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Mutational analysis of Mdm1p function in nuclear and mitochondrial inheritance.

Fisk HA, Yaffe MP - J. Cell Biol. (1997)

Bottom Line: Class I and II mutants also exhibited altered mitochondrial morphology, possessing primarily small, round mitochondria instead of the extended tubular structures found in wild-type cells.Genetic crosses of yeast strains containing different mdm1 alleles revealed complex genetic interactions including intragenic suppression, synthetic phenotypes, and intragenic complementation.These results support a model of Mdm1p function in which a network comprised of multimeric assemblies of the protein mediates two distinct cellular processes.

View Article: PubMed Central - PubMed

Affiliation: Department of Biology, University of California, San Diego, La Jolla, California 92093, USA.

ABSTRACT
Nuclear and mitochondrial transmission to daughter buds of Saccharomyces cerevisiae depends on Mdm1p, an intermediate filament-like protein localized to numerous punctate structures distributed throughout the yeast cell cytoplasm. These structures disappear and organelle inheritance is disrupted when mdm1 mutant cells are incubated at the restrictive temperature. To characterize further the function of Mdm1p, new mutant mdm1 alleles that confer temperature-sensitive growth and defects in organelle inheritance but produce stable Mdm1p structures were isolated. Microscopic analysis of the new mdm1 mutants revealed three phenotypic classes: Class I mutants showed defects in both mitochondrial and nuclear transmission; Class II alleles displayed defective mitochondrial inheritance but had no effect on nuclear movement; and Class III mutants showed aberrant nuclear inheritance but normal mitochondrial distribution. Class I and II mutants also exhibited altered mitochondrial morphology, possessing primarily small, round mitochondria instead of the extended tubular structures found in wild-type cells. Mutant mdm1 alleles affecting nuclear transmission were of two types: Class Ia and IIIa mutants were deficient for nuclear movement into daughter buds, while Class Ib and IIIb mutants displayed a complete transfer of all nuclear DNA into buds. The mutations defining all three allelic classes mapped to two distinct domains within the Mdm1p protein. Genetic crosses of yeast strains containing different mdm1 alleles revealed complex genetic interactions including intragenic suppression, synthetic phenotypes, and intragenic complementation. These results support a model of Mdm1p function in which a network comprised of multimeric assemblies of the protein mediates two distinct cellular processes.

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Intragenic interaction among mdm1 alleles. Genetic crosses were performed to obtain all possible pairwise combinations of  different mdm1 alleles. Organelle inheritance phenotypes were analyzed as described in Fig. 1. (A) Phenotypic classes of mdm1 diploids.  Numbers on the x and y axis indicate mdm1 allele of parental strains. Roman numeral refers to phenotypic class, and lower case “a” or  “b” refers to nuclear transmission phenotype, as described in Table II and in the text. “R” or “D” indicates the recessive or dominant  character of individual alleles. (B) Intragenic complementation is observed between mdm1-217 (Class IIIa) and mdm1-228 (Class IIIb).  Haploids and diploids harboring the mdm1-217 and mdm1-228 mutations were tested for growth on YPD at 37°C. (a–c) mdm1-217; (a)  MATa, (b) MATα, and (c) MATa/α; (d–f) mdm1-228; (d) MATa, (e) MATα, and (f) MATa/α; (g) mdm1-217/mdm1-228 (a/α); (h) mdm1-228/mdm1-217.
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Figure 5: Intragenic interaction among mdm1 alleles. Genetic crosses were performed to obtain all possible pairwise combinations of different mdm1 alleles. Organelle inheritance phenotypes were analyzed as described in Fig. 1. (A) Phenotypic classes of mdm1 diploids. Numbers on the x and y axis indicate mdm1 allele of parental strains. Roman numeral refers to phenotypic class, and lower case “a” or “b” refers to nuclear transmission phenotype, as described in Table II and in the text. “R” or “D” indicates the recessive or dominant character of individual alleles. (B) Intragenic complementation is observed between mdm1-217 (Class IIIa) and mdm1-228 (Class IIIb). Haploids and diploids harboring the mdm1-217 and mdm1-228 mutations were tested for growth on YPD at 37°C. (a–c) mdm1-217; (a) MATa, (b) MATα, and (c) MATa/α; (d–f) mdm1-228; (d) MATa, (e) MATα, and (f) MATa/α; (g) mdm1-217/mdm1-228 (a/α); (h) mdm1-228/mdm1-217.

