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Extrachromosomal inheritance in Schizosaccharomyces pombe. IV. Isolation and genetic characterization of mutants resistant to chloramphenicol and erythromycin using the mutator properties of mutant anar-8.

Spontaneous chloramphenicol (capr)- and erythromycin (eryr)-resistant mutants were isolated from strain ade7-50 h- and the antimycin-resistant mutant anar-8 ade 7-50 h- of Schizosaccharomyces pombe (Sch. p.). By mitotic segregation analysis all 154 capr- and 120 eryr-mutants derived from ade 7-50 h- proved to be recessive chromosomal, whereas all 108 capr- and 200 eryr-mutants originating from anar-8 were extrachromosomally inherited. The rate of spontaneous capr- and eryr-mutants was about hundredfold in anar-8 compared to ade 7-50 h-. Growth of capr- and eryr-mutants was not inhibited by chloramphenicol or erythromycin, respectively, in glucose-medium and only slightly in glycerol-medium at concentrations which completely inhibited anar-8. By mitotic segregation-, tetrad-, and mitotic haploidization-analysis the extrachromosomal inheritance of mutants derived from anar-8 was established. Segregational patterns of capr- and eryr-determinants during mitosis, meiosis, and mitotic haploidization of diploids are discussed.

Ascomycota

Chromosomal and extrachromosomal inheritance of erythromycin-resistance in the "petite-negative" yeast Kluyveromyces lactis.

Spontaneously arising erythromycin-resistant mutants were isolated in the "petite-negative" yeast Kluyveromyces lactis. Two independently arising mutants were studied, in one erythromycin-resistance was conferred by a single dominant nuclear gene, and in the other the resistance was extrachromosomally inherited. In fermentable medium growth of sensitive and resistant strains in presence or absence of erythromycin does not qualitatively change the cytochrome absorption spectra, whereas oxygen uptake of parental strains growing in glucose-medium is affected by the drug. The importance of "petite-negative" yeasts like Kluyveromyces lactis for the study of nucleo-cytoplasmic interrelations is discussed.

Chromosomes

Extrachromosomal inheritance of susceptibility to trypanosome infection in tsetse flies. I. Selection of susceptible and refractory lines of Glossina morsitans morsitans.

Differences in susceptibility to infection with Trypanosoma congolense between F1 families of Glossina morsitans morsitans indicated that susceptibility is maternally inherited in this species of tsetse fly. Twelve F1 families, six selected for susceptibility and six selected for refractoriness to infection, have been bred for up to 13 generations. The reciprocal differences demonstrated in the F1 generation persisted in these selected families over many generations, indicating that susceptibility/refractoriness to T. congolense infection is extrachromosomally inherited in G. m. morsitans. Repeated 'backcrossing' to males of the opposite strain showed that infection rates within families were independent of the contribution of the male parent. Susceptible families had a mean midgut infection rate of 76.9% and a mature (hypopharyngeal) infection rate of 47.9%. In the refractory families 88.9% of the flies failed to develop an infection, 11.1% had midgut infections and only 6.3% developed mature infections. Levels of midgut infection remained remarkably constant within families over generations, whether refractory or susceptible, while maturation rates varied between generations and between sexes. Males matured a significantly greater proportion of midgut infections than females in the susceptible families. It is suggested that the inheritance of susceptibility/refractoriness relates primarily to the establishment of midgut infections in G. m. morsitans, and that maturation of midgut infections is dependent upon environmental factors such as diet, and differences between sexes probably reflecting differences in rates of bloodmeal digestion.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals

[The origin of extrachromosomal inheritance factors (plasmids) in bacteria].

Present status of the problem of origin of extra-chromosomal inheritance elements is reviewed in the light of literary data and some results obtained by the author. The role of plasmids in the ecology of bacteria, in their adaptive variability and evolution is discussed. Extra-chromosomal inheritance is regarded as a particular case of biological inheritance.

Bacteria

Extrachromosomal inheritance in Schizosaccharomyces pombe. I. Evidence for an extrakaryotically inherited mutation conferring resistance to antimycin.

