`
`Page 1
`
`KASHIV EXHIBIT 1012
`IPR2019-00797
`
`

`

`
`THE CELL
`
`MOLECULAR BIOLOGY
`
`fourth
`
`edition
`
`Bruce Alberts
`
`Alexanderjohnson
`
`JuHan Lewis
`
`Martin Raff
`
`Keith Roberts
`
`Peter Walter
`
`Garland Science
`
`Tayler&Francis Group
`
`Page 2
`
`Page 2
`
`

`

`
`
`.
`Garland
`Vice President: Denise Schanck
`Managing Editor: Sarah Gibbs
`Senior Editorial Assistant: Kirsten Jenner
`Managing Production Editor: Emma Hunt
`Proofreader and Layout: Emma Hunt
`Production Assistant: Angela Bennett
`Text Editors: Marjorie Singer Anderson and Betsy Dilernia
`Copy Editor: Bruce Goatly
`Word Processors: Fran Dependahl, Misty Landers and Carol Winter
`Designer: Blink Studio, London
`Illustrator: Nigel Orme
`Indexer: Janine Ross and Sherry Granum
`Manufacturing: Nigel Eyre and Marion Morrow
`
`Bruce Alberts received his Ph.D. from Harvard University and is
`President of the National Academy of Sciences and Professor of
`Biochemistry and Biophysics at the University of California, San
`Francisco. Alexander Johnson received his Ph.D. from Harvard
`University and is a Professor of Microbiology and Immunology
`and Co-Director of the Biochemistry and Molecular Biology
`Program at the University of California, San Francisco.
`Julian Lewis received his D.Phil. from the University of Oxford
`and is a Principal Scientist at the London Research Institute of
`Cancer Research UK. Martin Raff received his MD. from McGill
`University and is at the Medical Research Council Laboratory for
`Molecular Cell Biology and Cell Biology Unit and in the Biology
`Department at University College London. Keith Roberts received
`his Ph.D. from the University of Cambridge and is Associate
`Research Director at the John Innes Centre, Norwich. Peter Walter
`received his Ph.D. from The Rockefeller University in New York and
`is Professor and Chairman of the Department of Biochemistry and
`Biophysics at the University of California, San Francisco, and an
`Investigator of the Howard Hughes Medical Institute.
`
`© 2002 by Bruce Alberts, Alexander Johnson, Julian Lewis,
`Martin Raff, Keith Roberts, and Peter Walter.
`© 1983, 1989, 1994 by Bruce Alberts, Dennis Bray, Julian Lewis,
`Martin Raff, Keith Roberts, and James D. Watson.
`
`All rights reserved. No part of this book covered by the copyright
`hereon may be reproduced or used in any format in any form or
`by any means—graphic, electronic, or mechanical, including
`photocopying, recording, taping, or information storage and
`retrieval systems~with0ut permission of the publisher.
`
`Library of Congress Cataloging~in~Publication Data
`Molecular biology of the cell / Bruce Alberts
`[et al.].—— 4th ed.
`p. cm
`
`Includes bibliographical references and index.
`ISBN 0-8153-3218—1 (hardbound) -— ISBN 0-8153~4072~9 (pbk.)
`1. Cytology. 2. Molecular biology. I. Alberts, Bruce.
`[DNLM2 1. Cells. 2. Molecular Biology ]
`QH581.2 .M64 2002
`571.6—~dC21
`
`2001054471 CIP
`
`Published by Garland Science, a member of the Taylor & Francis Group,
`29 West 35th Street, New York, NY 10001—2299
`
`Printed in the United States of America
`
`15141312111098765432
`
`Page 3
`
`Cell Biology Interactive
`Artistic and Scientific Direction: Peter Walter
`Narrated by: Julie Theriot
`Production, Design, and Development: Mike Morales
`
`Front cover Human Genome: Reprinted by permission
`from Nature, International Human Genome Sequencing
`Consortium, 409:860—921, 2001 © Macmillan Magazines
`Ltd. Adapted from an image by Francis Collins, NHGRI;
`Jim Kent, UCSC; Ewan Birney, EBI; and Darryl Leja,
`NHGRI; showing a portion of Chromosome 1 from the
`initial sequencing of the human genome.
`
`Chapter opener Portion of chromosome 2 from the
`genome of the fruit fly Drosaphila melanagasrelz
`(Reprinted with permission from MD. Adams et al.,
`Science 287:2185~2195, 2000. © AAAS.)
`
`In 1967, the British artist Peter Blake
`Back cover
`created a design classic. Nearly 35 years later Nigel
`Orme (illustrator), Richard Denyer (photographer), and
`the authors have together produced an affectionate
`tribute to Mr Blake’s image. With its gallery of icons and
`influences, its assembly created almost as much
`complexity, intrigue and mystery as the original.
`Drosophila, Ambidapsis, Dolly and the assembled
`company tempt you to dip inside where, as in the
`original, "a splendid time is guaranteed for all.”
`(Gunter Blobel, courtesy of The Rockefeller University; Marie
`Curie, Keystone Press Agency Inc; Darwin bust, by permission
`of the President and Council of the Royal Society; Rosalind
`Franklin, courtesy of Cold Spring Harbor Laboratory Archives;
`Dorothy Hodgkin, © The Nobel Foundation, 1964; James Joyce,
`etching by Peter Blake; Robert Johnson, photo booth
`self—portrait early 19303, © 1986 Delta I-Iaze Corporation all
`rights reserved, used by permission; Albert L. Lehninger,
`(unidentified photographer) courtesy ofThe Alan Mason
`Chesney Medical Archives of The Johns Hopkins Medical
`Institutions; Linus Pauling, from Ava Helen and Linus Pauling
`Papers. Special Collections. Oregon State University; Nicholas
`Poussin, courtesy of ArtTodaycom; Barbara McClintock,
`© David Micklos, 1983; Andrei Sakharov, courtesy of Elena
`Bonner; Frederick Sanger, © The Nobel Foundation, 1958.)
`
`Page 3
`
`