Mentions: To examine in greater detail the interactions between different mdm1 mutant alleles, all possible diploid combinations of the 10 new alleles were generated, and phenotypes of the resulting diploids were analyzed. Cells were examined for growth at 37°C and by fluorescence microscopy for mitochondrial and nuclear distribution. The consequent mutant phenotypes revealed by this analysis indicated a complex pattern of interactions (Fig. 5 A). First, certain combinations of dominant alleles yielded an apparently additive (and expected) phenotype. For example, the combination of either dominant Class II mutation with the dominant Class IIIb mutation, mdm1-227, produced diploids displaying a Class Ib phenotype. Second, some combinations of alleles yielded unexpected synthetic phenotypes. For example, the diploid obtained by crossing the recessive Class Ia mutant, mdm1-199, with the dominant Class II mutant, mdm1-202, displayed a Class Ib phenotype. Third, certain crosses led to intragenic suppression in which a phenotype caused by a dominant mutation was not observed in the diploid. An example of such intragenic suppression was the cross of the dominant Class II mutant, mdm1-202, with the dominant Class IIIa mutant, mdm1-200, resulting in a Class IIIb diploid that displayed no defect in mitochondrial distribution (suppression of the mitochondrial defect). A final type of interaction was intragenic complementation, in which the combination of two different alleles eliminated mutant phenotypes observed in one or both of the parental strains. An example of such complementation emerged from the mating of recessive Class IIIa mutant, mdm1-217, with recessive Class IIIb mutant, mdm1-228, to yield a diploid strain with wild-type growth and normal organelle distribution (Fig. 5 B).


Mutational analysis of Mdm1p function in nuclear and mitochondrial inheritance.

Fisk HA, Yaffe MP - J. Cell Biol. (1997)

Intragenic interaction among mdm1 alleles. Genetic crosses were performed to obtain all possible pairwise combinations of  different mdm1 alleles. Organelle inheritance phenotypes were analyzed as described in Fig. 1. (A) Phenotypic classes of mdm1 diploids.  Numbers on the x and y axis indicate mdm1 allele of parental strains. Roman numeral refers to phenotypic class, and lower case “a” or  “b” refers to nuclear transmission phenotype, as described in Table II and in the text. “R” or “D” indicates the recessive or dominant  character of individual alleles. (B) Intragenic complementation is observed between mdm1-217 (Class IIIa) and mdm1-228 (Class IIIb).  Haploids and diploids harboring the mdm1-217 and mdm1-228 mutations were tested for growth on YPD at 37°C. (a–c) mdm1-217; (a)  MATa, (b) MATα, and (c) MATa/α; (d–f) mdm1-228; (d) MATa, (e) MATα, and (f) MATa/α; (g) mdm1-217/mdm1-228 (a/α); (h) mdm1-228/mdm1-217.
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Figure 5: Intragenic interaction among mdm1 alleles. Genetic crosses were performed to obtain all possible pairwise combinations of different mdm1 alleles. Organelle inheritance phenotypes were analyzed as described in Fig. 1. (A) Phenotypic classes of mdm1 diploids. Numbers on the x and y axis indicate mdm1 allele of parental strains. Roman numeral refers to phenotypic class, and lower case “a” or “b” refers to nuclear transmission phenotype, as described in Table II and in the text. “R” or “D” indicates the recessive or dominant character of individual alleles. (B) Intragenic complementation is observed between mdm1-217 (Class IIIa) and mdm1-228 (Class IIIb). Haploids and diploids harboring the mdm1-217 and mdm1-228 mutations were tested for growth on YPD at 37°C. (a–c) mdm1-217; (a) MATa, (b) MATα, and (c) MATa/α; (d–f) mdm1-228; (d) MATa, (e) MATα, and (f) MATa/α; (g) mdm1-217/mdm1-228 (a/α); (h) mdm1-228/mdm1-217.
Mentions: To examine in greater detail the interactions between different mdm1 mutant alleles, all possible diploid combinations of the 10 new alleles were generated, and phenotypes of the resulting diploids were analyzed. Cells were examined for growth at 37°C and by fluorescence microscopy for mitochondrial and nuclear distribution. The consequent mutant phenotypes revealed by this analysis indicated a complex pattern of interactions (Fig. 5 A). First, certain combinations of dominant alleles yielded an apparently additive (and expected) phenotype. For example, the combination of either dominant Class II mutation with the dominant Class IIIb mutation, mdm1-227, produced diploids displaying a Class Ib phenotype. Second, some combinations of alleles yielded unexpected synthetic phenotypes. For example, the diploid obtained by crossing the recessive Class Ia mutant, mdm1-199, with the dominant Class II mutant, mdm1-202, displayed a Class Ib phenotype. Third, certain crosses led to intragenic suppression in which a phenotype caused by a dominant mutation was not observed in the diploid. An example of such intragenic suppression was the cross of the dominant Class II mutant, mdm1-202, with the dominant Class IIIa mutant, mdm1-200, resulting in a Class IIIb diploid that displayed no defect in mitochondrial distribution (suppression of the mitochondrial defect). A final type of interaction was intragenic complementation, in which the combination of two different alleles eliminated mutant phenotypes observed in one or both of the parental strains. An example of such complementation emerged from the mating of recessive Class IIIa mutant, mdm1-217, with recessive Class IIIb mutant, mdm1-228, to yield a diploid strain with wild-type growth and normal organelle distribution (Fig. 5 B).

