Eukaryotic Gen

EUKARIYOTIC GEN

Introduction

            Since 1953, when Crick, Watson and Wilkins provided insight into the double helical structure DNA, the steady snowfall of new knowledge about genes and gene structure has turned into a blizzard, a blizzard that all involved in genetics most endeavour to live with and work in. This book sets out to make life amidstthe rapid accumulation of new knowledge easier to mark a road through the snow. To do so is ambitious and beyond the skills of single individual. What we have therefore soughtto do has been to compile a book, with the chapters written by workers who can talk authoritatively in their field, that providing up to date information in a formal which reflects the comparative importance of different levels of information in terms of gene suquences, chromatin and chromosomes.

 

Figure 1. Picture Watson and Crick Explain DNA Model

            How can the present blizzard of the new information be accounted for? Essentially it result from the effeciency but comparative simplicity of the genetics code, consisting as it does of triplet codons of only four different bases and coding for little more than twenty different bits of translated informations. This coding simplicit, together with the coplementary nature of the squence on the two sister strands of the DNA helix, implies that if ways could be found to locate gene sequence with chromatin and determine base sequence within be isolated gene, then it would be possible to begin a study of genetics from precisely the opposite and to that which had occupied man’s attentions over the centuries. Instead of studying phenotype, familiar inhertance or at best, the expression of genes in their constituent proteins it has rather suddenly become possible to read out the genetic code itself and attempt to unscramble what is significant in the message so revealed. While it must be acknowledged that interpreting the facts about inhertance from knowledge of DNA squence is not by any means straightforward, and that many aspects of genetics will only be effectively understood by knowing that whole story from gene squence to phenotyphic character, yet it cannot be denied that the new approach has introduced a major revolution in the study of genetics and that our understanding of the processes of inhertance has made a considerable advance.

New technology has provided new insight

            It is wrothwhile to consider what the new techniques have been which have proved so valuable in genetics. Some of the technology is discussed in greater detail in the introduction to individual sections of this book, and to a lasser extent in some of the chapters themselves. But here it apropriate to consider the range of tchniques which have found application at many different levels in the genetics framework.

1.    At the level of the chromosome

Although the chromosomal location  of genes had been assumed since the turn of the century and known with certainty for a few decades, chromosomes have proved difficult to study. In most cells they were only visible during mitosis and many looked superficially similiar. Even the chromosomal location of large blocks of genes could not be readly ascertained as is revelead by the early ambiguity about the chromosomes involved in human mongolism. The great step forward came with chromosome banding. Following on from the application of fluorescence technique to the staining of chromosomes, it became apparent that particular cytochemical methods involving strong alkali or formamide would also specifically stain the centromeric chromatin. As such C banding became commonplaces; it also became clear that somewhat smiliar techniques, when followed by Giemsa staining, would reveal a banded pattern over the entire chromosome, called the G band pattern. What was particularly useful was that different chromosomes displayed different G band patterns , so permiting for the first time uniqeuivocal identification of particular chromosomes.

 

Figure 2. Chromosome 

Ascribing a gene to a particular chromosme or even a unique locus on a chromosomes remains difficult. Such assigment of genes has traditionally depend on linkage studies and measuremeant of cross-over frequencies where possible, but there are now staining techniques which in at least some circumstances, can reveal the location of spesific genes e.g the use of silver stain to localize active ribosomal RNA sequences. Genes which are repetitious and occur in large blocks such as the transfer RNA genes can be localized by autoradiography following in situ hybridization with radioactive cDNA. Some works has been carried out the location of unique structural gene sequences by autoradiography but the weak signal obtained renders the interpretation of such experiments with complementray cDNA extremely difficult. A quite different technology which can permit unequivocal assignment of a particular structural gene to an individual chromosome is that of cell fusion and the formation of bispesific hybrid cell. In particular by the exploitation of Sendai-virus medicated cell fusion between mouse and human cells, followed by the culture of the hybrids, often with of gradual loss of most of human chromosome set, cell lines have been obtained which the contain an’all-mouse’genome with the exception of a single human chromosome. Banding of the chromosome reveals the identity of the single human chromosome while a prolonged assay for spesific protein expression, say the timidyne kinase gene, permits unequivocal assignment of a particular structure gene to an individual chrosome.

 

Figure 3. DNA in Chromosome

            Useful as these advances in whole chromosome technology have been, they are certainly less startling than the developments in the fields of chromatinand actual gene sequence, and these latter areas are chiefly what this volume seeks to consider.

2.    At the level chromatin

Early theoretical attemps to understand the physical relationship betwwn histone and DNA formed an inner core and that the histone molecules, applied to the surface of the double helix could be regarded as the outher sheath. The true structure of chromatin was initially indicated by the simple but crucial experiments of Hewish and Burgoyne. These Australian workers digested rat liver chromatin with an endogeneous calcium active nuclease enzyme and produced the now familiar 200 base pair repeat ladder on electrophoresis of the DNA fragments from the digest. This result clearly implied that the DNA was in some way differentially protected from digestion, execpt for certain exposed sites located some 200 base pair apart. When this observation was put alongside the results of chromatin fine structure analysis in the electron miscroscope, the nation of histone bead as repeating structure in chromatin soon emerged. Nuclear magnetic resonance (n.m.r) studies soon esrabilished that, contrary to the earliest assumption about histone/DNA conformation, the DNA was wound on the outside of histone octamer. So was conceived what is now accepted as an accurate pictures of the subunit of most chromatin, the nucleosome. Some of the earlier chapters in this book provide an up to date view of nucleosome and how its structure is implicated with such other aspects of genetics as gene regulation and the arragement of chromatin chromosomes.

Its now clear the nucleosome are the invariable chromatin subunit in all transcriptionally inactive chromatin (expectsperm) and probably in most active chromatin as well. Arragement of the nucleosome in the higher oreder structure of chromatin is less clear but already exploitation of techniques such as nuclear magnetic resonance have provided persuasive evidence that nucleosome are arranged as helical solenoids, phraps each solenoid representing a loop or domain of DNA attached to backbone or scaffold in the intact chromosome. 

 

Figue 4. DNA in Chromatin

loading...