Molecular Genetics

molecular genetics

[mə′lek·yə·lər jə′ned·iks] (cell and molecular biology) The approach which deals with the physics and chemistry of the processes of inheritance.

Molecular Genetics

a branch of genetics and molecular biology concerned with learning the material bases of heredity and variation in living things by investigating on the subcellular molecular level the processes of the transmission, materialization, and alteration of genetic information and the methods of storing that information.

Molecular genetics became an independent discipline in the 1940’s as a result of the application of new physical and chemical methods to biology (X-ray diffraction analysis, chromatography, electrophoresis, high-speed centrifugation, electron microscopy, the use of radioactive isotopes). These methods made possible a deeper and more accurate study of the structures and functions of individual cell components and of the entire cell as a unified system. In addition, new ideas from chemistry, physics, mathematics, and cybernetics were introduced into biology. Molecular genetics to a large extent owes its rapid development to the transfer of the focus of genetic research from higher organisms (eucaryotes)—the principal subjects of classical genetics—to lower organisms (procaryotes)—bacteria, viruses, and many other microorganisms. The advantages of using simpler forms of life to solve genetic problems consist in the rapid succession of generations in these forms and the possibility of studying numerous individuals simultaneously; this leads to an increase in the resolving power and accuracy of genetic analysis. In addition, the relative simplicity of organization of bacteria, especially of viruses, facilitates elucidation of the molecular nature of genetic phenomena. The opinion sometimes expressed that molecular genetics and the genetics of microorganisms are one and the same is erroneous. Molecular genetics studies the molecular bases of genetic processes in both lower and higher organisms and does not include the specific genetics of procaryotes, which occupies a prominent place in the genetics of microorganisms.

During its short history, molecular genetics has made great strides, deepening and broadening our knowledge of the nature of heredity and variation; it has become the leading and most rapidly developing branch of genetics.

One of the main achievements of molecular genetics is the elucidation of the chemical nature of the gene. Classical genetics established that all hereditary potentials of organisms (their genetic information) are determined by discrete units of heredity called genes, which are located mainly in the chromosomes of the cell nucleus and in some organelles of the cytoplasm (plastids, mitochondria). However, the methods of classical genetics were unable to elucidate the chemical nature of the genes, which was noted as far back as 1928 by the outstanding Soviet biologist N. K. Kol’tsov, who substantiated the necessity of studying the mechanism of heredity on the molecular level. The first success in this area was achieved with the study of genetic transformation in bacteria. In 1944 the American scientist O. T. Avery and his associates discovered that hereditary characteristics of one type of pneumococcus could be transmitted to another, genetically different type by introducing into its cells the deoxyribonucleic acid (DNA) obtained from the first type. Subsequently, a similar genetic transformation by means of DNA was accomplished in other bacteria and recently in some multicellular organisms (flowering plants and insects).

Thus, it was shown that the genes consist of DNA. This conclusion was confirmed by experiments with DNA-containing viruses: it is sufficient to inject molecules of viral DNA into the cell of a susceptible host to cause the virus to reproduce; all the other components of the virus (proteins, lipides) lack infectious properties and are genetically inert. Similar experiments with viruses containing ribonucleic acid (RNA) instead of DNA have shown that the genes in these viruses consist of RNA. Clarification of the genetic roles of DNA and RNA served as a powerful stimulus to the study of nucleic acids by biochemical, physico-chemical, and X-ray diffraction methods.

In 1953 the American scientist J. Watson and the British scientist F. Crick proposed a model of the structure of DNA, hypothesizing that its gigantic molecules consist of a double helix made up of a pair of strands formed by nucleotides, arranged aperiodically but in a definite sequence. Each nucleotide of one strand is paired with an oppositely situated nucleotide of the other strand according to the rule of complementarity. Numerous experimental data have confirmed the Watson-Crick model. Somewhat later it was established that the molecules of various RNAs have an analogous structure but that they consist for the most part of a single polynucleotide strand. Later research, in which chemical and physicochemical methods were combined with precise genetic methods (for example, the use of various mutants and the phenomena of transduction and transformation) showed that different genes differ in the number of nucleotide pairs (from several dozens to 1,500 or more), as well as in the sequence of nucleotides, which is strictly determined for each gene and in which the genetic information is encoded. Genes consisting of RNA—in viruses of the RNA-type—have a fundamentally similar structure.