`

`
`
`
`
`
`
`
`FROM DNATO RNA
`
`FROM RNA TO PROTEIN
`
`THE RNA WORLD AND THE
`
`ORIGINS OF LIFE
`
`
`-' T-fliflfilmlifllm Hillifinslmll'lililr'll'mmflmmflfl '
`
`HOW CELLS READ THE
`GENOME: FROM
`DNA TO PROTEIN
`
`Only when the structure of DNA was discovered in the early 19505 did it become
`clear how the hereditary information in cells is encoded in DNA’s sequence of
`nucleotides. The progress since then has been astounding. Fifty years later, we
`have complete genome sequences for many organisms, including humans, and
`we therefore know the maximum amount of information that is required to pro-
`duce a complex organism like ourselves. The limits on the hereditary informa-
`tion needed for life constrain the biochemical and structural features of cells
`and make it clear that biology is not infinitely complex.
`In this chapter, we explain how Cells decode and use the information in their
`genomes. We shall see that much has been learned abOut how the genetic
`instructions written in an alphabet of just four "letters”-—the four different
`nucleotides in DNA—direct the formation of a bacterium, a fruitfly. or a human.
`Nevertheless, we still have a great deal to discover about how the information
`stored in an organism's genome produces even the simplest unicellular bacterium
`with 500 genes,
`let alone how it directs the deVeloprnent of a human with
`approximately 30,000 genes. An enormous amount of ignorance remains: many
`fascinating challenges therefore await the next generation of cell biologists.
`The problems cells face in decoding genomes can be appreciated by consid»
`ering a small portion of the genome of the fruit fly Drosophila melanogaster (Fig-
`ure 5-1]. Much of the DNA-encoded information present in this and other
`ganomes is used to specify the linear order—the sequence—of amino acids for
`Wary protein the organism makes. As described in Chapter 3, the amino acid
`quumce in turn dictates how each protein folds to give a molecule with a dis-
`tinctive shape and chemistry. When a particular protein is made by the cell, the
`lioflesponding region of the genome must therefore be accurately decoded. Addi-
`tlonal information encoded in the DNA of the genome specifies exactly when in
`the life of an organism and in which cell types each gene is to be expressed into
`protein. Since proteins are the main constituents of cells, the decoding of the
`genome determines not only the size, shape, biochemical properties, and behav-
`ior of cells, but also the distinctive features of each species on Earth.
`One might have predicted that the information present in genomes would be
`amlllged in an orderly fashion, resembling a dictionary or a telephone directory.
`
`
`
`———————-——fl
`Page 4
`
`29?
`
`Page 4
`
`

