Henry R. Mahler

Glycoproteins of the Synapse

A consideration of the current state of our knowledge (or should I say ignorance) of the glycoproteins in CNS synapses is pivotal to an understanding of the function of the complex carbohydrates of nerve tissues. Just as snyapses fulfill the crucial role in the establishment, construction, reconstruction, and reinforcement of neuronal pathways of communication, so do glycoproteins subserve the task of establishing and maintaining contact among cells and provide a framework for information transfer at the molecular level. Thus, they may provide key structural and sensory elements not only in the maintenance of synaptic contacts but also in their establishment during synaptogenesis and in their possible modification as a result of experience.

Studies in partially resolved bacteriophage-host systems. VII. Diamines, dyes, empty phage heads, and protoplast-infecting agent.

Abstract The photodynamic inactivation of bacteriophage T 2 and oX-174 in the presence of proflavine has been investigated. The reaction has been shown to be photochemical in nature and has been interpreted in terms of the formation of a reversible complex between the DNA of the phage and the dye. The approximate dissociation constants for these complexes have been estimated at 3·10 −6 M for T2 and 3·10 −7 M oX-174. This binding is competitively reversed in the presence of aliphatic diamines with no particular specificity f chain length. The approximate dissociation constant for cadaverine is 3.5·10 −3 M for the DNA of either phage. Some preliminary observations concerning the structural requirements of other photodynamically active dyes are presented: methylene blue and acridine organge, which as bifunctional amines bear a close resemblance to proflavine, are active; 9-aminoacridine and riboflavin, both of which are otherwise photochemically effective, lack the necessary structuralbifunctionality and are inert as photodynamic sensitizers. A brief survey has also been made of the ability of proflavine to inactivate both bacteriophages in the dark. This effect resembles photodynamic inactivation with respect to the concentration dependence and the amount of dye required to achieve half-maximal saturation. Dark inactivation is prevented competitively by aliphatic diamines; no chain-length specificityis observed with oX-174, but T2 is maximally protected by cadaverine. In this reaction the amine serves in a function presumably analogous to that responsible for the protection against thermal inactivation of π, the protoplast-infecting agent derived from T2, and for the shift in melting temperature of isolated T2 DNA.

Modulation of petite induction by low concentrations of ethidium bromide.

Abstract When starved, exponential phase cells of Saccharomyces cerevisiae are exposed to 10–25 μM ethidium bromide in buffer, the proportion of respiration deficient mutants in the population exhibits a rapid increase, followed by a pronounced decrease. This “self-rescue” by the mutagen can be dissociated from and studied independently of its mutagenic action and is shown to exhibit different requirements. These and related observations have been used to formulate a consistent model for mutagenesis by ethidium bromide and its modulation under a variety of conditions.

Biogenetic Autonomy of Mitochondria and Its Limits

The past ten years have seen such an almost explosive growth of efforts devoted to studies on mitochondrial biogenesis that they occupy at least as much contemporary journal space as do studies on mitochondrial function and energetics, their much more venerable companions. The results of this endeavor have been fairly spectacular: the documentation of the existence and the characteristics, in molecular terms, of a novel and separate system for the maintenance, duplication, and expression of genetic information within the organelle and its interaction with the classical nucleocytosolic system of the eukaryotic cell (for recent reviews see refs. 1–10).

Genetic Autonomy of Mitochondrial DNA

As a result of studies during the past ten years, the presence in the mitochondria of all “ordinary” eukaryotic cells — from protists to the most complex metazoa, including man — of a system for the storage, transmission and expression of genetic information can now be considered to rest on a firm experimental foundation (Ashwell and Work, 1970; Rabinowitz and Swift, 1970; Schatz, 1970; Preer, 1971; Boardman et al., 1971; Borst, 1972; Linnane et al., 1972; Sager, 1972; Mahler, 1973a). These investigations have also established that, although this second system is distinct and separate from the primary (nuclear-cell sap) system in the same cell — by virtue not only of its sequestration within the organelle but also by the properties of its components and of the reactions that they catalyze — it is by no means independent of it. Since these conclusions and the experiments on which they rest have been repeatedly and recently reviewed (see above), this presentation is restricted to a consideration of the nature, extent and limits of the biogenetic autonomy of the mitochondria of just one organism, Saccharomyces cerevisiae, baker’s yeast. It will make use of our own data, both published and some yet unpublished, as well as on studies by several groups here and abroad who are making use of the same experimental system. It will be convenient to broach the question in such a way as to work backwards from the gene products to the genes and to start with the translational products unique to mitochondria.