Classical genetics regarded the gene as a discrete and indivisible unit of heredity. The works of A. S. Serebrovskii and his students in the 1930’s, which first suggested the possibility of the divisibility of the gene, were of great significance in the reexamination of that concept. However, the resolving power of the methods of classical genetics was inadequate for the study of the fine structure of the gene. It was only with the development of molecular genetics in the 1950’s and 1960’s that it became possible to solve this problem. Through many studies, first conducted on bacteria and viruses and then on multicellular organisms, it became clear that the gene has a complex structure: it consists of tens or hundreds of sections—sites—which are capable of mutating and recombining independently. The limit of divisibility of a gene, and consequently the minimal size of a site, is one pair of nucleotides (in viruses containing one RNA strand, one nucleotide). Determination of the fine structure of genes has made possible a deeper insight into the mechanism of genetic recombination and the principles of the origin of gene mutations; it has also promoted elucidation of the mechanism of gene function.

Data on the chemical nature and fine structure of genes have made it possible to develop methods of isolating them. This was first done in 1969 by the American scientist J. Beckwith and his associates for one of the genes of Escherichia coli. Subsequently, the same was successfully accomplished in some higher organisms (amphibians). An even more significant achievement of molecular genetics was the first chemical synthesis of a gene (the one that encodes the alanine transfer RNA of yeasts), accomplished by H. Khorana in 1968. Studies of this kind are being conducted throughout the world. The latest biochemical methods, based on the phenomenon of reverse transcription( see below), have been successfully used for the extracellular synthesis of larger genes. Using these methods, S. Spiegelman, D. Baltimore, P. Leder, and their associates (USA) have made great progress in artificially synthesizing the genes that determine protein structure in hemoglobin molecules of rabbits and humans. Similar studies have recently been conducted elsewhere, including the USSR.

Thus, molecular genetics has already explained, in theory, how genetic information received by offspring from parents is recorded and stored, although much work is still required to decipher the detailed content of that information for each individual gene.

Determination of the DNA structure has paved the way for experimental investigation of the biosynthesis of DNA molecules —that is, their replication. The process of DNA replication is the basis for the transfer of genetic information from cell to cell and from generation to generation—that is, it determines the relative constancy of genes. Study of DNA replication has led to the important conclusion of the template nature of DNA biosynthesis: in order for biosynthesis to take place, the presence of a completed DNA molecule is necessary, upon which, as on a template, the new DNA molecules are synthesized. In this process, the double helix of DNA unwinds, and on each of its strands a new, complementary strand is synthesized; as a result, the daughter DNA molecules consist of one old and one new strand (semiconservative replication). The protein that induces unwinding of the double helix of DNA and the enzymes that carry out the biosynthesis of nucleotides and their linkage have been identified. Undoubtedly, there are mechanisms in the cell that regulate DNA synthesis. The means of such regulation are still largely unclear, but it is evident that regulation is largely determined by genetic factors.

Molecular genetics has also achieved outstanding success in solving that most important question, already formulated by classical genetics, of how the gene determines a character, or how the materialization of genetic information occurs. The “one gene—one enzyme” hypothesis, formulated in 1941 by G. Beadle and E. Tatum, served as the starting point. This hypothesis made possible the following restatement of the question: How do the genes—that is, actually the segments of the DNA molecule —determine the chemical structure and properties of the proteins that are specific to a given organism? Discovery of the chemical structure of DNA and protein made it possible to correlate these two types of biopolymers, which led to the concept of the genetic code, according to which the sequence of the four different nucleotides in DNA determines the sequence of the 20 different amino acids in the protein molecule. All the properties of the protein molecule depend on the sequence of its amino acids (its primary structure).

The principles on which the genetic code is based were deciphered in 1962 by F. Crick and his associates in genetic experiments with mutants of a single bacterial virus. It turned out that each trio of nucleotides in the DNA chain (triplet, codon) determines precisely which of the 20 amino acids will occupy a given place in the polypeptide chain of the protein to be synthesized —that is, each triplet encodes a specific amino acid. Subsequent studies completely elucidated the genetic code and established the nucleotide composition of all the triplets that encode amino acids. Scientists have also established the composition of the initiating codon that determines the start of synthesis of a given polypeptide chain and of the three termination codons that end synthesis. The genetic code is universal for all living things—that is, it is identical for all organisms, from viruses to higher animals and man. The segment of the DNA molecule that constitutes one gene determines, as a rule, the amino-acid sequence in the molecule of a single protein (or in one polypeptide chain if the given protein consists of several such chains).

The decipherment of the genetic code was instrumental in elucidating the mechanism of protein biosynthesis, a process that includes the transfer of genetic information contained in the DNA to the molecules of messenger RNA (mRNA). This process, the essence of which is the synthesis of mRNA on the DNA template, is called transcription. Messenger RNA then becomes associated with special cell structures called ribosomes, on which the polypeptide chain is synthesized in accordance with the information contained in the mRNA molecule. This process of the synthesis of polypeptide chains through the mediation of mRNA is called translation.

