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The inhibition of polo kinase by matrimony maintains G2 arrest in the meiotic cell cycle.

Xiang Y, Takeo S, Florens L, Hughes SE, Huo LJ, Gilliland WD, Swanson SK, Teeter K, Schwartz JW, Washburn MP, Jaspersen SL, Hawley RS - PLoS Biol. (2007)

Bottom Line: Loss-of-function mtrm mutants result in precocious NEB.The meiotic defects observed in mtrm/+ heterozygotes are fully suppressed by reducing the dose of polo+, demonstrating that Mtrm acts as an inhibitor of Polo.Our data suggest a model in which the eventual activation of Cdc25 by an excess of Polo at stage 13 triggers NEB and entry into prometaphase.

View Article: PubMed Central - PubMed

Affiliation: Stowers Institute for Medical Research, Kansas City, Missouri, United States of America.

ABSTRACT
Many meiotic systems in female animals include a lengthy arrest in G2 that separates the end of pachytene from nuclear envelope breakdown (NEB). However, the mechanisms by which a meiotic cell can arrest for long periods of time (decades in human females) have remained a mystery. The Drosophila Matrimony (Mtrm) protein is expressed from the end of pachytene until the completion of meiosis I. Loss-of-function mtrm mutants result in precocious NEB. Coimmunoprecipitation experiments reveal that Mtrm physically interacts with Polo kinase (Polo) in vivo, and multidimensional protein identification technology mass spectrometry analysis reveals that Mtrm binds to Polo with an approximate stoichiometry of 1:1. Mutation of a Polo-Box Domain (PBD) binding site in Mtrm ablates the function of Mtrm and the physical interaction of Mtrm with Polo. The meiotic defects observed in mtrm/+ heterozygotes are fully suppressed by reducing the dose of polo+, demonstrating that Mtrm acts as an inhibitor of Polo. Mtrm acts as a negative regulator of Polo during the later stages of G2 arrest. Indeed, both the repression of Polo expression until stage 11 and the inactivation of newly synthesized Polo by Mtrm until stage 13 play critical roles in maintaining and properly terminating G2 arrest. Our data suggest a model in which the eventual activation of Cdc25 by an excess of Polo at stage 13 triggers NEB and entry into prometaphase.

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Reducing the Dose of polo+ Suppresses mtrm Defects, and Increasing the Dose of polo+ Partially Mimics the Effects of mtrm(A) Schematic diagram of the polo gene (black boxes depict the five exons) indicating the insertions sites for the two polo alleles (polo16–1 and poloKG03033).(B) Summary of the genetic interaction of mtrm and polo mutants as examined by assaying the frequency of nondisjunction of X and 4th chromosomes. As shown by Harris et al. [9], mtrm/+ heterozygotes display high levels of nondisjunction for both achiasmate X and 4th chromosomes (42% and 37%, respectively) when compared to mtrm+/mtrm+ females. However, simultaneously reducing the dose of polo, as a result of heterozygosity for either the two P element insertion site mutants or a deficiency that uncovers polo (Df(3L)rdgC-co2) suppresses the meiotic phenotype of mtrm/+ heterozygotes.(C) Expression of the UASP-polo+ transgene in mtrm+/mtrm+ females results in a dose-dependent increase in the frequency of achiasmate nondisjunction for both the X and the 4th chromosomes. However, two weaker alleles of polo, polo01673 and polo1, showed little or no suppression of the segregational defect (unpublished data). The polo1 mutant, which is the weakest of the known polo mutants (it is viable over a deletion) is the result of a point mutation at base pair 725, V242E, in the kinase domain. Although polo01673 is recessive lethal, it must retain some degree of function because it complements at least one other hypomorphic allele of polo, polox8. The results indicate that reduction of polo+ dosage rescues mtrm defects and the suppressive effect of a given polo mutant correlates with the severity in the reduction of Polo function.
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pbio-0050323-g003: Reducing the Dose of polo+ Suppresses mtrm Defects, and Increasing the Dose of polo+ Partially Mimics the Effects of mtrm(A) Schematic diagram of the polo gene (black boxes depict the five exons) indicating the insertions sites for the two polo alleles (polo16–1 and poloKG03033).(B) Summary of the genetic interaction of mtrm and polo mutants as examined by assaying the frequency of nondisjunction of X and 4th chromosomes. As shown by Harris et al. [9], mtrm/+ heterozygotes display high levels of nondisjunction for both achiasmate X and 4th chromosomes (42% and 37%, respectively) when compared to mtrm+/mtrm+ females. However, simultaneously reducing the dose of polo, as a result of heterozygosity for either the two P element insertion site mutants or a deficiency that uncovers polo (Df(3L)rdgC-co2) suppresses the meiotic phenotype of mtrm/+ heterozygotes.(C) Expression of the UASP-polo+ transgene in mtrm+/mtrm+ females results in a dose-dependent increase in the frequency of achiasmate nondisjunction for both the X and the 4th chromosomes. However, two weaker alleles of polo, polo01673 and polo1, showed little or no suppression of the segregational defect (unpublished data). The polo1 mutant, which is the weakest of the known polo mutants (it is viable over a deletion) is the result of a point mutation at base pair 725, V242E, in the kinase domain. Although polo01673 is recessive lethal, it must retain some degree of function because it complements at least one other hypomorphic allele of polo, polox8. The results indicate that reduction of polo+ dosage rescues mtrm defects and the suppressive effect of a given polo mutant correlates with the severity in the reduction of Polo function.

