Zelek S. Herman

Functional profiling of the Saccharomyces cerevisiae genome

Determining the effect of gene deletion is a fundamental approach to understanding gene function. Conventional genetic screens exhibit biases, and genes contributing to a phenotype are often missed. We systematically constructed a nearly complete collection of gene-deletion mutants (96% of annotated open reading frames, or ORFs) of the yeast Saccharomyces cerevisiae. DNA sequences dubbed ‘molecular bar codes’ uniquely identify each strain, enabling their growth to be analysed in parallel and the fitness contribution of each gene to be quantitatively assessed by hybridization to high-density oligonucleotide arrays. We show that previously known and new genes are necessary for optimal growth under six well-studied conditions: high salt, sorbitol, galactose, pH 8, minimal medium and nystatin treatment. Less than 7% of genes that exhibit a significant increase in messenger RNA expression are also required for optimal growth in four of the tested conditions. Our results validate the yeast gene-deletion collection as a valuable resource for functional genomics.

Systematic screen for human disease genes in yeast

High similarity between yeast and human mitochondria allows functional genomic study of Saccharomyces cerevisiae to be used to identify human genes involved in disease. So far, 102 heritable disorders have been attributed to defects in a quarter of the known nuclear-encoded mitochondrial proteins in humans. Many mitochondrial diseases remain unexplained, however, in part because only 40–60% of the presumed 700–1,000 proteins involved in mitochondrial function and biogenesis have been identified. Here we apply a systematic functional screen using the pre-existing whole-genome pool of yeast deletion mutants to identify mitochondrial proteins. Three million measurements of strain fitness identified 466 genes whose deletions impaired mitochondrial respiration, of which 265 were new. Our approach gave higher selection than other systematic approaches, including fivefold greater selection than gene expression analysis. To apply these advantages to human disorders involving mitochondria, human orthologs were identified and linked to heritable diseases using genomic map positions.

Integrative Analysis of the Mitochondrial Proteome in Yeast

In this study yeast mitochondria were used as a model system to apply, evaluate, and integrate different genomic approaches to define the proteins of an organelle. Liquid chromatography mass spectrometry applied to purified mitochondria identified 546 proteins. By expression analysis and comparison to other proteome studies, we demonstrate that the proteomic approach identifies primarily highly abundant proteins. By expanding our evaluation to other types of genomic approaches, including systematic deletion phenotype screening, expression profiling, subcellular localization studies, protein interaction analyses, and computational predictions, we show that an integration of approaches moves beyond the limitations of any single approach. We report the success of each approach by benchmarking it against a reference set of known mitochondrial proteins, and predict approximately 700 proteins associated with the mitochondrial organelle from the integration of 22 datasets. We show that a combination of complementary approaches like deletion phenotype screening and mass spectrometry can identify over 75% of the known mitochondrial proteome. These findings have implications for choosing optimal genome-wide approaches for the study of other cellular systems, including organelles and pathways in various species. Furthermore, our systematic identification of genes involved in mitochondrial function and biogenesis in yeast expands the candidate genes available for mapping Mendelian and complex mitochondrial disorders in humans.

Effects of dietary phytic acid (phytate) on the incidence and growth rate of tumors promoted in Fischer rats by a magnesium supplement

Abstract A Fischer rat tumor model was used to examine the effects of (i) dietary magnesium supplementation on tumor incidence, rate of tumor growth and latent periods for tumor appearance and death, and (ii) addition of phytate to the magnesium-supplemented diet on the same evaluation endpoints. Fifty, three-month old female Fischer rats were assigned to each of three isocaloric test diets fed ad libitum. The diets tested were: (i) certified rodent chow 5002 alone, (ii) chow plus 1.4% magnesium oxide (MgO), and (iii) chow plus 1.4% MgO plus 12.6% phytate. After two weeks each rat was injected subcutaneously with a syngeneic, tumorigenic cell line (1×10 6 cells per animal). Respective test diets were continued for an additional 23 weeks following cell injection. Rats were sacrificed when tumors reached a limit length or width of 4 cm. The effect of magnesium supplementation was evaluated by comparing the chow group to the MgO test group. The effect of phytate addition was evaluated by comparing the chow+MgO group to the chow+MgO+phytate group. Dietary magnesium supplement exhibited a statistically significant tumor-promoting effect by all four evaluation endpoints measured. Dietary phytate, by the same four evaluation endpoints and all statistical parameters, nullifed the tumor-potentiating effects induced by MgO. Phytate addition highly significantly lowered tumor growth rate (p

On the use of the character projection operator in the determination of the symmetry of molecular orbitals and in the construction of hybrid bond orbitals

The utility of the character projection operator in determining the symmetry of molecular orbitals and in the construction of hybrid bond orbitals is discussed. Examples from the authors work are mentioned. These are topics which this author discussed in detail with Jean-Louis Calais during the past 25 years.

Protection by dietary ascorbate of guinea pigs from neurolathyrism

The protection afforded by dietary vitamin C from the toxic effects of intraperitoneal injection of an aqueous solution of an air-dried ethanol extract of Lathyrus sativus peas is demonstrated in guinea pigs fed an ascorbate-fortified diet in contrast to those fed a diet devoid of vitamin C. Weakness and flaccid paraparesis occurred in most of the ascorbate-depleted animals, whereas all guinea pigs on the ascorbate-supplemented diet remained active and appeared well following injection of the extract. These results extend previous work on the role of dietary ascorbate in the protection of guinea pigs from neurolathyrism. In particular, spastic paralysis in some ascorbate-depleted guinea pigs is reported for the first time. Since guinea pigs, like all primates, including humans, do not synthesize vitamin C, their use as experimental models can partially simulate human neurolathyrism.

Ionic crystals: A simple and safe lecture demonstration of the preparation of NaI from its elements

A simple and safe classroom demonstration showing the production of sodium iodide (NaI) crystals from elemental sodium and elemental (molecular) iodine is presented. The demonstration, which is quite impressive, naturally fits into the discussion of ionic bonding and the alkali halide crystals. It is best suited for a classroom or laboratory containing fewer than 75 students. Suggestions are made for using the product in other topics covered in the course.

A Model of Early Cancer: The Effect of Ascorbate and Its Oxidation or Degeneration Products on Tumor Incidence and Growth in Syngeneic Guinea Pigs Given Minimal Numbers of Tumor Cells

In a series of experiments, minimal numbers of tumor cells were injected into guinea pigs, and the effect of administering vitamin C in their drinking water on tumor onset and tumor growth rate was investigated. In some of the work, air was bubbled through the water to ascertain if ascorbate or its oxidation products had any different effect on the onset times and growth rates. In all experiments, it was found that adding vitamin C to the drinking water significantly increases the tumor onset time but has no discernible effect on the tumor growth rate once the tumor is palpable. No significant change in the tumor-free survival times or the tumor growth rates results from bubbling air through the ascorbate-supplemented drinking water.