaphase transition in fission yeast: Dis1 is implicated in force balance in metaphase bipolar spindle. Mol. Biol. Cell
9, 3211–3225.
Nakamura, T., Nakamura-Kubo, M., Hirata, A. & Shimoda, C. (2001) The S. pombe spo3+ gene is required for assembly of the forespore membrane and genetically interacts with psy1+ encoding syntaxin-like protein. Mol. Biol. Cell
12, 3955–3972.
Nakamura-Kubo, M., Nakamura, T., Hirata, A. & Shimoda, C. (2003) The fission yeast spo14+ gene encoding a functional homologue of budding yeast Sec12 is required for the development of forespore membranes. Mol. Biol. Cell
14, 1109–1124.
Nakase, Y., Nakamura, T., Hirata, A., et al. (2001) The S. pombe spo20+ gene encoding a homologue of S. cerevisiae Sec14 plays an important role in forespore membrane formation. Mol. Biol. Cell
12, 901–917.
Neiman, A.M. (1998) Prospore membrane formation defines a developmentally regulated branch of the secretory pathway in yeast. J. Cell Biol.
140, 29–37.
Novick, P., Field, C. & Schekman, R. (1981) Order of events in the yeast secretory pathway. Cell 25, 461–469.[CrossRef][Medline]
Okazaki, K., Okayama, H. & Niwa, O. (2000) The polyubiquitin gene is essential for meiosis in fission yeast. Exp. Cell Res. 254, 143–152.[CrossRef][Medline]
Pelham, H.R. (1999) SNAREs and the secretory pathway—lessons from yeast. Exp. Cell Res. 247, 1–8.[CrossRef][Medline]
Rothman, J.E. & Orci, L. (1992) Molecular dissection of the secretory pathway. Nature 355, 409–415.[CrossRef][Medline]
Sha, B., Phillips, S.E., Bankaitis, V.A. & Luo, M. (1998) Crystal structure of the S. cerevisiae phosphatidylinositol-transfer protein. Nature 391, 506–510.[CrossRef][Medline]
Sollner, T., Whiteheart, S.W., Brunner, M., et al. (1993) SNAP receptors implicated in vesicle targeting and fusion. Nature 362, 318–324.[CrossRef][Medline]
Tanaka, K. & Hirata, A. (1982) Ascospore development in the fission yeasts S. pombe and S. japonicus. J. Cell Sci.
56, 263–279.
Vardy, L. & Toda, T. (2000) The fission yeast -tubulin complex is required in G1 phase and is a component of the spindle assembly checkpoint. EMBO J.
19, 6098–6111.[CrossRef][Medline]
Woods, A., Sherwin, T., Sasse, R., MacRae, T.H., Baines, A.J. & Gull, K. (1989) Definition of individual components within the cytoskeleton of Trypanosoma brucei by a library of monoclonal antibodies. J. Cell Sci.
93, 491–500.
Yoo, B.Y., Calleja, G.B. & Johnson, B.F. (1973) Ultrastructural changes of the fission yeast (S. pombe) during ascospore formation. Arch. Microbiol. 91, 1–10.
Received: 10 May 2004
Accepted: 17 September 2004
This article has been cited by other articles:
|
|
 |
|
  T. Nakamura, H. Asakawa, Y. Nakase, J. Kashiwazaki, Y. Hiraoka, and C. Shimoda Live Observation of Forespore Membrane Formation in Fission Yeast Mol. Biol. Cell, August 1, 2008; 19(8): 3544 - 3553. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 A. Yamamoto, K. Kitamura, D. Hihara, Y. Hirose, S. Katsuyama, and Y. Hiraoka Spindle checkpoint activation at meiosis I advances anaphase II onset via meiosis-specific APC/C regulation J. Cell Biol., July 28, 2008; 182(2): 277 - 288. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
Y. Nakase, M. Nakamura-Kubo, Y. Ye, A. Hirata, C. Shimoda, and T. Nakamura Meiotic Spindle Pole Bodies Acquire the Ability to Assemble the Spore Plasma Membrane by Sequential Recruitment of Sporulation-specific Components in Fission Yeast Mol. Biol. Cell, June 1, 2008; 19(6): 2476 - 2487. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
Y. Ye, M. Fujii, A. Hirata, M. Kawamukai, C. Shimoda, and T. Nakamura Geranylgeranyl Diphosphate Synthase in Fission Yeast Is a Heteromer of Farnesyl Diphosphate Synthase (FPS), Fps1, and an FPS-like Protein, Spo9, Essential for Sporulation Mol. Biol. Cell, September 1, 2007; 18(9): 3568 - 3581. [Abstract] [Full Text] [PDF]
|
|
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | ADVANCED SEARCH | TABLE OF CONTENTS |