In crosses of [ANTr8] with auxotrophic strains, resistance to antimycin segregates almost 50:50 in random spore analysis with a slight preponderance for the sensitivity allele. Tetrad analysis, however, shows all possible types of tetrads (2:2; 3:1; 1:3; 4:0; 0:4 resistant versus sensitive) with an excess of 2:2 segregations and sectoring of colonies on antimycin medium indicating an extrachromosomal mode of inheritance. The overall ratio of resistant versus sensitive spores is the same as compared with random spore data. Using a mutant blocked in meiosis (mei 1) mitotic segregation of stable diploids is achieved, leading to a ratio of 20% resistant to 80% sensitive clones. Possible reasons for the bias in transmission of the resistance determinant is discussed.

Antimycin A

Extrachromosomal inheritance in Schizosaccharomyces pombe. III. Isolation and characterization of paromomycin-resistant mutants.

In the antimycin--resistant mutant anar-8 of the fission yeast Schizosaccharomyces pombe (Sch.p.) spontaneous mutants were isolated showing high resistance to the aminoglycoside antibiotic paromomycin. All mutants were resistant to the structurally related antibiotic neomycin. Tetrad analysis, mitotic segregation analysis, and mitotic haploidization revealed extrachromosomal, very likely mitochondrial inheritance. In contrast to the rapid segregation of mitochondrial markers in zygotic clones of Saccharomyces cerevisiae (S.c.) the heteroplasmic state of diploids proved to persist for at least 50 generations after zygote formation. Stationary cultures of the paromomycin-resistant mutants parr-106 and parr-112 contain up to 6% respiratory-deficient mutants, but no reversion to paromomycin-sensitivity was observed among 1700-1800 colonies tested. The ability of mutant anar-8 to produce spontaneously respiratory-deficient mutants could be separated from the antimycin-resistant phenotype of anar-8.

Ascomycota

Extrachromosomal inheritance in Schizosaccharomyces pombe. II. Evidence for extrakaryotically inherited respiratory deficient mutants.

In contrast to the wild-type, mutant [ANTr8] is able spontaneously to throw off stable respiratory deficient mutants. The frequency of these mutants is considerably enhanced by treatment with ethidium bromide (EB) or the azo-dye Janus green (JG). An unstable cell state with a petite-like phenotype is found in both mutant [ANTr8] and wild-type after EB-treatment. However, only in the mutant is this unstable cell state followed by the appearance of stable respiratory deficient (RD) mutants. Formation of microcolonies is observed both in [ANTr8] and wild-type. RD mutants were isolated after EB treatment. Three of them (mit-12, mit-25, and mit-30) were analyzed and mit-25 characterized in more detail.

Ascomycota

Extrachromosomal inheritance of susceptibility to trypanosome infection in tsetse flies. II. Susceptibility of selected lines of Glossina morsitans morsitans to different stocks and species of trypanosome.

Two lines of Glossina m. morsitans, selected for susceptibility and refractoriness to infection with a single stock of Trypanosoma congolense, have now been shown to be susceptible or refractory to different stocks of T. congolense and, also, to different stocks of T. b. brucei and T. b. gambiense. The mean midgut infection rates of the susceptible line obtained in different experiments with T. congolense, T. b. brucei and T. b. gambiense were, respectively, 66%, 56% and 55%; the corresponding mature (hypopharynx or salivary gland) infection rates were 37%, 23% and 0%. The highest mature infection rates obtained in individual experiments with susceptible flies were 65% (T. congolense) and 40% (T. b. brucei). Mean T. congolense and T. b. brucei midgut infection rates obtained with the refractory line were 29% and 33% respectively, the mature infection rates being 12% and 7%, all significantly lower than the corresponding rates in the susceptible line. Development of midgut infections in susceptible flies appears to take place irrespective of trypanosome stock or form. There is some evidence to suggest that higher infection rates can be obtained with flies infected and maintained on mammals rather than on in vitro feeding systems. Susceptible flies matured a significantly greater proportion of their midgut T. congolense and T. b. brucei infections than did the refractory line, which suggests that maturation of infections is influenced by the susceptibility status of the fly. However, the apparent inability of these flies to develop mature infections of a major T. b. gambiense genetic grouping suggests that maturation of infections established in the midgut is a phenomenon primarily associated with trypanosome genotype.

