As of 2001, a total of 89 ran expressed sequence tags (ESTs) have been reported in the Berkeley Drosophila Genome Project database while only a single ran-like EST has been identified. The ran like EST (GH25818) was isolated from an adult head cDNA library. In comparison, ran ESTs have been isolated from both embryo and adult tissues (61, embryo; 18, adult head; and 1 cell culture). The differences in the number of reported ran and ran-like ESTs suggest that the two genes may have different spatial/temporal expression dynamics during development. To test this hypothesis, their embryonic expression patterns were tested on 0- to 18-h embryo collections via whole-mount in situ hybridizations (Koizumi, 2001).
Localization of ran mRNA during embryonic development revealed two temporal phases of expression. Initially, ran expression is found in the pre-cellular blastoderm, suggesting that it is supplied maternally. Localization of ran transcripts in progressively older embryos revealed that shortly after the onset of gastrulation, there is a significant and rapid loss of ran mRNA throughout the embryo. By embryonic stage 7, no expression was observed. The second temporal phase of ran expression occurs in CNS neuroblasts but only during their late sublineage-forming mitotic divisions. No expression was observed in NBs or any other cell during stages 8-11 indicating that ran gene expression is restricted from NBs during the formation of their early sublineages. During germ-band contraction (stage 12), ran transcripts were detected in large-diameter cells that line the cephalic lobes and the ventral/ventral-lateral surface of the developing ventral cord ganglia. The size and location of ran-expressing cells suggest that they are predominantly NBs. By stage 16, ran transcripts were detected in only a few putative neural precursor cells (NBs and/or ganglion mother cells). The fact that ran CNS expression is progressively reduced, starting at early stage 14, further suggests that its expression is predominately restricted to NBs and ganglion mother cells and not to post-mitotic neurons or glia. In situ hybridization of third instar larval tissues with ran -specific probes reveals that ran is also expressed ubiquitously, albeit at low levels, in both larval tissues and imaginal discs (Koizumi, 2001).
ran-like in situ hybridization analysis of embryos, obtained from overnight collections, also identified a maternal and zygotic pattern of gene expression. Similar to ran, ran-like transcripts were detected uniformly distributed in syncytial blastoderm embryos, and starting just after cellularization there was a rapid decline in the ran-like in situ signal. By stage 5, little or no ran-like in situ staining was observed. However, differing from the neuronal precursor gene expression pattern of ran, zygotic ran-like expression was detected in salivary gland cells and in the epithelial cells lining the tracheal tubular network (Koizumi, 2001).
The GTPase Ran regulates multiple cellular functions throughout the cell cycle, including nucleocytoplasmic transport, nuclear membrane assembly, and spindle assembly. Ran mediates spindle assembly by affecting multiple spindle assembly pathways: microtubule dynamics, microtubule motor activity, and spindle pole assembly. Ran is predicted to facilitate spindle assembly by remaining in the GTP-bound state around the chromatin in mitosis. This study directly tests the central tenet of this hypothesis in vivo by determining the cellular localization of Ran pathway components in Drosophila embryos. During mitosis, RCC1, the nucleotide exchange factor for Ran, is associated with chromatin, while Ran and RanL43E, an allele locked in the GTP-bound state, localizes around the spindle. In contrast, nuclear proteins redistribute throughout the embryo upon nuclear envelope breakdown (NEB). Thus, in vivo RanGTP has the correct spatial localization within the cell to modulate spindle assembly (Trieselmann, 2002).
To investigate the significance of ran-restricted CNS expression, its misexpression was targeted to different temporal windows of CNS development. In addition, a dominant- negative mutant form of ran was targeted to the developing CNS and to the larval eye/antenna imaginal disc to assess the role of ran-dependent functions. Embryonic CNS misexpression of the mutant, but not wild-type, ran results in larval death. Neither wild-type nor mutant ran misexpression had any detectable effect on embryonic CNS lineage specification, nuclear transport of a number of CNS-specific transcription factors or axonal guidance. However, expression of the dominant-negative mutant ran in the developing eye/antenna disc did result in a severe adult eye phenotype marked by apoptosis of photoreceptor, cone and pigment cells (Koizumi, 2001).
