5.2 :Creating Variation Let's look next at the causesof mutations and the different ways mutations can alter DNA. There are many causesof mutations. Radioactiveparticles pass through our bodies every day, for example, and if one of these particles strikes a molecule of DNA, it can damage the molecule's structure. Ultraviolet solar radiation strikes our skin cells and can causemutations to arise as thesecells divide. Manv chemicalsalso can interfere with DNA replication and lead to . Whenever a copiesits DNA, there is a small chance it may misread the sequenceand add the wrong nucleotide. Our cells have proofreading proteins that can fix most of these errors, but they some- times let mistakesslip bv. Mutations alter DNA in severaldifferent ways: . Point mutation: A single base changes from one nucleotide to another (also known as a substitution). ' : A segmentof DNA is inserted into the middle of an existing sequence. The insertion may be as short as a singlebase or as long as thousandsof bases (including entire ). . : A segment of DNA may be deleted accidentally.A small portion of a genemay disappear,or an entire set of genesmay be removed. ' Duplication: A segmentof DNA is copieda secondtime. A small duplicationcan produce an extra copy of a region inside a . Entire genescan be duplicated. In somecases, even an entire genomecan be duplicated. . Inversion: A segmentof DNA is flipped around and insertedbackward into its original position. . fusion: TWo are joined together as one. ' Aneuploidy: Chromosomesare duplicatedor lost, leading to abnormal levelsof ploidy. One way that mutations can give rise to geneticvariation is by altering the DNA within the coding region of a gene (the region that encodesa protein).An alteredcod- ing region may lead to a protein with a different sequenceof amino acids,which may fold into a different shape.This changecould causethe protein to perform its original activity at a faster or slower rate, or it may acquire a different activity.

Pointmutation lnversion TGCATTGCGTAGGC Y TGCATTCCGTAGGC

Insertion Chromosomefusion TGCATTTA6GC TGCATTCCGTAGGC A CCGJ Genomeduplication Deletion iiiliiiltiiliiiitii rGcArrckqrAGGc iiillliiliii!iitn Y TGCATTTAGGC ffffli|tiiiiiiHii.- iiB:tiir; fiiill;iiitli!ltfi--^ Geneduplication !-i ii fr it ii ffi lilliitlii$itttitFigure5.13 DNAcan experience # iiii!liiiil!*ji?iHseveraldifferent kinds of mutations, suchas point mutations, insertions, ffiFf:tuxil deletions,and duplications.

t.2 MUTATToNs:cREATTNG vARrATroN 133 Genesand in Bacteriaand Archaea

All livingthings use DNAor RNAto storegenetic information. But can be locatedthousands of basepairs away from genesthey con- eukaryotesare differentfrom bacteriaand archaeain the organi- trolin .Bacteria and archaea have self-splicing introns but zationof their DNAas wellas in how they replicateDNA and pass lackthe abundantspliceosomal introns found in eukaryotes,which it down to their offspring.The name eukaryotepoints to one of the requirea groupof proteinscalled the splrceosometo removethem most obviousthings that set eukaryotesapart from other forms of fromtranscripts. Bacteria and archaea thus lack the alternative splic- life.In Greek,it means"true kernel," referring to the nucleusin which ingfound in eukaryotes.As a result,they alwaysproduce the same eukaryoteskeep their DNA tightly coiled. Bacteria and archaea lack a proteinfrom any givengene. nucleus,and that trait has earnedthem the nameprokaryote, mean- Replicationin bacteriaand archaeais alsosimpler. They do not ing"before the kernel." performmitosis or .They do not havefull-blown sexual repro- fheword prokaryotecame into use beforethe dawn of molecular duction,in whichmales and females produce that combine systematics.Based on what is now knownabout the relationshipof in a newoffspring. Instead, bacteria and archaeatypically grow until thethree domains, a numberof leadingmicrobiologists argue against they are largeenough to divide.They then builda secondcopy of using the term .We know now that archaea are more their circularchromosome and then the two DNA moleculesare closelyrelated to eukaryotesthan they are to bacteria,which means draggedto eitherend of the dividingcell. The two daughtercells are that if a cladeincludes archaea and bacteria*butnot eukaryotes- identicalto the original,except for mutationsthat ariseduring DNA it wouldnot be monophyletic(Pace 2OO9). In general,taxonomists replication. avoidgiving names to paraphyletictaxa (Chapter4). Criticsargue Bacteriaand archaea have many of the samekinds of mutations that usingthe wordprokaryote makes as much senseas mammalo- found in eukaryotes,such as point mutationsand insertions.But gistsgiving a namefor the cladethat includesall mammals-except theycannot acquire genetic variation as a consequenceof reproduc- for rodents.So in this book,we'll avoid using prokaryote and simply tion the way we see in eukaryotes(i.e., independent assortment of referto bacteria andarchaea. chromosomes).As we'llsee in Chapter10, this difference can havea In bacteriaand archaea,a singlecjrcular chromosome floats majoreffect on howmutations spread through populations. withinthe cell.lt is not constrainedby a nucleus,nor is it tightly Beneficialmutations that increasethe survivalor reproductive spooledaround histones. That doesnot meanthis DNAis simplya rateof bacteriacan sweep quickly through a populationof microbes, loosetangle. Bacteria and archaeaproduce proteins that keepsec- thanksto naturalselection. As we'llsee in Chapter6, scientistshave tions of DNA organizedin twistedloops, Like eukaryotes, bacteria used microbesto perform important experimentson evolution, andarchaea can regulate gene expression by unwindingand winding observingnatural selection in action.And, as we'llsee in Chapter18, theirDNA. bacteriacan rapidlyevolve resistance antibiotics, turning what were Like eukaryotes,bacteria and archaeahave genetic regulatory onceeasily treated diseases into seriousthreats to publichealth. regionsupstream from their genes. factors can trig- Onereason that antibiotic resistance can spread so quicklyis that ger dramaticchanges in gene expressionin bacteriaand archaea bacteriaare not limitedsimply to passingdown their genes to their throughregulatory cascades. Bacteria and archaeacan altertherr descendants(known as vertical gene transfer). lt's also possible for geneexpression in responseto signalsfrom their environment.As one individualmicrobe to "donate"DNA to another,through a pro- a result,some speciescan producespores when conditionsturn cesscalled . stressful.Others can produce toxins when they sense other microbes One way for genesto move from one microbe to another is via competingfor resources. plasmids,which are small ringlets of DNAthat areseparate from the Overall,however, gene regulation is lesscomplex in bacteriaand mainbacterialchromosome. Under certain conditions. a microbe will archaeathan it rsin eukaryotes.Bacteria and archaea lack enhancers translateplasmid genes and assemblea tube calleda pilus,which (shortregions of DNAthat helpin transcription),for example,which linksthe genesto a neighboringcell. The donorcell can then pump B Promoter Terminator Ribosomes NucleoidDNA DNA template Plasma I membrane I Transcription Y Capsule Protein-coding sequence Cellwall RNAtranscript