Thus, the transmission of genetic information occurs according to the scheme DNA → RNA →; protein. This basic assumption (dogma), whose correctness has been established by many experiments with various organisms, was significantly amplified in 1970. The American scientists H. Temin and D. Baltimore discovered that in the reproduction of certain RNA-containing viruses that cause tumors in animals genetic information is transmitted from the RNA of the virus to the DNA. A similar reverse transcription is carried out by special enzymes contained in these viruses. The phenomenon of reverse transcription has also been discovered in some healthy animal and human cells. It is presumed that reverse transcription plays an important role in the origin of some forms of malignant tumors and in leukemias and possibly also in the processes of differentiation during the normal development of organisms. It must be emphasized that the discovery of reverse transcription does not contradict the basic assumption of molecular genetics that genetic information is transmitted from nucleic acids to proteins but cannot be transmitted from proteins to nucleic acids.

A remarkable achievement of molecular genetics is the discovery of the genetic mechanisms of regulating protein synthesis in the bacterial cell. As was shown in 1961 by the French scientists F. Jacob and J. Monod, the biosynthesis of protein in bacteria is under dual genetic control. On the one hand, the molecular structure of each protein is determined by a corresponding structural gene; on the other hand, the possibility of the synthesis of that protein is determined by a special regulator gene, which encodes a special regulatory protein that is capable of combining with a specific segment of DNA, the operator gene, during which time the regulatory protein can “turn on” or “turn off’ the functioning of the structural genes directed by the operator gene. A system of one or more structural genes and their operator gene constitutes an operon.

The ability of regulatory proteins to combine with an operator gene depends on compounds of low molecular weight, called effectors, which interact with these proteins. Effectors enter the cell from outside or are synthesized by the cell and signal the necessity for the synthesis by that cell of certain proteins or the cessation of synthesis. There are two types of regulatory proteins: repressor proteins, which, upon combining with an operator gene, block protein synthesis (negative regulation) and activator proteins, which, upon combining with an operator gene, induce protein synthesis (positive regulation). With negative regulation, the repressor in some cases is found in an active form before interacting with the effector and, upon combining with the operator gene, obstructs transcription of the structural genes of the operon (and consequently also synthesis of the corresponding proteins). The effector converts the repressor to an inactive form, the operator gene is freed, and the transcription of structural genes (and hence also the synthesis of the proteins encoded by these genes) becomes possible. In other cases, the interaction of repressor and effector converts the repressor to its active form, in which it is capable of combining with the operator gene, which leads to blocking of protein synthesis. With positive regulation, only the active form of the activator protein, which is capable of combining with the operator gene, determines protein synthesis. The active form of the activator protein is also determined by the activator’s interaction with the effector.

In multicellular organisms, the genetic regulation of protein synthesis is more complex and has not been sufficiently studied. However, it is clear that here, too, an important role is played by feedback similar to that described in bacteria for the system effector—regulator protein—operator gene; moreover, in a number of cases, hormones serve as the signaling substances.

With the development of molecular genetics, our understanding of mutation—that is, of change in genetic information—has expanded. It has been shown that mutations consist either of substitutions of certain nucleotides or of insertions or deletions of nucleotides in the DNA molecule. Mutations arise as a result of accidental errors in DNA replication and the effects of mutagens, various physical and chemical agents that damage nucleic acids. They are also caused by changes in mutator genes, which encode enzymes that participate in replication, correct genetic injuries, and so on. Alterations in the chemical structure of DNA caused by mutagens represent either direct mutations or lead to errors in the course of subsequent DNA replication, which, in turn, result in mutations. A significant portion of molecular injuries to DNA caused by mutagens does not give rise to mutations but is corrected (repaired). The essence of the phenomenon of repair lies in the fact that all organisms have genes that encode special enzymes capable of “recognizing” injured segments of DNA,“cutting them out” of the molecule, and substituting undamaged ones. Some of these enzymes have been identified and the mechanisms of their action established, but the repair process is not yet completely understood.