`

`PM
`
`w'1'! III'IIIIIIIIMJ'|||IIIII'IIIHIIII iIIIIIIIE IIIII 'II I
`
`I.
`iIIIIIIIIIIIIIIIII I III III IIIIIIIIIII
`
`"Iil'iIi III IIIIII'III
`III 'III
`WI
`IIIIIIIIIIIII
`.IIIIIIIIII
`III
`: III I II.IIIII IIIIII
`
`
`
`color code for sequence 5:
`of genes identified
`
`I
`
`Y = S. cerevisiee
`
`13 or more
`
`"one
`
`I “I 12
`$312:J 23:11,:er
`of corresponding
`cDNAs identified
`in databases
`
`P :i/
`
`%Gc content [—1
`5
`
`2
`
`56
`
`:|— transposabie eiements
`
`|
`
`i
`
`I a
`I
`
`I
`
`II
`
`‘
`
`I
`
`III
`
`.
`
`E.
`
`I5
`
`i
`
`I
`
`I';
`
`'
`
`known and predicted genes
`
`identified on top strand of DNA
`Ignown and predicted genes
`Identlfled on bottom strand of DNA
`
`I
`
`-
`
`EII
`
`i_—_I
`100,000 nucleotide pairs
`
`t
`a
`=
`w = aeiegans
`
`300
`
`Chapter 6 : HOW CELLS READ THE GENOME: FROM DNA TO PROTEIN
`
`Page 5
`
`Page 5
`
`

`

`m
`
`I '
`
`
`
`Figure 6-! (opposite page) Schematic depiction of a portion oi chromosome 2 from the genome of
`the fruit fly Drosophilo melonogasterThls figure represents approximately 3% of the total Drosophilo genome.
`arranged as six contigUOUs segments.As summarized in the key, the symbollc representations are: rainbow-colored
`bor: G—C base»pair content: black vertical lines of various thicknesses: locations of transposable elements. with
`thicker bars indicating clusters of elements: colored boxes: genes (both known and predicted) coded on one strand
`of DNA (boxes above the midline} and genes coded on the other strand (boxes below the midline).The length of
`each predicted gene includes both its exons (protein-coding DNA) and its introns (non—coding DNA) (see Figure
`4-25).As indicated in die key. the height of each gene box is proportional to the number of cDNAs in various
`databases that match the geneAs described in Chapter 8. cDNAs are DNA copies of mRNA molecules. and
`large collections of the nucleoride sequences of cDNAs have been deposited in a variety of databases.The higher
`the number of matches between the nucleotide sequences of cDNAs and that of a particular predicted gene. the
`higher the confidence that the predicted gene is transcribed into RNA and is thus a genuine geneThe color of
`each gene box (see color code in the key) indicates whether a closely related gene is known to occur in other
`organisms. For example. HWY means the gene has close relatives in mammals. in the nematode worm
`Coenorhobditis elegans, and in the yeast Socchommyces cererisioe. MW indicates the gene has close relatives in
`mammals and the worm but not. in yeast. (From Mark D. Adams et al.. Science 237:2l 85—2 | 95. 2000. ©AAAS.)
`
`although the genomes of some bacteria seem fairly well organized, the genomes
`of most multiccllular organisms, such as our Drosophila example. are surpris-
`ingly disorderly. Small bits of coding DNA [that is, DNA that codes for protein}
`are interspersed with large blocks of seemingly meaningless DNA. Some sections
`of the genome contain many genes and others lack genes altogether. Proteins
`that work closely with one another in the cell often have their genes located on
`different chromosomes. and adjacent genes typically encode proteins that have
`little to do with each other in the cell. Decoding genomes is dierefore no simple
`matter. Even with the aid of powerful computers, it is still difficult for researchers
`to locate definitively the beginning and end of genes in the DNA sequences of
`complex genomes, much less to predict when each gene is expressed in the life
`of the organism. Although the DNA sequence of the human genome is known. it
`will probably take at least a decade for humans to identify every gene and deter-
`mine the precise arnino acid sequence of the protein it produces. Yet the cells in
`our body do this thousands of times a second.
`The DNA in genomes does not direct protein synthesis itself. but instead
`uses RNA as an intermediary molecule.When the cell needs a particular protein.
`the nucleotide sequence of the appropriate portion of the immensely long DNA
`molecule in a chromosome is first copied into RNA [a process called transcrip—
`tion). It is these RNA copies of segments of the DNA that are used directly as
`templates to direct the synthesis of the protein (a process called translation).
`The flow of genetic information in cells is therefore from DNA to RNA to protein
`(Figure 6-2). All cells. from bacteria to humans. express their genetic informa-
`tion in this way—a principle so fundamental that it is termed the central dogma
`ofmolecular biology.
`Despite the universality of. the central dogma. there are important variations
`in the way information flows from DNA to protein. Principal among these is that
`RNA transcripts in eucaryotic cells are subject to a series of processing steps in
`the nucleus. including RNA splicing, before they are permitted to exit from the
`nucleus and be translated into protein. These processing steps can critically
`change the “meaning" of an RNA molecule and are therefore crucial for under—
`standing how eucaryotic cells read the genome. Finally. although we focus on
`the production of the proteins encoded by the genome in this chapter. we see
`that for some genes RNA is the final product. Like proteins, many of th esc RNAs
`fold into precise three-dimensional structures that have structural and catalytic
`roles in the cell.
`We begin this chapter with the first step in decoding a genome: the process
`thranscription by which an RNA molecule is produced from the DNA of a gene.
`We then follow the fate of this RNA molecule through the cell. finishing when a
`Correctly folded protein molecule has been formed. At the end of the chapter. we
`consider how the present, quite complex. scheme of information storage. tran-
`scription. and translation might have arisen from simpler systems in the earliest
`Stages of cellular evolution.
`
`HOW CELLS READ THE GENOME; FROM DNA TO PROTEIN
`
`DNA replication
`DNA repair
`
`(geneticrecombination) DNA
`
`3'
`
`RNA synthesis
`liranscriptionl
`
`5’
`
`RNA
`
`AW?
`protein synthesis
`{translation}
`
`PROTEIN
`"
`'
`
`amino acids
`
`CDOH
`
`Figure 6-2 The pathway from DNA
`to protein. The flow of genetic
`information from DNA to RNA
`(tianscription) and from RNA to protein
`(translation) occurs in all living cells.
`
`30]
`
`
`Page 6
`
`Page 6
`
`