Animals

Extrachromosomal inheritance in Schizosaccharomyces pombe. VII. Studies by zygote clone analysis on transmission, segregation, recombination, and uniparental inheritance of mitochondrial markers conferring resistance to antimycin, chloramphenicol, and erythromycin.

Crosses involving mitochondrial markers conferring resistance to antimycin (anar, AR), chloramphenicol (capr, CR), and erythromycin (eryr, ER) in cis- and trans-configuration were studied by zygote clone analysis. Mutant anar-8, from which all other drug--resistant isolates were derived, exhibits a highly biased transmission (6.8% anar) in an analysis of 100 individual zygote clones. Important results of zygote clone analyses were:--Zygote clones may contain one, two, three, or four mitochondrial genotypes.--The proportion of the two parental and the two recombinant genotypes in individual zygote clones can vary almost over the entire range of percentages.--Proportions of the two corresponding recombinant types in individual clones are usually unequal.--Transmission rates of markers are higher in trans- than in cis-crosses, indicating additivity of bias by two mutated alleles in coupling.--Transmission rates are different for the three markers both in cis- and trans-crosses, being lowest for CR and highest for ER.--Up to more than 80% uniform clones, expressing only one genotype, can be produced in cis- and trans-crosses. In cis-crosses always the double-sensitive parental type becomes uniform, in trans-crosses this may be the case for parental and/or recombinant genotypes. A tentative map is presented using data from cis- and trans-crosses, including a correction by omission of uniform clones. Phenomena of transmission, segregation, and formation of uniform clones are discussed with special regard to the difference brought about by fission versus budding. A comparison with relevant data from Saccharomyces cerevisiae and other organisms is presented.

Anti-Bacterial Agents

Quantitative models of hybrid dysgenesis: rapid evolution under transposition, extrachromosomal inheritance, and fertility selection.

A model of the P-M system of hybrid dysgenesis is presented which incorporates single-site transposition of P factors in M cytotype, determination of offspring cytotype by both maternal cytotype and maternal or offspring nuclear genotype, and strong fertility selection in dysgenic individuals. The conditions required for the initial invasion of P factors into a pure M population, information concerning stable polymorphisms, and results of numerical iterations depicting the dynamic, nonequilibrium behavior of the system are summarized. While conditions for initial increase are independent of the rate of cytotype switching, the rate of evolution is accelerated by increased production of dysgenic individuals. If the transposition rate is sufficiently high to overcome the fertility barrier opposing P factors introduced into M populations, then convergence to high frequencies of the P factor occurs very rapidly. Under intense fertility depression, the phase of rapid increase may be preceded by an extended period of gradual increase at low frequencies.

Animals

Inheritance of extrachromosomal ribosomal DNA during the asexual life cycle of Dictyostelium discoideum: examination by use of DNA polymorphisms.

Wild-type isolates of Dictyostelium discoideum exhibited differences in the size of restriction fragments of the extrachromosomal 88-kilobase ribosomal DNA (rDNA) palindrome. Polymorphisms in rDNA also were found among strains derived solely from the NC4 wild-type isolate. These variations involved EcoRI fragments II, III, and V; they included loss of the EcoRI site separating fragments II and V and deletion and insertion of DNA. More than one rDNA form can coexist in the same diploid or haploid cell. However, one or another parental rDNA tended to predominate in diploids constructed, using the parasexual cycle, between haploid NC4-derived strains and haploid wild-type isolates. In some cases, most if not all of the rDNA of such diploids were of one form after ca. 50 generations of growth. Segregant haploids, derived from diploids that possessed predominantly a single rDNA allele, possessed the same allele as the diploid and did not recover the other form. This evidence implies that replication does not proceed from a single chromosomal or extrachromosomal copy of the rDNA during the asexual life cycle of D. discoideum.

Alleles