Synthesis of Mitochondrial Proteins

The recent discovery that the ubiquitous energy-transducing organelle of eukaryotic cells, the mitochondrion, contains its own DNA, distinct from that of the nucleus (for recent reviews, see 1-4), has generated intensive investigations in a number of related areas. They have made accessible possible answers to such fundamental questions as the nature and extent of the genetic and biogenetic autonomy of the organelle; the origin, mode of transmission, and significance of extrachromosomal, non-Mendelian hereditary determinants; and the whole chain of events that must intervene in the course of the expression of the mitochondrial genome. Central to this area is the problem of mitochondrial protein synthesis, both in vivo and in vitro; known now for some 13 years (5,6), it has been raised from its former status of relative obscurity as a laboratory curiosity to one of the fashionable fields of inquiry of contemporary biochemistry and molecular biology. Since the topic has been the subject of several recent and comprehensive reviews (7ߝ11), I shall restrict my presentation, in the main, to our own investigations with the unicellular eukaryote Saccharomyces cerevisiae or baker’s yeast.

Cytochrome b of the Respiratory Chain

As originally pointed out by Keilin and Hartree (1939), to whom we owe most of the basic definitions in this general field, cytochrome b can be distinguished from the other cytochromes of the respiratory chain, i.e., c and aa 3, on the basis of the following characteristic properties: an oxidation reduction potential permitting its ready reduction by succinate in the presence of the respiratory chain, or chemically by ascorbate, and its oxidation by cytochrome c., an absorption spectrum with its α-, β-, and γ-bands centered around 564, 530, and 432 nm, respectively, and a lack of reaction with typical heme ligands such as CN−. The reported CN−-insensitive oxidation by O2 of the reduced form of the non-covalently attached heme (ferro-protoporphyrin IX) prosthetic group is now known not to be a property of membrane-bound cytochrome b. Since then, a large number of additional cytochromes of the b type (cytochromes B) have been discovered in animal, plant, fungal, and bacterial cells and given rise to a bewildering array of both trivial (i.e., cytochromes b 1,b 2,...b 6) and systematic (e.g., cytochromes b-556, b 556, b-558, or b 558, and b-562 or b 562 of E. coli) designations. The discussion in this essay will be restricted to the classical form of cytochrome b tightly associated with the inner membrane of animal and fungal mitochondria, where it contributes to two segments of their respiratory chain (see below); about 80% of the cytochrome forms part of ubiquinol-cytochrome c reductase (complex III) and the remainder is associated with succinate-cytochrome c reductase (complex II). The reader is referred to a recent review (von Jagow and Sebald, 1980) for a discussion of nonmitochondrial and bacterial cytochromes of the b type and to an article by Bowyer and Trumpower (1981a,b) for a comparison of the mitochondrial bc 1 complex with a similar entity found in photosynthetic bacteria.

Specification and Expression of Mitochondrial Cytochrome b

Recent studies on the structure, organization, and expression of the mitochondrial gene for (apo)cytochrome b in Saccharomyces cerevisiae and related species have provided answers to some old questions and revealed some hitherto unexpected complexities of regulatory and, possibly, evolutionary significance.* To the first set belong, most importantly, the primary structure of the protein, a problem that had defied solution by protein sequencing techniques, but was solved by Nobrega and Tzagoloff (1980) by DNA sequencing: the polypeptide chain consists of 385 amino acids, corresponding to a molecular weight of 44,000. Another question that has been resolved without ambiguity concerns the type and number of polypeptides responsible for the multiplicity of cytochrome b species implicated in the b c 1 segment (Complex III, coenzyme QH2: cytochrome b oxidoreductase) of the mitochondrial respiratory (electron transfer) chain (reviewed by von Jagow and Sebald, 1980). Because single, defined, revertible mutational lesions in the coding segments of a unique and discrete gene, known as cob, are now known to result in the complete elimination or alteration of all forms of the cytochrome in question (Claisse et al., 1978; Mahler et al., 1978; Alexander et al., 1979; Haid et al., 1979; Kreike et al., 1979), all such cytochromes must have been synthesized with the same primary sequence. The different species observed may therefore be the consequence of placing the same polypeptide in different environments, of posttranslational modification, or a combination of these two effects. Related to this problem, and equally accessible by a combination of genetic and biochemical techniques, is the localization of sites of interaction between the polypeptide on the one hand, and heme or various characteristic inhibitors of Complex III (such as antimycin A, diuron, funiculosin and mucidin) on the other.

The influence of social values on public policy determination

ysis of values which influence public policy decision-making at two levels of government-international and metropolitan. The immediate object of the study is to test the applicability of different methods of behavioral analysis to the study of this problem. The ultimate purpose is to develop better understanding of the factors which impede or contribute toward effective cooperation among relatively autonomous governmental units in the solution of common problems.