`

`Figure 6—3 Genes can be expressed
`with different efficiencies. GeneA is
`transcribed and translated much more
`efficiently than gene B.This allows the
`amount of proteinA in the cell to be
`much greater than that of protein B.
`
`gene 8
`gene A
`fl
`
`_— T
`
`4—“
`
`RANSLATION
`
`THANSLATlON
`
`lTFlANSCRIPTION
`
`— RNA — RNA
`
`JDNA
`l TRANSCRIPTION
`
`FROM DNA T0 RNA
`
`Transcription and translation are the means by which cells read out, or express,
`the genetic instructions in their genes. Because many identical RNA copies can
`be made from the same gene, and each RNA molecule can direct the synthesis
`of many identical protein molecules, cells can synthesize a large amount of
`protein rapidly when necessary. But each gene can also be transcribed and
`translated with a different efficiency, allowing the cell to make vast quantities of
`some proteins and tiny quantities of others (Figure 6~3). Moreover, as we see in
`the next chapter. a cell can change [or regulate} the expression of each of its
`genes according to the needs of the moment-mmosl' obviously by controlling
`the production of its RNA.
`-
`
`Portions of DNA Sequence Are Transcribed into RNA
`
`The first step a cell takes in reading out a needed part of its genetic instructions
`is to copy a particular portion of its DNA nucleotide sequence—a gene—into an
`RNA nucleotide sequence. The information in. RNA. although copied into another
`chemical form. is still written in essentially the same language as it is in DNA—
`the language of a nucleotide sequence. Hence the name transcription.
`Like DNA, RNA is a linear polymer made of four different types of nucleotide
`subunits linked together by phosphodiester bonds {Figure 6-4]. It differs from
`DNA chemically in two respects:
`[1]
`the nucleotides
`in RNA are
`ribonucleotides—that is, they contain the sugar ribose [hence the name ribonu-
`cleic acid) rather than deorryribose; (2] although, like DNA, RNA contains the
`bases adenine (A), guanine (G), and cytosine (C), it contains the base uracil [U]
`instead of the thymine [T] in DNA, Since U, like T, can base-pair by hydrogen-
`bonding with A [Figure 6—5],
`the complementary base—pairing properties
`described for DNA in Chapters 4 and 5 apply also to RNA (in RNA. G pairs with
`C, and A pairs with U]. It is not uncommon, however, to find other types of base
`pairs in RNA: for example, G pairing with U occasionally.
`Despite these small chemical differences, DNA and RNA differ quite dra-
`matically in overall structure. Whereas DNA always occurs in cells as a double-
`stranded helix, RNA is single-stranded. RNA chains therefore fold up into a
`variety of shapes, just as a polypeptide chain folds up to form the final shape of
`a protein (Figure 6—6). As we see later in this chapter, the ability to fold into com-
`plex three-dimensional shapes allows some RNA molecules to have structural
`and catalytic functions.
`
`Transcription Produces RNA Complementary to
`One Strand of DNA
`
`Ali of the RNA in a cell is made by DNA transcription, a process that has cer-
`tain similarities to the process of DNA replication discussed in Chapter 5.
`
`302
`
`Chapter 5 : HOW CELLS READ THE GENOME: FROM DNATO PROTEIN
`
`
`Page 7
`
`Page 7
`
`