Blocking Ran function(s) in mammalian cultured cells has been achieved via the misexpression of different dominant-negative mutant forms of Ran. The mutant Ran proteins have single or double amino acid substitutions (G19V and/or Q69L) that alter the Ran protein in such a way that it can bind but not hydrolyze GTP (Coutavas, 1993; Ren, 1994; Lounsbury, 1996). These dominant-negative mutant alleles have been shown to have profound effects on multiple nuclear events, including progression of the cell cycle, RNA transport, and DNA replication (Ren, 1994; Melchior, 1998).
The restricted expression of ran to CNS NBs undergoing late lineage-generating divisions and its absence from post-mitotic neurons or glia suggests that ran may be required for a temporally defined event in neural development. Alternatively, ran may be required in all neural cells, but due to the high rate of NB mitotic divisions, the maternal supply is insufficient for CNS stem cells to complete their lineage development (Koizumi, 2001).
As an initial step toward determining the significance of the temporally restricted CNS expression, the effects of ran misexpression outside the temporal boundaries of its wild-type expression window was studied. The effects of the dominant-negative ran mutant, ranG19V;Q69L, on different aspects of embryonic development and on adult eye development was studied. The targeted ectopic expression was accomplished by using the yeast Gal4/UAS system. Both wild-type and mutant ran ORFs were inserted into the P-element based UAS vector to create the following constructs: P[UAS.ranwild-type] and P[UAS.ranG19V;Q69L], respectively. Multiple independent transformant lines were generated with the above P-element constructs with integration sites on either the second or third chromosomes. For misexpression in the developing nervous system, Gal4 drivers were used that can activate the expression of the UAS transgenes in either NBs or post-mitotic neurons. For NB expression, pros.Gal4 and pdm.Gal4 drivers were used; for post-mitotic neurons an elav.Gal4 driver was used. Pan-neural embryonic transgene expression was also achieved using scabrous. Gal4. The effects of UAS.ranwild-type and UAS.ranG19V;Q69L misexpression on early segmentation was also studied using a hairy.Gal4 driver. The effects of wild-type and mutant ran misexpression were assessed by three criteria: viability, changes in the sub-cellular localization of nuclear factors, and defects/changes in nervous system structure as assessed by axonal patterning. To confirm that these Gal4 drivers activated the different UAS.ran transgenes, misexpression was monitored via in situ hybridization (Koizumi, 2001).
The misexpression of wild-type ran with the Gal4 drivers used in this study did not result in any detectable abnormal phenotype in either the CNS or in other aspects of Drosophila biology. However, expression of UAS.ranG19V;Q69L under the control of each of the above Gal4 drivers, with the exception of the scabrous driver, resulted in either late first, early second instar larval or pupal mortality. No abnormal behavioral phenotypes were detected in the first instar larva harboring any of the different promoter-driven mutant ran transgene combinations. After hatching from the egg case, larva exhibited wildtype motility and eating behavior. The prosperoGal4/UAS.ranG19V;Q69L transformant adults failed to exit the pupa case and inspection of the pharate adults failed to identify any morphological defects. The scabrous.Gal4/UAS.ranG19V;Q69L transgene combination proved to be viable (Koizumi, 2001).
To determine if ectopic expression of the wild-type or mutant ran transgenes leads to defects in the subcellular distribution of developmentally regulated nuclear proteins, embryos were stained with antibodies to the Castor, Pdm1, Hb, Prospero and Even-skipped transcription factors. Immunostains of all Gal4/UAS combinations (both wild-type and mutant Ran) failed to reveal any defects in the subcellular distribution of these proteins. To assess if other aspects of neural development were affected, such as axonal outgrowth, axon markers (monoclonal antibodies BP102 and 22C10) were employed to immunostain embryos containing the different Gal4/UAS combinations. Again, within the resolution of this analysis, no defects in axonal outgrowth or axon fascicle development during embryonic development were detected in any of the Gal4/UAS combinations tested. However, the possibility can not be ruled out that misexpression of mutant ran triggers an embryonic defect, albeit subtle, which ultimately results in larval lethality (Koizumi, 2001).