Mentions: mtrm/+ heterozygotes display a substantial defect in the processes that ensure the segregation of achiasmate homologs. We show here that these meiotic defects are strongly suppressed by simultaneous heterozygosity for strong loss-of-function alleles of polo. (Our impetus for searching for a genetic interaction between mtrm and polo came from the finding that the mutants in the mei-S332 gene were partially suppressed by polo mutants [12].) We measure meiotic mis-segregation by assaying X and 4th chromosomal nondisjunction in females of the genotype FM7/X where FM7 is a balancer chromosome that fully suppresses X chromosomal exchange. (The 4th chromosome is obligately achiasmate.) As shown in Figure 3B, FM7/X; mtrm/+ females typically show frequencies of X and 4th chromosome nondisjunction in the range of 35%–45%, which is more than 100-fold above control values.


The inhibition of polo kinase by matrimony maintains G2 arrest in the meiotic cell cycle.

Xiang Y, Takeo S, Florens L, Hughes SE, Huo LJ, Gilliland WD, Swanson SK, Teeter K, Schwartz JW, Washburn MP, Jaspersen SL, Hawley RS - PLoS Biol. (2007)

Reducing the Dose of polo+ Suppresses mtrm Defects, and Increasing the Dose of polo+ Partially Mimics the Effects of mtrm(A) Schematic diagram of the polo gene (black boxes depict the five exons) indicating the insertions sites for the two polo alleles (polo16–1 and poloKG03033).(B) Summary of the genetic interaction of mtrm and polo mutants as examined by assaying the frequency of nondisjunction of X and 4th chromosomes. As shown by Harris et al. [9], mtrm/+ heterozygotes display high levels of nondisjunction for both achiasmate X and 4th chromosomes (42% and 37%, respectively) when compared to mtrm+/mtrm+ females. However, simultaneously reducing the dose of polo, as a result of heterozygosity for either the two P element insertion site mutants or a deficiency that uncovers polo (Df(3L)rdgC-co2) suppresses the meiotic phenotype of mtrm/+ heterozygotes.(C) Expression of the UASP-polo+ transgene in mtrm+/mtrm+ females results in a dose-dependent increase in the frequency of achiasmate nondisjunction for both the X and the 4th chromosomes. However, two weaker alleles of polo, polo01673 and polo1, showed little or no suppression of the segregational defect (unpublished data). The polo1 mutant, which is the weakest of the known polo mutants (it is viable over a deletion) is the result of a point mutation at base pair 725, V242E, in the kinase domain. Although polo01673 is recessive lethal, it must retain some degree of function because it complements at least one other hypomorphic allele of polo, polox8. The results indicate that reduction of polo+ dosage rescues mtrm defects and the suppressive effect of a given polo mutant correlates with the severity in the reduction of Polo function.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2100146&req=5