The study of repair has fostered new approaches to investigating the recombination mechanism of linked genes (that is, genes lying on one chromosome), which is one of the causes of combinatory variation; the latter, along with mutations, plays an important role in evolution. Classical genetics showed that the recombination of linked genes occurs by an exchange of segments between homologous chromosomes (crossing-over), but the detailed mechanism of such an exchange remained unknown. Experimental data of the past ten to 15 years have made it possible to view intrachromosomal and intragenal (intersite) recombination as an enzymatic process that occurs when DNA molecules interact. Recombination occurs through breaks of pieces of polynucleotide strands and their reattachment in new combinations. In this case, breaks with subsequent reattachments may occur simultaneously in the two DNA strands (crossing-over) or within only one strand (half crossing-over). Cross-over and repair require the synthesis of injured segments and restoration of disrupted phosphate bonds. These are effected by the appropriate enzymes.

Molecular genetics has influenced all the biological sciences through its remarkable discoveries. It provided the basis for the development of molecular biology; it significantly accelerated progress in biochemistry, biophysics, cytology, microbiology, virology, and developmental biology; and it revealed new approaches to understanding the origin of life and the evolution of the organic world. At the same time, molecular genetics, which has made possible a fuller understanding of the nature of the most important life processes and which is successfully continuing to investigate them, makes no claim whatever that it can solve many problems, including genetic ones, that concern the entire organism much less those that concern aggregates of organisms—populations, species, biocoenoses, and so on—where principles prevail that require methods of study other than those used by molecular genetics.

The achievements of molecular genetics, which have made an enormous theoretical contribution to general biology, will undoubtedly be widely used in agriculture and medicine. (Examples include genetic engineering by replacement of harmful genes with beneficial ones, including artificially synthesized ones; management of the mutation process; control of viral diseases and malignant tumors by intervening in the processes of replication of nucleic acids and oncogenic viruses; and direction of the development of organisms by acting on the genetic mechanisms of protein synthesis.) Successes attained on test subjects confirm future practical applications of the achievements of molecular genetics. Thus, in genetically well-studied species of bacteria, it is possible to obtain a mutation of any gene, to deprive a cell of any gene, to introduce any gene into a cell, and to regulate the functions of many genes.

Although the genetic properties of the cells of eucaryotes have been insufficiently studied on the molecular level, the first attempts to introduce certain genes into mammalian cells by means of viruses have been successful, and hybridization of somatic cells has been accomplished. For example, in 1971 the American scientist C. Merril and his associates cultivated in vitro the cells of a man suffering from galactosemia (such cells are incapable of producing one of the enzymes necessary for the utilization of lactose, which is the cause of this serious hereditary disease) and introduced into those cells a bacterial virus that was noninfectious to them and contained the gene that encodes the enzyme in question. As a result, the cells were “cured” and began synthesizing the missing enzyme and transmitting that ability to succeeding cell generations. The data of molecular genetics are already being used to produce medications for the prevention and treatment of neoplasms, leukemias, viral infections, and radiation injuries; they are also used in the search for new mutagens.

REFERENCES

Wagner, R., and H. Mitchell. Genetika i obmen veshchestv. Moscow, 1958. (Translated from English.)
Molekuliarnaia genetika, part 1. Moscow, 1964. (Collection of articles translated from English.)
Kol’tsov, N. K.“Nasledstvennye molekuly.” Biull. Moskovskogo ob-va ispytatelei prirody: Otdel biologicheskii, 1965, vol. 70, fasc. 4, pp. 75-104.
Bresler, S. E. Vvedenie v molekuliarnuiu biologiiu, 3rd ed. Moscow-Leningrad, 1973.
Watson, J. Molekuliarnaia biologiia gena. Moscow, 1967. (Translated from English.)
Herskowitz, I. Genetika. Moscow, 1968. (Translated from English.)
Khesin, R. B.“Enzimologiia geneticheskikh protsessov.” In Voprosy molekuliarnoi genetiki i genetiki mikroorganizmov. Moscow, 1968.
Ratner, V. A. Printsipy organizatsii i mekhanizmy molekuliarno-geneticheskikh protsessov. Novosibirsk, 1972.
Stent, G. S. Molecular Genetics. San Francisco, 1971.
Eigen, M. “Self-organization-of Matter and the Evolution of Biological Macromolecules.” Naturwissenschaften, 1971, vol. 58, fasc. 10.
Baltimore, D.“Viral RNA-dependent DNA Polymerase.” Nature, 1970, vol. 226, no. 5252.
Temin, H., and S. Mizutani.“RNA-dependent DNA Polymerase in Virions of Rous Sarcoma Virus.” Nature, 1970, vol. 226, no. 5252.
Kacian, D. L. et al.“In Vitro Synthesis of DNA Components of Human Genes for Globins.” Nature: New Biology, 1972, vol. 235, no. 58.

S. M. GERSHENZON and E. I. CHEREPENKO

molecular genetics