Bottom Line: Class I and II mutants also exhibited altered mitochondrial morphology, possessing primarily small, round mitochondria instead of the extended tubular structures found in wild-type cells.Genetic crosses of yeast strains containing different mdm1 alleles revealed complex genetic interactions including intragenic suppression, synthetic phenotypes, and intragenic complementation.These results support a model of Mdm1p function in which a network comprised of multimeric assemblies of the protein mediates two distinct cellular processes.

View Article: PubMed Central - PubMed

Affiliation: Department of Biology, University of California, San Diego, La Jolla, California 92093, USA.

ABSTRACT
Nuclear and mitochondrial transmission to daughter buds of Saccharomyces cerevisiae depends on Mdm1p, an intermediate filament-like protein localized to numerous punctate structures distributed throughout the yeast cell cytoplasm. These structures disappear and organelle inheritance is disrupted when mdm1 mutant cells are incubated at the restrictive temperature. To characterize further the function of Mdm1p, new mutant mdm1 alleles that confer temperature-sensitive growth and defects in organelle inheritance but produce stable Mdm1p structures were isolated. Microscopic analysis of the new mdm1 mutants revealed three phenotypic classes: Class I mutants showed defects in both mitochondrial and nuclear transmission; Class II alleles displayed defective mitochondrial inheritance but had no effect on nuclear movement; and Class III mutants showed aberrant nuclear inheritance but normal mitochondrial distribution. Class I and II mutants also exhibited altered mitochondrial morphology, possessing primarily small, round mitochondria instead of the extended tubular structures found in wild-type cells. Mutant mdm1 alleles affecting nuclear transmission were of two types: Class Ia and IIIa mutants were deficient for nuclear movement into daughter buds, while Class Ib and IIIb mutants displayed a complete transfer of all nuclear DNA into buds. The mutations defining all three allelic classes mapped to two distinct domains within the Mdm1p protein. Genetic crosses of yeast strains containing different mdm1 alleles revealed complex genetic interactions including intragenic suppression, synthetic phenotypes, and intragenic complementation. These results support a model of Mdm1p function in which a network comprised of multimeric assemblies of the protein mediates two distinct cellular processes.

Show MeSH
Related in: MedlinePlus