Given the lack of detectable abnormal phenotype accompanying the wild-type or mutant ran transgene expression, the possibility exists that the levels of transgene expression were not sufficient to trigger cellular or developmental embryonic defects. To address this possibility, transformants were generated that contained multiple wild-type or mutant Ran Gal4/UAS transgene combinations. Analysis of these lines revealed no detectable differences between embryos and larva harboring single or multiple copies of the Gal4/UAS combinations. Nevertheless, embryonic expression of Ran G19V;Q69L does produce larval and pupal lethality, confirming that the Gal4/UAS.ranG19V;Q69L transgene combinations did in fact work. The data suggest that larval cells and/or postmitotic neurons may have a greater requirement for Ran dependent processes. Currently it is not known why the effects of the ectopic expression are delayed. Perhaps the high levels of endogenous Ran expression are sufficient to overcome the immediate effects of the different Gal4 driven UAS.ranG19V;Q69L combinations. It is also possible that other ran -independent nucleo-cytoplasmic transport mechanisms may compensate for a deficiency in ran function during embryonic development. Ran-independent nuclear transport has been observed in other systems (Nakielny, 1997; Fagotto, 1998; Sachdev, 2000; Koizumi, 2001 and references therein).
During embryonic development, perdurance of Ran function from the maternal expression of both ran genes may be sufficient to over-ride the misexpression of the dominant-negative mutant Ran. To avoid this possibility, we assessed the effects of ran misexpression during larval development in imaginal cells that should have little or no maternal contribution. To accomplish this, the misexpression of wild-type and mutant ran was targeted to different stages of adult eye development. The targeted misexpression of UAS.ran transgenes to the eye-imaginal disc takes advantage of the fact that the eye is not essential for viability and that disruption of the ommatidial organization is a sensitive indicator of phenotypic effects. In addition, different Gal4 drivers are available to target misexpression within different temporal windows of eye development. Three different spatial/temporal eye disc Gal4 drivers were employed: (1) dppdisc-.Gal4 to target misexpression to undifferentiated cells within the morphogenetic furrow, (2) pGMR.Gal4 to target all cells behind the morphogenetic furrow, and (3) sevenless.Gal4 to drive misexpression in a subset of differentiating photoreceptor cells and non-neuronal cone and pigment cells. As in the embryo, the ectopic misexpression of wildtype ran during eye development resulted in no detectable mutant phenotype. In all cases the ommatidial organization of the various Gal4/UAS.ran combinations was indistinguishable from wild-type eyes. In addition, the misexpression of mutant ran in retinal precursors within the morphogenetic furrow, P[dppdisc.Gal4]/P[UAS.ranG19V;Q69L], did not induce detectable changes in eye development. Expression of Ran G19V;Q69L in developing photoreceptors using a sevenless promoter resulted in an eye phenotype: close examination of eye cross-sections showed a disrupted ommatidial structure with absence of photoreceptor cells including the R7 cell. Expression of mutant ran behind the morphogenetic furrow via the P[GMR.Gal4] transgene resulted in a severe eye phenotype characterized by a complete loss of photoreceptor, cone and pigment cells. The severe phenotype was partially reverted in the presence of a constitutively active p35 genetic background. p35 has been shown to prevent programmed cell death in Drosophila, and acts by binding to and inhibiting the proteolytic activity of caspases. The partial rescue suggests that the loss of ommatidial structures is caused by programmed cell death. However, misexpression of mutant ran via the GMR promoter did not appear to affect the development of interommatidial bristles (Koizumi, 2001).
Interommatidial bristles are mechanosensory organs composed of four cells that are derived from a single sensory organ precursor (SOP). Unlike the photoreceptor and cone cells, the appearance of SOPs is not related to the distance from the morphogenetic furrow. The final cell divisions generating the interommatidial bristles take place between 6 h and 15 h after puparium formation. Photoreceptor cell determination occurs behind the furrow, during third instar development. Because all eye lineages come from equivalent precursors, cell death induction due to Ran G19V; Q69L expression occurs after the transit of the morphogenetic furrow and subsequent to or parallel to the induction of interommatidial bristles. It is not clear from these results whether the bristles cells are more resistant to RanG19V;Q69L or whether Ran is specifically required for a transport step in photoreceptor cells that does not take place in bristle cells (Koizumi, 2001).
RanGTP is important for chromosome-dependent spindle assembly in Xenopus extracts. This study reports on experiments to determine the role of the Ran pathway on microtubule dynamics in Drosophila oocytes and embryos. Females expressing a dominant-negative form of Ran have fertility defects, suggesting that RanGTP is required for normal fertility. This is not, however, because of a defect in acentrosomal meiotic spindle assembly. Therefore, RanGTP does not appear to be essential or sufficient for the formation of the acentrosomal spindle. Instead, the most important function of the Ran pathway in spindle assembly appears to be in the tapering of microtubules at the spindle poles, which might be through regulation of proteins such as TACC and the HURP homolog, Mars. One consequence of this spindle organization defect is an increase in the nondisjunction of achiasmate chromosomes. However, the meiotic defects are not severe enough to cause the decreased fertility. Reductions in fertility occur because RanGTP has a role in microtubule assembly that is not directly nucleated by the chromosomes. This includes microtubules nucleated from the sperm aster, which are required for pronuclear fusion. It is proposed that following nuclear envelope breakdown, RanGTP is released from the nucleus and creates a cytoplasm that is activated for assembling microtubules, which is important for processes such as pronuclear fusion. Around the chromosomes, however, RanGTP might be redundant with other factors such as the chromosome passenger complex (Cesario, 2011).