`

`HOCH2 0
`
`H
`
`
`
`OH
`
`HOCI-i2 0
`
`OH
`
`H mH
`
`OH OH
`
`OH H
`
`{A}
`
`{3;
`
`
`
`3r
`
`5|
`
` N
`
`adenine
`C \ H
`
`EEPM
`used in ribonucleic
`acid {RNA}
`
`3%
`used in deoxyribonucleic
`acid lDNAl
`
`O
`t
`
`HC/
`Hé'
`
`“NH
`d
`\N/ {*0
`|
`H
`
`.ur'atill
`Used in RNA
`
`..
`3 \
`
`O
`l
`C/ ~“NH
`Hc”
`é
`\N/ KC)
`l
`H
`
`rhyming.
`used in DNA
`
`Figure 6-4 The chemical structure of RNA. (A) RNA contains the
`sugar ribose. which differs from deoxyribose.the sugar used in DNA. by the
`presence of an additional —OH group. (3) RNA contains the base uracil.
`which differs from thymine. the equivalent base in DNA, by the absence of 3
`‘CH3 group. {C}A short length of RNA.The phosphodiester chemical
`linkage between nucleotides in RNA is the same as that in DNA.
`
`
`
`Transcription begins with the opening and unwinding of a small portion of the
`DNA double helix to expose the bases on each DNA strand. One of the two
`strands of the DNA double helix then acts as a template for the synthesis of an
`
`RNA molecule. As in DNA replication. the nucleotide sequence of the RNA chain
`
`is determined by the complementary base-pairing between incoming
`
`nucleotides and the DNA template. When a good match is made. the incoming
`
`ribonucieotide is covalently linked to the growing RNA chain in an enzymati-
`
`cally catalyzed reaction. The RNA chain produced by transcription—the tron-
`
`script—is therefore elongated one nucleotide at a time. and it has a nucleotide
`
`sequence that is exactly complementary to the strand of DNA used as the tem-
`
`plate (Figure 6—7].
`
`Transcription. however, differs from DNA replication in several crucial ways.
`
`Unlike a newly formed DNA strand, the RNA strand does not remain hydrogen-
`
`bonded to the DNA template strand. Instead, just behind the region where the
`
`ribonucleotides are being added, the RNA chain is displaced and the DNA helix
`
`re—forms. Thus, the RNA molecules produced by transcription are released from
`
`the DNA template as single strands. In addition. because they are copied from
`
`only a limited region of the DNA, RNA molecules are much shorter than DNA
`
`molecules. A DNA molecule in a human chromosome can be up to 250 million
`
`nucleotide-pairs long; in contrast, most RNAs are no more than a few thousand
`
`nucleotides long. and many are considerably shorter.
`
`The enzymes that perform transcription are called RNA polymerases. Like
`
`the DNA polymerase that catalyzes DNA replication (discussed in Chapter 5).
`
`RNA polymerases catalyze the formation of the phosphodiester bonds that link
`
`the nucleotides together to form a linear chain. The RNA polymerase moves
`
`Stepwise along the DNA, unwinding the DNA helix just ahead of the active site
`
`for polymerization to expose a new region of the template strand for comple-
`
`mentary base-pairing. In this way, the growing RNA chain is extended by one
`
`nucleotide at a time in the 5’-to-3’ direction [Figure 6—8}. The substrates are
`
`nucleoside triphosphates (ATP. CTP, UTP. and GTP]; as for DNA replication. a
`
`hydrolysis of high-energy bonds provides the energy needed to drive the reac-
`
`tion forward (see Figure 5—4}.
`The almost immediate release of the RNA strand from the DNA as it is syn
`thesized means that many RNA copies can be made from the same gene in a
`
`
`
`Figure 6-5 Uracil forms base pairs with adenine. The absence of a
`methyl group in U has no effect on basespairing: thus. U—A base pairs closely
`resemble T—A base pairs (see Figure 4—4].
`
`5’
`
`sugar—phosphate backbone
`
`3‘
`
`393
`
`FROM DNATO RNA
`
`
`
`Page 8
`————_
`
`Page 8
`
`

