M. Sela

General aspects of nucleic acid uptake by mammalian cells have been the subject of several reviews during the last few years (PAGANO, 1970; BHARGAVA and SHANMUGAM, 1971; DUBES, 1971; RYSER, 1967). These reviews covered methods used for the infection of cells by viral nucleic acids as well as interaction of mammalian cells with non-viral nucleic acids. This article is restricted to a discussion of experiments with poliovirus RNA and focuses special attention on the steps following the uptake of RNA into a cell, aspects that were not discussed in earlier review articles. The fate of input RNA once inside the cell is determined by the host cell but experimental conditions can be chosen to favor the survival of input RNA and the induction of a virus growth cycle by interfering with host-cell meta­ bolism through events that, in the case of infection with intact virus, might be controlled by viral proteins.

Immunoglobulin gene expression appears to include a number of unique features (Cohn, 1971; Gaily and Edelman, 1972; Hood et aI. , 1975). First, a variety of genetic and protein structural evidence suggests that two discrete genes - both a variable region gene and a constant region gene - specify each heavy chain and each light chain. This constitutes the twogene-one polypeptide hypothesis. Second, a single differentiated lymphocyte or plasma cell appears to express only one heavy chain allele and one light chain allele at a time. This is the only example of allelic exclusion known in mammalian cells except for X chromosome inactivation. Third, during the course of lymphocyte differentiation, there may be a switch of the heavy chain constant region gene expression but no change in the heavy chain variable region gene expression. Rarely, normal or malignant cells have been found which express two different heavy chain subclasses simul­ taneously (e. g. see Sledge et aI. , 1976). Fourth, the vast number of different antibodies which can be made by an individual animal has raised the question of whether the generation of diversity occurs during evolution or within the animal, i. e. germ line vs somatic variation. Other aspects of immunoglobulin gene ex­ pression may be similar to regulation of gene expression in many eukaryotic cells.

At the end of the last century and the beginning of this century, the prob­ lems of immunity in lower vertebrates and the influence of environmental temperature attracted attention for the first time (ERNST, 1890; WIDAL and SICARD, 1897; METCHNIKOFF, 1901). However, relatively little work has been done on this subject until recently. The early investigators were chiefly in­ terested in the immuno-pathological problems. They immunized various species of lower vertebrates essentially with bacterial vaccines; agglutinating, neutralizing and protective antibodies were detected in their blood. The in­ fluence of environmental temperature on the immune response was investigated, since this subject represented great economical and theoretical importance. Epizootic diseases were observed to occur in relation to the cold season of the year, when the decrease or spontaneous increase of water temperature occurred (SCHAPERCLAUS, 1965; BESSE et al. , 1965; KLONTZ et al. , 1965 WOOD,1966). The immunological deficiency of fish, caused by their natural or experimental stay in cold water, is now evident for both humoral and cellular immunity. In this review we will focus on two points: We shall attempt (1) to explain the mechanism by which the environmental temperature influences the immune resistance of fish to pathogens, (2) to determine the chronological location of this temperature-sensitive stage in the process of antibody formation, and to make some approaches to the general antibody formation mechanism.

This volume is dedicated to the memory of the late Professor WERNER BRAUN, one of the most devoted and active members of the Editorial Board of the Current Topics in Microbiology and Immunology, who passed away, after suffering a heart attack, in November 1972. Dr. WERNER BRAUN was born in Berlin, Germany, on November 16,1914. During his highschool days in Berlin he did research work on problems of genetics as a young guest in the Kaiser-Wilhelm-Institut fur Biologie, in the department of Prof. R. GOLDSCHMIDT. I remember his colourful description of his discussions during this period, while still a teen-ager, with OTTO WAR­ BURG. He studied biology and medicine at the University of G6ttingen and received a Ph. D. degree in biology in 1936. In the same year he left Nazi Germany and came to the United States first as a Guest Investigator in Genetics at the University of Michigan at Ann Arbor, and then in Berkeley, where he carried out his work in the Depart­ ments of Zoology and of Veterinary Science until 1948. He was engaged during this period in the study of problems concerned with physiological genetics, bacterial variation, immunology and biochemistry.

Expression of an immune response is the net result of complex synergis­ tic and antagonistic activities performed by a variety of cell types. It includes macrophages, T and B populations which may interact in performance of a response, and suppressor cells interfering with it. Accordingly, a lack of res­ ponse may not necessarily indicate absence of immunocompetent cells, but rather nonexpression of competence. Thus, one should consider two possible situations, which are by no means mutually exclusive, to account for immuno­ logic unresponsiveness: (a) one or more of the cell populations composing the synergistic unit is absent or immature, and (b) an antagonistic unit which interferes with the response is dominating. In view of this, an approach to development of immune reactivity necessitates parallel surveys of development of cells with the potential to perform, as well as of cells which can suppress the response. Classification of the various cell types has been based so far on their phenotypic properties (e. g. , membrane antigen markers, cell receptors, pro­ duction and secretion of immunoglobulins, etc. ). Genotypically, T and B cells may represent either separate, independent cell lines, or different stages of development within the same cell lineage.