The Ran pathway has a variety of targets, which leads to effects on kinetochores, centrosomes and microtubule-associated proteins(Kalab, 2008). RanGTP is potentially an important molecule for spindle assembly in acentrosomal oocytes because it has been identifiedas a key factor for chromatin-induced spindle formation in Xenopus extracts. Surprisingly, the current results suggest that RanGTP might be more important for microtubule assembly in other circumstances, such as when centrosomes are present or when microtubules assemble without direct contact with the chromosomes (Cesario, 2011).
Diffusion of RanGTP from its source, the chromatin, into the cytoplasm, where it is converted into RanGDP, can create a gradient that regulates microtubule organization. Drosophila oocytes contain two key regulators of the Ran pathway in distinct locations. RCC1, as expected, is located tightly around the karyosome in mature oocytes. RanGAP localization is more complex than expected because it is present in many clusters, possibly vesicles, within the oocyte, suggesting that conversion of RanGTP to RanGDP might be regulated and only occur in certain locations. This could mean that a gradient of RanGTP is not established in the oocyte. A candidate protein responsible for generating the concentrations of RanGAP is Ran binding protein 2 (RanBP2; also known as Nup358). This protein is found within the nuclear envelope and binds to RanGAP (Hutten, 2008). Following NEB, RanGAP could be anchored to RanBP2-containing cytoplasmic vesicles (Cesario, 2011).
Ran has an unusual localization pattern in oocytes; concentrating around the outside of the spindle. By contrast, Ran overlaps with the spindle in Drosophila mitotic cells. It has not been determined whether these concentrations of Ran are in the GDP or GTP state. However, it the basis of the localization patterns of wild-type and mutant proteins is open to speculation. From this type of evidence, it has been suggested that the bulk of Ran on the embryonic spindle is in the GTP state. Similarly, the bulk of the Ran localized around the outside of the meiotic spindle might be in the GTP form. The pattern of mutant RanQ69L staining suggests it enters RanGAP-containing vesicles but does not leave because it is not hydrolyzed. Thus, the wild-type Ran that localizes adjacent to the clusters of RanGAP could be the GDP form of the protein that has left RanGAP-containing vesicles (Cesario, 2011).
RCC1 and RanGTP were found to be required for chromatin-induced spindle assembly in Xenopus extracts. In such extracts, expression of RanT24N blocks spindle assembly and high concentrations of RCC1 or expression of a GTP-locked form of Ran leads to spindle formation in the absence of chromosomes and centrosomes (Cesario, 2011).
This analysis of RanGTP function in Drosophila oocytes is based on these and numerous other studies in which expression of the ranT24N mutation effectively reduces the concentration of RanGTP. It is believed that the ranT24N mutant had the desired effect of reducing RanGTP production, for four reasons. First, expression of RanT24N in somatic cells caused embryonic or early larval lethality. Second, RanT24N localized tightly to the meiotic chromosomes, consistent with the expectation that this form of Ran remains bound to RCC1 because it has a low rate of GTP exchange. The high affinity of RanT24N for RCC1 causes a reduction in the production of RanGTP. Third, the spindle organization defects observed in ranT24N oocytes were similar to defects seen in mars1 mutant oocytes, a protein known to be regulated by the Ran pathway. Fourth, ranT24N caused dramatic disruptions in chromosome-independent microtubule assembly assays, such as pronuclear fusion. Because these chromosome-dependent (meiotic spindle) and -independent functions occur in the same cytoplasm, it is concluded that the reduction in RanGTP levels sufficient to block pronuclear fusion were not sufficient to block acentrosomal spindle assembly (Cesario, 2011).