`

`
`
`Figure 6-6 RNA can fold into specific structures. RNA is largely single-stranded. but it often contains short
`stretches of nucleotides that can form conventional base-palrs with complementary sequences found elsewhere
`on the same molecule. These interactions. along with additional “nonconventional” base-pair interactions,allow an
`RNA molecule to fold into a three-dimensional structure that is determined by its sequence of nucleotides.
`(A) Diagram of a folded RNA structure showing only conventional base-pair interactions: (B) structure with both
`conventional (red) and nonconventional {green} base-pair interactions: {C} structure of an actual RNA. a portion of
`a group I
`intron (see Figure 6—36). Each conventional base-pair interaction is indicated by a"rung" in the double
`helix. Bases in other configurations are indicated by broken rungs.
`
`relatively short time, the synthesis of additional RNA molecules being started
`before the first RNA is completed [Figure 6—9}. When RNA polymerase molecules
`follow hard on each other’s heels in this way. each moving at about 20
`nucleotides per second {the speed in eucaryotes}, over a thousand transcripts
`can be synthesized in an hour from a single gene.
`Although RNA polymerase catalyzes essentially the same chemical reaction
`as DNA polymerase, there are some important differences between the two
`enzymes. First, and most obvious, RNA polymerase catalyzes the linkage of
`ribonucleotides, not deoxyribonucleotides. Second, unlike the DNA polymerases
`involved in DNA replication, RNA polymerases can start an RNA chain without
`a primer. This difference may exist because transcription need not be as accu-
`rate as DNA replication [see Table 5~1. p. 243]. Unlike DNA, RNA does not per—
`manently store genetic information in cells. RNA polymerases make about one
`mistake for every 10‘1 nucleotides copied into RNA [compared with an error rate
`for direct copying by DNA polymerase of about one in 107' nucleotides), and the
`consequences of an error in RNA transcription are much less significant than
`that in DNA replication.
`Although RNA polymerases are not nearly as accurate as the DNA poly-
`merases that replicate DNA, they nonetheless have a modest proofreading
`mechanism. If the incorrect ribonucleotide is added to the growing RNA chain.
`the polymerase can back 11 p. and the active site of the enzyme can perform an
`excision reaction that mimics the reverse of the polymerization reaction, except
`that water instead of pyrophosphate is used (see Figure 5—4}. RNA polymerase
`hovers around a misincorporated ribonucleotide longer than it does for a cor—
`
`rect addition, causing excision to be favored for incorrect nucleotides. However,
`
`RNA polymerase also excises many correct bases as part of the cost for improved
`accuracy.
`
`Cells Produce Several Types of RNA
`The majority of genes carried in a cell's DNA specify the amino acid sequence of
`proteins; the RNA molecules that are copied from these genes [which ultimately
`direct the synthesis of proteins) are called messenger RNA (mRNAJ molecules.
`
`304
`
`Chapter 6 :HOW CELLS READ THE GENOME: FROM DNA TO PROTEIN
`
`
`lmmscmpno“
`
`
`template strand
`
`RNA
`Figure H, DNAtmnsmpfion
`Produces a Single-Stranded RNA
`mOIBwle that is comP'ementarY
`t° ""8 5'th °f DNA-
`
`Page 9
`
`Page 9
`
`