pbio-0050323-g003: Reducing the Dose of polo+ Suppresses mtrm Defects, and Increasing the Dose of polo+ Partially Mimics the Effects of mtrm(A) Schematic diagram of the polo gene (black boxes depict the five exons) indicating the insertions sites for the two polo alleles (polo16–1 and poloKG03033).(B) Summary of the genetic interaction of mtrm and polo mutants as examined by assaying the frequency of nondisjunction of X and 4th chromosomes. As shown by Harris et al. [9], mtrm/+ heterozygotes display high levels of nondisjunction for both achiasmate X and 4th chromosomes (42% and 37%, respectively) when compared to mtrm+/mtrm+ females. However, simultaneously reducing the dose of polo, as a result of heterozygosity for either the two P element insertion site mutants or a deficiency that uncovers polo (Df(3L)rdgC-co2) suppresses the meiotic phenotype of mtrm/+ heterozygotes.(C) Expression of the UASP-polo+ transgene in mtrm+/mtrm+ females results in a dose-dependent increase in the frequency of achiasmate nondisjunction for both the X and the 4th chromosomes. However, two weaker alleles of polo, polo01673 and polo1, showed little or no suppression of the segregational defect (unpublished data). The polo1 mutant, which is the weakest of the known polo mutants (it is viable over a deletion) is the result of a point mutation at base pair 725, V242E, in the kinase domain. Although polo01673 is recessive lethal, it must retain some degree of function because it complements at least one other hypomorphic allele of polo, polox8. The results indicate that reduction of polo+ dosage rescues mtrm defects and the suppressive effect of a given polo mutant correlates with the severity in the reduction of Polo function.
Mentions: mtrm/+ heterozygotes display a substantial defect in the processes that ensure the segregation of achiasmate homologs. We show here that these meiotic defects are strongly suppressed by simultaneous heterozygosity for strong loss-of-function alleles of polo. (Our impetus for searching for a genetic interaction between mtrm and polo came from the finding that the mutants in the mei-S332 gene were partially suppressed by polo mutants [12].) We measure meiotic mis-segregation by assaying X and 4th chromosomal nondisjunction in females of the genotype FM7/X where FM7 is a balancer chromosome that fully suppresses X chromosomal exchange. (The 4th chromosome is obligately achiasmate.) As shown in Figure 3B, FM7/X; mtrm/+ females typically show frequencies of X and 4th chromosome nondisjunction in the range of 35%–45%, which is more than 100-fold above control values.

Bottom Line: Loss-of-function mtrm mutants result in precocious NEB.The meiotic defects observed in mtrm/+ heterozygotes are fully suppressed by reducing the dose of polo+, demonstrating that Mtrm acts as an inhibitor of Polo.Our data suggest a model in which the eventual activation of Cdc25 by an excess of Polo at stage 13 triggers NEB and entry into prometaphase.

View Article: PubMed Central - PubMed

Affiliation: Stowers Institute for Medical Research, Kansas City, Missouri, United States of America.

ABSTRACT
Many meiotic systems in female animals include a lengthy arrest in G2 that separates the end of pachytene from nuclear envelope breakdown (NEB). However, the mechanisms by which a meiotic cell can arrest for long periods of time (decades in human females) have remained a mystery. The Drosophila Matrimony (Mtrm) protein is expressed from the end of pachytene until the completion of meiosis I. Loss-of-function mtrm mutants result in precocious NEB. Coimmunoprecipitation experiments reveal that Mtrm physically interacts with Polo kinase (Polo) in vivo, and multidimensional protein identification technology mass spectrometry analysis reveals that Mtrm binds to Polo with an approximate stoichiometry of 1:1. Mutation of a Polo-Box Domain (PBD) binding site in Mtrm ablates the function of Mtrm and the physical interaction of Mtrm with Polo. The meiotic defects observed in mtrm/+ heterozygotes are fully suppressed by reducing the dose of polo+, demonstrating that Mtrm acts as an inhibitor of Polo. Mtrm acts as a negative regulator of Polo during the later stages of G2 arrest. Indeed, both the repression of Polo expression until stage 11 and the inactivation of newly synthesized Polo by Mtrm until stage 13 play critical roles in maintaining and properly terminating G2 arrest. Our data suggest a model in which the eventual activation of Cdc25 by an excess of Polo at stage 13 triggers NEB and entry into prometaphase.

Show MeSH
Related in: MedlinePlus