Adrian M. Srb

Immunofluorescent localization of a phase-specific protein in Neurospora tetrasperma perithecia

Neurospora tetrasperma perithecial sections were stained with fluorescein isothiocyanate-conjugated antibodies produced against a Neurospora phase-specific perithecial protein. Analysis of the sections demonstrates the localization of the protein antigen in the non-cellular material surrounding the asci.

Selection for increased growth rate in inter-and intra-strain crosses of Neurospora

ANYONE studying quantitative inheritance almost automatically enCounters complex problems which can usually be solved only partially or solved in the light of certain assumptions. An organism and experimental procedure chosen to minimise some of these complexities would be extremely desirable. The haploid inheritance in heterothallic Neurosporas, the short life cycle, the absence of heterosis and the ability to produce large numbers of offspring, both sexually and asexually, make certain species of this genus well suited for model studies of quantitative inheritance. Problems of gene action, mutagenesis and recombination have been widely studied in microorganisms but little has been done with polygenic systems. Genetic variability must exist for a selection program to be effective, but storage of variability in heterozygotes is not possible in haploids. Bodmer and Parsons (1962) pointed out that the necessity for storing potential genetic variability is as important for microorganisms as for other organisms. The only mechanism capable of storing such variability, however, in the absence of heterozygosis or heterocaryosis, is the balanced polygenic complex. Although very few selection experiments have been conducted on lower forms, an extensive literature has accumulated pertaining to selection in higher plants and animals. Chapman (1961) has compiled an extensive bibliography pertaining to selection in laboratory animals. Kojima and Kelleher (1961) have presented general information from Drosophila and mice experiments relevant to the planning of selection experiments and the understanding of genetic changes accompanying these experiments. Polygenic inheritance in Xeurospora was studied by Pateman (1955), (igg), Pateman and Lee (ig6o), Lee and Pateman (1961), and Lee (i 962). In studying the effect of selection on ascospore size, they showed that phenotypic effects of polygenes on ascospore length were not simply additive but included a considerable amount of gene interaction. Linkage of part of the polygenic activity to the albino locus was also demonstrated. Simchen and Jinks (1964), Simchen (1965) and Croft and Simchen

The isolation of mutants affecting ascus development in neurospora crassa and their analysis by a zygote complementation test.

The mutant gene peak-2 [pk-2) in Neurospora crassa is located in linkage group V. In the haploid vegetative mycelium it determines dichotomous branching and colonial growth habit. It also affects the sexual reproductive apparatus by determining abnormal, non-linear asci, for the most part eight-spored, in contrast to the linear, eight-spored asci of wild type. In this system, pk-2 acts as a zygote recessive. That is, asci initiated by zygotes that are homozygous wild type ( + / + ), or are heterozygous ( + /pk-2), are morphologically normal, showing the linear arrangement of ascospores; asci initiated by homozygous mutant zygotes (pk2jpk-2) are morphologically abnormal, with the arrangement of ascospores differing markedly from linearity (Murray & Srb, 1959, 1962). Controlled reciprocal crosses have shown that heterozygotes develop normal ascus morphology whether the mutant or the wild type has been used as maternal parent (Srb, 1962). Maternal effects, therefore, do not offer a plausible alternative explanation to this apparent instance of zygotic control of ascus morphology. Although the ultimate differentiation of the ascus may be influenced by the meiotic products of the zygote and by their immediate mitotic derivatives, the zygote nucleus at least has a determinative influence on the initiation of ascus development. This conclusion is based on the cytological observation that a characteristic difference in the cellular shape of wild or peak-2 asci is apparent even before meiosis begins. The foregoing observations and their interpretation focus on the single diploid cell, the zygote, that is known to exist in the normal life cycle of Neurospora. Having, then, a genetic system at least partly under the control of the diploid nucleus, the possibility arises for utilizing some of the procedures of diploid genetic analysis for carrying out studies of the ascus of Neurospora. The most obvious possibility is that of diploid complementation analysis. Specifically, if one has two abnormal ascus mutants of independent origin (e.g. mutant-a and mutant-b), and if each acts as a zygote recessive in matings with wild type, then the mating giving a zygote mut-ajmut-b should provide a test for functional identity or non-identity of the pertinent genetic material. The initial test would be exceedingly simple— mere observation for the appearance of normal or abnormal asci. Normal asci would indicate complementation and be interpreted as the consequence of functional non-identity. Thus, in principle, the zygote complementation scheme just

Inheritance of growth rate in Neurospora crassa: reverse selection in an improved strain.