`

`
`
`flap in
`closed
`position
`
`.
`.
`
`rrbonuclootide
`_
`_
`active site
`triphosphate
`5’ channel
`tunnel
`
`newly synthesized
`RNA transcript
`
`short region of
`DNNRNA helix
`
`
`
`
`RNA polymerase
`
`iaws in closed
`configuration
`
`DNA double
`helix
`
`DNA
`.
`
`rewinding
`
`
`
`ribonuciootida
`
`if triphosphetes
`
`direction of
`transcription
`
`
`
`Figure 6-8 DNA is transcribed by
`the enzyme RNA polymerase. The
`RNA polymerase (pale blue) moves
`stepwise along the DNA. unwinding the
`DNA helix at its active site. As ll:
`
`progresses. the polymerase adds
`nucleotides (here.srnali "T"5ltapesl one by
`one to the RNA chain at the
`
`polymerization site using an exposed
`DNA strand as a template.The RNA
`transcript is thus a single-stranded
`complementary copy of one of the twu
`DNA strandsThe polymerase has a
`rudder (see Figure 6-i I) that displaces
`the newly Formed RNA. allowing the two
`strands of DNA behind the polymerase to
`rewind.A short region of DNAIRNA helix
`(approximately nine nucleotides in length)
`is therefore formed only transiently. and a
`"window" of DNAFRNA helix therefore
`
`moves along the DNA with the
`polymeraseThe incoming nucleotides are
`In the form of ribonucleoside
`
`criphosphates (ATP. UTF'. CTR and GTP).
`and the energy stored in their
`phosphate-phosphate bonds provides the
`driving force for the polymerization
`reaction {see Figure 5-4). {Adapted from a
`figure kindly supplied by Robert Landick.)
`
`305
`
`The final product of a minority of genes, however. is the RNA itself. Careful
`analysis of the complete DNA sequence of the genome of the yeast S. cereyisine
`
`has uncovered well over 750 genes [somewhat more than 10% of the total num-
`
`ber of yeast genes] that produce RNA as their final product, although this num-
`
`ber includes multiple copies of some highly repeated genes. These RNAs, like
`
`proteins. serve as enzymatic and structural comporrents for a wide variety of
`
`processes in the cell. In Chapter 5 we encountered one of those RNAs. the tem—
`
`plate carried by the enzyme telomerase. Although not all of their functions are
`
`known. we see in this chapter that some small nuclear-RNA (snRNA) molecules
`
`direct the splicing of pre-mRNA to form mRNA, that ribosomni RNA {rRNA}
`
`molecules form the core of ribosomes. and that transfer RNA (IRNA) molecules
`
`form the adaptors that select amino acids and hold them in place on a ribosome
`
`for incorporation into protein [Table 6—1].
`
`Each transcribed segment of DNA is called a transcription unit. In eucary-
`ores. a transcription unit typically carries the information of just one gene. and
`therefore codes for either a single RNA molecule or a single protein {or group of
`
`related proteins if the initial RNA transcript is spliced in more than one way to
`
`produce different mRNAs]. In bacteria. a set of adjacent genes is often trans-
`
`cribed as a unit; the resulting rnRNA molecule therefore carries the information
`
`for several distinct proteins.
`
`Overall, RNA makes Up a few percent of a cell’s dry weight. Most of the RNA
`
`in cells is rRNA: mRNA comprises only 3—5% of the total RNA in a typical mam-
`
`malian cell. The mRNA population is made up of tens of thousands of different
`
`species, and there are on average only 10—15 molecules of each species of mRNA
`
`present in each cell.
`E
`"ill...
`iii-ll
`
`
`
`
`
`
`
`3’
`
`
`
`Figure 6-9 Transcription of two genes as observed under the
`electron microscope. The micrograph Show rnany molecules of RNA
`polymerase simultaneously transcribing each of two adjacent genes.
`Molemles of RNA polymerase are visible as a series of dots along the DNA
`with the newly synthesized transcripts (fine threads} attached to them.The
`RNA molecules (ribosomal RNAs} shown in this example are not translated
`into protein bur: are instead used directly as components of ribosomes. the
`machines on which translation.taices place.The particles at the 5’ end (the
`free end) of each rRNA transcript are believed to reflect the beginnings of
`ribosome assembly. From the lengths of the newly synthesized transcripts. It
`can be deduced that the RNA polymerase molecules are transcribing from
`left to right. {Courtesy of Ulrich Scheer.)
`
`FROM DNA T0 RNA
`
`
`
`Page 10
`___————-——
`
`Page 10
`
`