Unlike the results in Xenopus extracts, expression of the dominant-negative GDP-locked variant of Ran had relatively mild effects on Drosophila oocyte spindle assembly and karyosome organization. Meiosis I spindles were bipolar in ranT24N oocytes. Indeed, reducing the RanGTP concentration in the oocyte was not sufficient to severely affect either meiotic division, because meiosis II spindles and female meiotic products could be seen in the embryos. The most important defect was that the meiosis I spindle often had non-tapered poles, and was associated with the abnormal localization of proteins necessary for pole formation such as Mars or TACC (Cullen, 2001). These results are consistent with experiments in embryos that found depletion of RanGTP causes defects in spindle pole organization and chromosome organization and congression (Silverman-Gavrila, 2006). Abnormal spindle morphology in oocytes could be the reason for the disorganized karyosome phenotype and nondisjunction of achiasmate chromosomes. Loss of RanGTP could result in a failure to activate Aurora A, which phosphorylates TACC. In embryos, TACC localization to the centrosomes depends on phosphorylation by Aurora A (Barros, 2005). TACC initially binds all microtubules, but as the spindle matures, TACC is phosphorylated and localizes to the poles. Expression of ranT24N in oocytes might cause a reduction in Aurora A activity, resulting in a failure to phosphorylate TACC and localize it to the poles. Further studies are needed, however, because the role of Aurora A in Drosophila female meiosis is not known. In addition, Mars might have a role in promoting the dephosphorylation of TACC (Tan, 2008). Overall, RanGTP might have a specific role in organizing spindle poles but might not be required for chromosome-promoted spindle assembly in oocytes (Cesario, 2011).
Expression of ranT24N did not block spindle assembly. Conversely, expression of the GTP-locked mutant, ranQ69L, did not induce an uncoupling between spindle assembly and the chromosomes, as it does in Xenopus oocytes. Thus, RanGTP might not be sufficient to initiate spindle assembly in Drosophila oocytes. Surprisingly, ranQ69L oocytes showed loss-of-function spindle phenotypes similar to ranT24N mutant oocytes. Interestingly, manipulation of RanGTP levels with T24N or Q69L mutations in mammals has similar phenotypes. For example, the expression of either form of Ran in mouse oocytes resulted in similar meiosis II spindle phenotypes. These results suggest that the effects of manipulating RanGTP levels in an intact oocyte are not easily predicted by experiments in Xenopus extracts. Other factors such as protein localization might play important roles in regulating the Ran pathway. Also it cannot be ruled out that the Ran pathway functions differently in oocyte meiosis, such as if active Ran is not GTP dependent (Cesario, 2011).
It is suggested that there could be two reasons for the similarity of the ranQ69L and ranT24N phenotypes. First, expressing the GTP-locked ranQ69L mutation can inhibit the binding of RCC1 to the chromatin, causing a reduction in RanGTP near the chromatin. Alternatively, the phenotypes of the ranQ69L mutant oocyte might be associated with defects in the organization of membranes or vesicles. For example, expression of the ranQ69L mutation caused Lamin, RanGAP and RanQ69L to colocalize in globular structures throughout the oocyte. The transmembrane protein Axs has been proposed to be a component of a membranous structure surrounding the meiotic spindle. With the caveat that the link between membranous structures and spindle assembly is not known, the ranQ69L mutation might cause defects in membranous structures that have a role in spindle organization. It was also found that Ran and Axs are closely associated, although at the light microscope level it is difficult to determine if Ran is inside or outside the Axs staining (Cesario, 2011).
Similar to the oocytes, the phenotype of the ranQ69L mutant zygotes might be associated with defects in membrane structure. In ranQ69L mutants, a single cluster of DNA and microtubules could be observed in the center of the zygote. A strikingly similar phenotype has been observed in dominant-negative Ketel mutants; Ketel is the Drosophila homolog of importin-β. In the Ketel dominant mutants, meiosis I and II occur and the female and male pronuclei come together, but they interact abnormally because of defects in the nuclear envelopes. Subsequently, the first mitotic division fails and the chromosomes disintegrate within a large aggregate of microtubules. Similar to the Ketel mutant, ranQ69L could cause abnormal interactions among nuclear envelope proteins in the zygote, causing a failure in the first mitotic division (Cesario, 2011).