A CLASSICAL method of determining limits to selection responses has been to measure response when selection is applied in the opposite direction. If genotypic fixation of the selected lines has occurred, then response in either direction will not be possible unless new genetic variation arises or is induced. Numerous studies on the nature of selection limits and related phenomena have been conducted on a variety of organisms. In many instances genetic variability still existed in selected lines, yet additional selection responses were not realised. Presumably, this genetic variability could not contribute to further selection responses. Experimental evidence in support of this notion has been offered and encompasses various phenomena. In certain instances, natural selection in the form of differential fertility was found to be opposing artificial selection (Falconer, Lee and Pateman, 1961). A. Robertson (1955) reported that the maintenance of high genetic variance, although response to selection had ceased, could occur as a result of a negative correlation between the character selected and fitness components. Clayton and A. Robertson (ig,) attributed unfixable genetic variation in selected lines to the selection of heterozygotes for a lethal gene. The possibility that a physiological limit may set upper limits on a selected character was examined by Falconer and King On crossing individual strains of mice which presumably had reached selection limits, they observed a renewed response to selection for increased and decreased body size. Lines having reached apparent selection limits due to the absence of free genetic variability might still possess potential genetic variability. Utilisation of this potential genetic variability, however, would depend on breaking up rather tight linkages to get recombinant individuals of superior phenotype (Mather, 1943). Theoretical treatments of linkage and selection, although quite complex, have been partially examined by Fraser (i7, 1960) using Monte Carlo methods. Recent quantitative inheritance studies in various Neurosporas have shown linear growth rate to be under the control of several to many genes (Papa, Srb and Federer, 1966). The slow and gradual increase in growth rate during artificial selection was indicative of a polygenic

A study of L-glutamine: D-fructose 6-phosphate amidotransferase in certain developmental mutants of Neurospora crassa

SummaryWild type and 22 mutant strains of Neurospora crassa were compared for L-glutamine: D-fructose 6-phosphate amidotransferase (GFAT: EC 2.6.1.16) specific activity. The mutant genes map at eleven functionally nonallelic loci, mutations at which result in altered mycelial and ascus morphologies. The enzyme GFAT, which is involved in the synthesis of the important mycelial wall constituent chitin, was investigated as a possible causal factor in determining the altered morphologies. When compared with the wild-type (control) activity, a statistically increased GFAT activity was detected in crude extracts of six mutant strains. Two of these strains carry mutant alleles mapping at the peak locus (peak-2 and clock) and the remaining four carry mutant genes mapping at four other loci. GFAT activity was not significantly different from that of wild type in crude extracts prepared from nine other strains carrying mutant alleles that map at the peak locus, or from seven strains carrying the remaining mutant alleles mapping at other loci. In the two cases tested, including that of peak-2, the activity increase segregated with the mutant morphological phenotype when cultures derived from ascospores of tetrads isolated from crosses of the mutants with wild type were assayed.Further studies with crude extracts showed that, compared with wild type, GFAT activity of peak-2 is higher at all growth times between 12 and 72 hours, and the enzyme is more thermolabile at 35°C. The higher GFAT activity of peak-2 was shown probably not to be due to differential feedback inhibition.The peak locus is almost certainly not a structural gene for the enzyme GFAT, since only two out of the eleven strains carrying mutant alleles at the peak locus that were assayed for GFAT activity showed increased activity compared with that of wild type. For the four nonallelic mutants showing higher GFAT activities than wild type, evidence is still insufficient to determine whether or not the appropriate loci are structural genes for GFAT. The increased GFAT activity where found is consistent with, and possibly contributory to, the altered morphologies.

Unbiased second-division segregation in Neurospora

Results of crosses of a large number of Neurosporas of different origin, including several distinct strains of N. sitophila , were utilized to re-examine the question of whether certain wild-type Neurosporas other than N. crassa show biases in the two types of second-division segregation. Segregations for alleles of the mating type and peak loci on a wide variety of genetic backgrounds gave little evidence for excess either of symmetrical or asymmetrical ‘post-reduction’ asci.

Recurrent changes in colony morphology dependent on intracolony selection of prototrophs occurring in red yeast 1

Recurrent changes in colony morphology dependent on intracolony selection of prototrophs occurring in red yeast 1

Tartaric Acid Metabolism of Neurospora crassa

The growth of wild type Neurospora crassa is stimulated by various organic acids including tartaric, tartronic, and mesoxalic acids. Evidence is presented that this organism converts d- or l-tartaric acid to tartronic and mesoxalic acid, probably by fixation of CO2.