`

`#—
`
`TABLE 6—! Principal Types of RNAs Produced in CellsH
`
` TYPE OF RNA
`FUNCTlON
`
`mRNAs
`rRNAs
`
`tRNAs
`
`snRNAs
`
`snoRNAs
`
`messenger RNAs. code for proteins
`rihosomal RNAs. form the basic structure of the
`ribosome and catalyze protein synthesis
`transfer RNAs, central to protein synthesis as adaptors
`between mRNA and amino acids
`
`small nuclear RNAs. function in a variety of nuclear
`processes, including the splicing of pre—mRNA
`small nucleolar RNAs, used to process and chemically
`modify rRNAs
`
`function in diverse cellular processes, including
`Other noncoding
`telornere synthesis, X-chromosorne inactivation,
`RNAs
`and the transport of proteins into the ERfl
`
`Signals Encoded in DNA Tell RNA Polymerase Where
`to Start and Stop
`
`To transcribe a gene accurately, RNA polymerase must recognize where on the
`genome to start and where to finish. The way in which RNA polymerases per—
`form these tasks differs somewhat between bacteria and eucaryotes. Because
`
`the process in bacteria is simpler, we look there first.
`The initiation of transcription is an especially important step in gene expres-
`sion because it is the main point at which the cell regulates which proteins are
`to be produced and at what rate. Bacterial RNA polymerase is a multisubunit
`complex. A detachable subunit. called sign-ta. (or) factor, is largely responsible for
`its ability to read the signals in the DNA that tell it where to begin transcribing
`[Figure 6—10}. RNA polymerase molecules adhere only weakly to the bacterial
`DNA when they collide with it, and a polymerase molecule typically slides
`rapidly along the long DNA molecule until it dissociates again. However, when
`the polymerase slides into a region on the DNA double helix called a promoter,
`a special sequence of nucleotides indicating the starting point for RNA synthe-
`sis, it binds tightly to it. The polymerase, using its 0 factor, recognizes this DNA
`sequence by making specific contacts with the portions of the bases that are
`exposed on the outside of the helix {Step l in Figure 6—10}.
`After the RNA polymerase binds tightly to the promoter DNA in this way, it
`opens up the double helix to expose a short stretch of nucleotides on each
`strand (Step 2 in Figure 6—10}. Unlike a DNA hellcase reaction [see Figure 5—15).
`this limited opening of the helix does not require the energy ofATP hydrolysis.
`Instead. the polymerase and DNA both undergo reversible structural changes
`that result in a more energetically favorable state. With the DNA unwound, one
`of the Mo exposed DNA strands acts as a template for complementary base-
`pairing with incoming ribonucleotides (see Figure 6—?], two of which are joined
`together by the polymerase to begin an RNA chain. After the first ten or so
`nucleotides of RNA have been synthesized [a relatively inefficient process dur-
`ing which polymerase syntheSizes and discards short nucleotide oligomers]. the
`(5 factor relaxes its tight hold on the polymerase and evenutally dissociates from
`it. During this process, the polymerase undergoes additional structural changes
`that enable it to move forward rapidly, transcribing without the 6 factor (Step 4
`in Figure 6—10}. Chain elongation continues {at a speed of approximately 50
`nucleoti ties! sec for bacterial RNA polymerases] until the enzyme encounters a
`second signal in the DNA, the terminator {described below]. where the poly-
`merase halts and releases both the DNA template and the newly made RNA
`chain (Step 7 in Figure 6—10]. After the polymerase has been released at a termi-
`nator. it reassociates with a free 5 factor and searches for a new promoter, where
`it can begin the process oftranscription again.
`Several structural features of bacterial RNA polymerase make it particularly
`adept at performing the transcription cycle just described. Once the 0 factor
`
`306
`
`Chapter 6 : HOW CELLS READ THE GENOME: FROM DNA TO PROTEIN
`
`Page 11
`
`
`
`
`
`Page 11
`
`

`

`RNA
`
`oiactor
`.
`
`Figure 6-H! The transcription cycle
`of bacterial RNA polymerase. In step
`Prommer
`I. the RNA polymerase holoenzyme (core
`k polymerase plus 6 factor) forms and then
`
`DNA
`
`locates a promoter (see Figure 6-—| 2).The
`
`1
`
`FINA polymerase
`
`fig
`\
`
`7
`
`RNA
`
`posizlons the poiymerase on the promoter and the template DNA has been
`unwound and pushed to the active site, a pair of moveable jaws is thought to
`clamp onto the DNA [Figure 6—11}. When the first 10 nucleotides have been
`transcribed. the dissociation of 0 allows a flap at the hack of the polymerase to
`
`rudder
`
`site of nucleotide
`addition
`
`newly synthesized
`rudder
`RNA tranScript
`
`
`template
`DNA
`strand
`
`polymerase unwinds the DNA at the
`position at which transcription is to begin
`(step 2) and begins transcribing (step 3).
`This i