There are two chromosome segregation mechanisms in Drosophila females. The first is the segregation of bivalents connected by chiasmata, which is how most chromosomes segregate. The second is of the chromosomes that lack chiasmata. This includes the small fourth chromosome, which always lacks crossovers, and larger chromosomes, which lack a crossover in approximately 5% of meioses. Homologous pairs can be forced into the achiasmate system with balancers that suppress crossing over. In all these cases, homologous chromosomes segregate correctly even though they are not connected by chiasmata. Expression of ranT24N had only mild effects on chiasmate segregation, but had a severe effect on the segregation of achiasmate X-chromosomes. These results suggest that the spindle pole organization defects caused by low RanGTP levels affect chromosome segregation (Cesario, 2011).
Unlike assembly of the meiosis I spindle, expression of ranT24N blocked two other types of microtubule assembly. First, ranT24N mutants had a defect in the fusion of the female and male pronuclei. This was the most probable cause of the fertility defect in ranT24N mutants. Several genes with roles in microtubule assembly are also required for pronuclear fusion, including subito. This process depends on the assembly of a microtubule array that is nucleated by the centrosome donated by the sperm, and acts to draw the female pronucleus towards the male pronucleus. Second, ranT24N suppressed the formation of the ectopic spindles that form in a neomorphic subito mutant (sub?NT) . The formation of these spindles occurs after NEB, consistent with a dependence on release of RanGTP from the nucleus. Both of these examples involve assembly of microtubules without direct interaction with the chromosomes and suggest that the assembly and bundling of microtubules in the oocyte cytoplasm depend on RanGTP (Cesario, 2011).
One characteristic of the ectopic spindles in subδNT mutants is that they form in discrete clusters within the oocyte. Because RanGAP appears in clusters, it is possible that RanGTP is not in a gradient or distributed evenly in the cytoplasm. Thus, an interesting possibility is that the regions containing ectopic spindles are where the concentration of RanGTP-dependent spindle assembly factors are at their highest (Cesario, 2011).
These experiments revealed a surprising mutual suppression by the sub?NT and ranT24N mutations. Although both mutants have decreased fertility, the double mutant is fertile. One interpretation is that RanGTP regulates Subito, and the subδNT mutation bypasses the dependence on RanGTP. However, this study found that ran mutations did not affect Subito localization or the formation of the central spindle. A more probable explanation is that there are two independent spindle assembly pathways in the oocyte and the loss of spindle assembly factors in ranT24N zygotes is balanced by the enhanced spindle assembly activity present in the sub?NT mutant. Expression of ranT24N might suppress the sterility phenotype of sub?NT by abolishing ectopic spindles, whereas sub?NT might suppress the reduced fertility phenotype of ranT24N by overcoming the defects in microtubule assembly needed for processes such as pro-nuclear fusion (Cesario, 2011).
Using dominant-negative mutations, this study has found that RanGTP is required for the fertility of Drosophila females. No evidence was found that RanGTP is required, or sufficient, for the initiation of acentrosomal spindle assembly in Drosophila oocytes. A role was detected in organizing the spindle poles that could be explained by RanGTP regulation of proteins such as Mars/Hurp, TACC and Aurora A. These defects, however, would not be expected to have a severe effect on fertility. A similar conclusion was drawn from expressing a dominant-negative form of Ran in mouse oocytes or when RCC1 was depleted from Xenopus oocytes. By analyzing mutants similar to the ones used in this study, only mild defects in meiosis I spindle assembly were found, such as a delay establishing bipolarity. The failure to observe evidence supporting a role for RanGTP in acentrosomal spindle assembly might be explained by a predominant chromosome-dependent pathway in oocytes involving the chromosome passenger complex (CPC). The CPC is required for chromosome-dependent spindle assembly in Xenopus egg extracts (Cesario, 2011).
These results suggest that, compared with chromosome-mediated spindle assembly, RanGTP has a greater role in microtubule organization when centrosomes are present or when at a distance from the chromosomes. It is suggested that NEB preceding the assembly of the meiosis I spindle releases RanGTP into the cytoplasm, which results in a cytoplasm enriched for active spindle assembly factors. The restriction of RanGAP to vesicle-like structures could leave a considerable amount of RanGTP in the cytoplasm. This activity has only a minor role in meiotic spindle assembly, but is crucial for the early events of embryogenesis. The microtubule array facilitating pronuclear fusion assembles while the nuclear envelope is intact. Therefore, the oocyte might accumulate and store RanGTP during the meiotic divisions when there is no nuclear envelope in order to support pronuclear fusion, when the nuclear envelope is intact (Cesario, 2011).
Reference names in red indicate recommended papers.
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date revised: 20 March 2012
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