1.1 Classification of Togaviruses The family, Togaviridae, is composed of the alphaviruses, the flaviviruses, rubella (a rubivirus), and the pestiviruses (Fenner, 1976). Of these four genera, two (the alpha- and flaviviruses) are transmitted by blood-sucking arthropods, specif­ ically mosquitoes and ticks. Among the togaviruses, extensive studies of defective interfering (DI) particles have so far been carried out only with Sindbis virus (SV) and Semliki Forest virus (SFV), both members of the alphavirus genus. Since these viruses are so similar, in most cases it will be assumed that what is true of one is also true of the other. 1.2 Definition of Defective Interfering (DI) Particles Defective interfering viral particles, as defined by Huang (1973), have the follow­ ing properties: (1) they are deletion mutants and therefore lack large amounts of the genetic material present in the standard virus; (2) they contain the same viral structural proteins as standard virus; (3) they are unable to replicate alone; however, they are replicated in cells co-infected with standard virions; and (4) at the same time as they require standard virus to replicate, they inhibit the replication of standard virus and hence are interfering.

Prominent progress in molecular biology was only made when it became possible to separate functionally distinct molecules by taking advantage of their biophysical properties. Likewise, the analysis of the functions of hetero­ geneous populations of immunocompetent cells, as to the functional properties of their various subpopulations, can not be done until these can be isolated in reasonably pure form by selective fractionation. During the last few years significant advances have been made in this field, and cells have been separated according to size, density or charge (MILLER et aI. , 1969; SHORTMAN, 1968; ANDERSSON, 1973 c), or by taking advantage of more specific surface markers to allow selective depletion or enrichment of a given subpopulation of cells (WIGZELL and ANDERSSON, 1971). Although separation techniques have been used in a variety of cellular systems, they have been particularly useful in the study of reticuloendothelial cells and primarily in the study of cells partici­ pating in the immune responses. Quite extensive reviews have been written which well cover the methods used for separation of cells and the results obtained with the various approaches (WIGZELL and ANDERSSON, 1971; SHORTMAN, 1972). To review this work is becoming a more and more voluminous task. As data rapidly accumulate, we will not try to make such a complete review.

The study of the genetic regulation of immune response to natural multidetermi­ nant immunogens was undertaken by the method of bidirectional selective breed­ ing of High or Low antibody responder lines of mice. Five Selections are described: Selection I, carried out for agglutinin responsiveness to sheep erythrocytes and pigeon erythrocytes alternated in each generation. Selection II, carried out for agglutinin responsiveness to sheep erythrocytes repeated in each generation. Selection III and Selection IV performed respectively for agglutinin response to flagellar or somatic antigens of Salmonella typhimurium and Salmonella oranienburg alternated in each generation. Selection V, performed for passive agglutinin response to bovine serum albumin and rabbit gamma globulin alternated in each generation. In each Selection the character investigated is polygenic. High and Low responder lines diverge progressively during the selective breeding. The maximal interline separation (selection limit) is reached in the 7th-16th generations. High and Low responder lines at selection limit are considered homozygous for the character submitted to se~ection. Their variance is therefore only due to environ­ mental effects. The difference in agglutinin titre between High and Low lines is 220-fold in Selection I, 103-fold in Selection II, 90-fold in Selection III, 85-fold in Selection IV and 275-fold in Selection V. The partition of genetic and environmental variances in the foundation popu­ lations of the five Selections is established. The proportion of genetic variance is 60% in Selection I; 49% in Selection II; 51% in Selection III; 47% in Selection IV and 76% in Selection V.

The processes involved in herpesvirus replication, latency, and oncogenic transformation, have, in general, been rather poorly defined. A primary reason for this is the size and complexity of the herpesvirus genome. Undoubtedly, a better understanding of the functions of the viral genome in infected and transformed cells will be achieved through studies with temperature-sensitive (ts) mutants of herpesviruses since, theoretically, any essential gene function can be affected by mutants of this type. A. The Herpesviruses A consideration of the genetic analysis of members of the herpesvirus group necessitates a description, albeit brief, of the properties of the group and, most importantly, of their genetic material. The herpesviruses comprise a group of relatively large (100-150 nm), enveloped viruses. The envelope surrounds an icosahedral capsid enclosing a core which contains double­ stranded DNA (ROIZMAN, 1969). The group is thus defined on the basis of a common virion morphology. In addition to a common structure, members of the group share a number of biological properties such as a similar replicative cycle, the ability to cause latent and chronic infections, and the ability to induce antigenic modifications of infected cell membranes. Several herpes­ viruses have been associated recently with malignancies in man and animals (KLEIN, 1972). Herpesviruses are ubiquitous and have been described in over 30 different species (HUNT and MELENDEZ, 1969; WILDY, 1971; FARLEY et aI. , 1972; KAZAMA and SCHORNSTEIN, 1972; NAHMIAS et aI. , 1972; ROlZMAN et aI. , 1973). Their widespread occurrence in nature suggests a common ancestor.