genetics of fungi

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SEMINAR ON GENETICS OF FUNGI PRESENTED BY priyanka das(M.Sc Mb)

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CONTENTS 1.YEAST INDUCTION OF MUTATION 2.CYTOPLASMIC INHERITANCE Definition Patterns of inheritance Comparison Extra nuclear inheritance in yeast Significance 3.BIO-CHEMICAL MUTANTS 4.REFERENCES

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YEAST INDUCTION OF MUTATION The induction of mutation in yeast by hydrogen peroxide The inactivation and mutation to respiratory deficiency of yeast cells by H2O2 are shown to vary progressively with the phase of cell growth. Respiratory deficient mutants isolated from the wild-type population are of two types, one of which is much more sensitive to H2O2 but forms only a small fraction of the mutant sub-population. Based upon the response of the more resistant type, mutation frequency increases appear to result from selection of pre-existing mutants in log phase populations, while induction occurs in stationary phase cells. All the induced mutants are extranuclear and of the resistant type, and show a wide range of suppressiveness in crosses to respiratory competent cells. This may indicate mitochondrial DNA is altered to different extents by H2O2. A small increase in forward mutation of nuclear genes was also found after H2O2 treatment.

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A haploid strain of the yeast, Saccharomyces cerevisiae, wild-type with regard to radiation sensitivity, was irradiated with 254-nm ultraviolet (UV) light and then exposed to X-rays after incubation for 0-6 hr. The cells were incubated either on nutrient medium or salt agar between the treatments. Loss of reproductive ability and mutation to canavanine resistance were measured. Incubations up to about 3 hr on nutrient medium between the treatments led to synergism with respect to cell inactivation and antagonism with respect to mutation, but after 4-6 hr the two treatments acted independently. Incubation on salt agar did not cause any change in the survival curves, but there was a strong suppression of X-ray-induced mutation with increasing UV fluence. On the basis of these results, we suggest that mutation after combined UV and X-ray exposure is affected not only by the induction and suppression of DNA repair processes, but also by radiation-induced modifications of cell-cycle progression and changes in the expression of the mutant phenotype. INDUCTION OF MUTTION AFTER SPLIT DOSE RADIATION

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EFFECT OF TEMPERATURE The budding yeast S.cerevisiae have been used to identify mutants that interfered with the cell cycle (cdc mutants). Leland hartwell & Paul Nurse- nobel prize. The screen was for conditional heat sensitive cdc mutations,which are wild type at low temperature but mutant at high . They result from alpha changes that lead to protein shape changes at high temperature.

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These mutants can be propagated at r.t. (permissive temperature) and then shifted to higher temperature(restrictive temperature) ,at which they express the mutant phenotype. The set of mutant derived from this type of screen has enabled researchers to define many of the proteins that regulate the highly programmed progression through the cell cycle. CONT…

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C YTOPLASMIC INHERITANCE INHERITANCE- A thing that is inherited (derive from ones parents). PATTERNS OF INHERITANCE- Based on the different possible location of that single gene. 1.Autosomal inheritance- based on variation of single gene on regular chromosomes (autosomes). 2.Sex-linked inheritance -based on variation of single genes on the sex determining chromosomes. 3.Cytoplasmic inheritance -based on variation of single genes on organelle chromosome.

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C YTOPLASMIC INHERITANCE- It is inheritance of characteristics coded for by genes found in the cytoplasm, typically from the mitochondria or chloroplasts .Because the cytoplasm is usually contributed entirely by one parent, the characteristics encoded by these genes are usually inherited from only that parent.

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A. Extra nuclear inheritance in eukaryotes 1.Maternal inheritance- shell coiling in limnaea 2.Extra nuclear inheritance by cellular organelles Mitochondria of yeast Mitochondria of poky strain of Neurospora 3.Uniparental inheritance Chlamydomonas reinhardi B. EXTRA NUCLEAR INHERITANCE IN PROKAR YOTES 1.Episomes 2.Plasmids

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EXTRA NUCLEAR INHERITANCE B Y MITOCHONDRIA OF Y EAST S.cerevisiae- unicellular, ascomycota fungi Life cycle- diploid & haploid adult aiternates - former reproduce by ascospores ,latter by isogametes Can grow with or without O2 In absence of O2 , yeast obtain energy for growth & metabolism through fermentation, in which mitochondria are not involved In presence of O2 , mitochondria carry out aerobic respiration & facilitate a faster growth rate than is the case in the absence of O2.

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Characteristics of petite mutants- When yeast cells are spread on agar medium containing glucose, a very low proportion of the cells give rise to colonies that are much smaller than wild type colonies. DISCOVERER-Boris Ephrussi (French) The small colonies are called petites, wild type colonies sometimes are called grandes. BIOCHEMICAL ANALYSIS-shows that petites have cytochrome deficiencies & therefore are essentially incapable of carrying out aerobic respiration, they must obtain their energy primarily from fermentation, which is a less efficient process. In an unmutagenized population of cells,0.1 to 1% of the cells spontaneously become petite. In presence of intercalating mutagen such as ethidium bromide,acridine dye, or euflavine, 100% of the cells become petite.

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Yeast petite system is useful in studies of non-mendelian inheritance because yeast cells that lack mitochondrial function can still survive and grow. Such mutations to respiratory deficiencies in yeast are automatically conditional mutants. CONT…

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Different types of petite mutants- Crosses between petites & wild type were made to determine how petites are inherited. Yeast are haploid &crosses involve fusing two cells ,one of mating type a &one of mating type alpha to produce a diploid zygote. That zygote can be grown into a colony to check its phenotype. When the zygote goes through meiosis , the 4 haploid meiotic products-the sexual spores (ascospores) are contained within an ascus. Tetrad analysis can be done ,that is ,the analysis of all 4 products of meiosis to determine segregation patterns.

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Segregational petites- Some petites ,when crossed with wild type. 2:2 segregation of wild type :petite in tetrads Segregation pattern is that found for nuclear gene mutation, so called nuclear petites. Pet¯

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Neutral petites- Shows non-mendelian inheritance rho¯N rho¯N * rho+N Diploids all produce wild type colonies When these diploids go through meiosis,all resulting tetrads show a 0:4 ratio of petite:wild type.while nuclear markers segregate 2:2 The name neutral refers to that fact ,that this class of petites does not affect the wild type. This result is example of uniparental inheritance. This phenomenon is not maternal inheritance,however because the two haploid cell that fuse to produce the the diploid are the same size &contribute equal amount of cytoplasm.

Cotinued…………. : 

Cotinued…………. Prolonged treatment with an intercalating agent , the majority of petites produced are neutral petites. The mitochondrial genome is implicated in these mutant because cytochromes are attend ,because this is evidence for extra nuclear inheritance & because the mitochondria are the other site in the yeast cell in which genetic material is found.

SUPPRESS PETITES : 

SUPPRESS PETITES Inheritance pattern is different from other types : when {rho+} / { rho-s} diploid is formed ,it has respiratory properties intermediate between those of normal & petite strains. If this diploid divides mitotically a no. of times ,the diploid population produced is up to 99% petites.

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The name suppressive ,then refers ,to the fact that this class of petites overwhelms the w.t. so that a respiratory deficient phenotype result. Sporulation of any of the petites in that population produce tetrads with a 4:0 ratio of petite : w.t. Sporulation of any of the few w.t. diploid in the population produce tetrads with a 0:4 ratio of petites :w.t . These petites have changes in mt DNA . Petites colony results due to deficiencies in the enzymes involved aerobic respiration.

UTILITY : 

UTILITY Advantages of many self reproducing cellular organelles in eukaryotes : 1} It prevents total loss of organelles due to a single mutation in nuclear genes. 2} It provides a reservoir of cytoplasmic mutation which would become useful under adverse environmental condition.

Biochemical mutants : 

Biochemical mutants The term mutants refers to an organism in which either base sequence of DNA has been changed. Those definition are actually the same since the base sequence of DNA determines the amino acid sequence of a protein. The chemicals & physical properties of each protein are determined by its amino acid sequence ,so a single amino acid change is capable of attiring the activity of ,or even completely inactivativating ,a protein.

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Continued…………… For e.g.. A protein whose 3 – D structure is determined entirely by an interaction between one positively charged amino acid { for e.g., lysine} & one negetively charged amino acid ( aspartic acid ). A substitution of methionine , which is uncharged ,for the lysine would clearly destroy the 3 – D structure.

Biochemical mutants of yeast cells with impaired oxidation – : 

Biochemical mutants of yeast cells with impaired oxidation – Under the influences of trypafauine , camphor & U. V light yeast cells produces biochemical mutants with impaired oxidation . This impairment of the oxidation apparatus is regularly transmitted to succeeding occur a period of many months. An attempt was made to develop method for specific coloration of colonies ,in order to distinguish the mutant colonies .by there color , form the colonies of normal forms surroundings there . The best results where obtained with the leucolase of methylene blue.

Temperature sensitive mutants of yeast exhibiting a rapid inhibition of protein : 

Temperature sensitive mutants of yeast exhibiting a rapid inhibition of protein Certain temperature sensitive (t5) mutants of yeast which cannot the corrected by nutritional supplementation exhibited a rapid cessation of protein synthesis after a shift to the restrictive temperature . Genetic & biochemical test permitted a division of these mutants in to 4 classes . This division was based upon genetic complementation patterns among the mutants & are investigation of glucose incorporation in to macromolecules & polyribosome content in the mutants after a shift to the restriction temperature . A study of these parameters in the parent strains (ts+) in the presences of certain well characterized inhibition allowed a tentative identification of the biochemical defects in each of the four classes .

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Continued………… The properties of the mutants in class IA were consistent with the hypothesis that they result from a defect in the initiation polypeptide chains or in RNA synthesis ; mutants in class IB from a defect in the elongation of polypeptide chains . Mutants in class ii A from a defect in energy metabolism ; & mutants in class ii B from a lesion affecting membrane function. Ctoplasmic inheritance – it is inheritance of characteristics coded for by genes found in the cytoplasm ,typically from the mitochondria or chloroplast ,because the cytoplasm is usually contributed entirely by one parent ,the characteristics encoded by these genes are usually inherited from only thet parent.

REFERANCES : 

REFERANCES 1.Genetics-Analysis of genes & genomes(6th edition) Daniel L.Hartl,Elizabeth jones 2.Introduction to genetic analysis(8th edition) Griffith.wessler.suzuki.miller 3.Genetics-B.D.Singh 4.www.google.com 5.www.pubmedcentral.nih.gov 6.www.Springerlink.com 7.www.sciencelink.com

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