It_ Translation I Y-2 .;e€ Porypeptide@

BoxFigure 5.1.1 A: Bacteriaand archaea differ from eukaryotes, like withina nuclearmembrane. B: The process of geneexpression and humans,in thatthe genetic material within the cell is not contained regulationis muchsimpler than in eukaryotes.

'134 cHAprERFtvE RAw MATERTAL:HERITABLE vaRtATtoN AMoNG tNDrvrDUALs i :opy of the plasmidthrough the pilus,and oftenpumps a copyof genesin species that are unlike their close relatives but are homolo- s:'ne of itschromosomal DNA as well. In effect, plasmids are genetic gousto genesfound in distantlyrelated clades. These studies indi- ::-asites,using bacteria and archaea as their hosts and spreading to catethat horizontalgene transfer has been a majorelement of evolu- -=,vhosts through the piliencoded in their own genes. However, they tion.In E. coli,Ior example, 80 percentof all the genes in its :r alsocarry genesencoding beneficial traits, such as antibiotic showevidence of horizontalgene transfer at somepoint since the -:srstance,which can provide advantages to theirhosts, lastcommon ancestor of bacteria(Dagan and Martin 2007). As we Virusescan alsocarry out horizontalgene transfer. As they rep- sawin Chapter4,horizonlal gene transfer is promptingscientists to :3te, some virusescan accidentallyincorporate host genesinto revisetheir concepts of speciesand of theoverall shape of the tree '-.rr owngenome. When they infect a newhost, they can insert those oflife. r:res intotheir new host's chromosome. Comparedto bacteriaand archaea, eukaryotes appear to have Inmanycases, horizontal gene transfer is a deadend. The donated experiencedrelatively little horizontal gene transfer. There are a a:res areharmf ul to the recipientcell, which dies or growstoo slowly numberof possibleexplanations for thisdifference. One is oppor- ': competewith other individuals.But if a microbeacquires a useful tunity:the complexity of eukaryoticDNA replication may not afford g:re, naturalselection can favor it. Evidencefor successfulhorizon- theopportunity to takeup foreigngenes. In bacteriaand archaea, a :a genetransfer can be foundin studieson the spreadof antibiotic contiguousset of genes may form a functionalunit, called an operon, ':srstance:the samegene often turns up in differentspecies. lt's also inwhich they are all controlled by the same upstream regulatory ele- :':ssibleto identifycases of horizontalgene transfer that occurred ments.The entire operon can be insertedinto a newhost, where it - llionsof yearsago by performinglarge-scale comparisons of DNA maybe ableto providea usefulfunction. Eukaryotes lack operons, - bacteriaand archaea, In a numberof cases,scientists have found however,sothat foreign genes are less likely to beuseful in a newcell.

Vertical genetransfer: Theprocess parentce,@ of receivinggenetic material from an ancestor.

A Horizontal genetransfer: Any process o'n"!'-l',=t="j.1-1 in whichgenetic material is transferred W to anotherorganism without descent. Y Plasmids:Molecules of DNA,found mostoften in bacteria,that canrepli- cateindependently of chromosomal @@ DNA. Plasmid /\

ldentical daughtercells

Horizontal 9ene transfer

/ \ Box Figure 5.1.2 Bacteriareproduce by dividingin two. As they preparefor the division,they separate the two strandsof DNA and add a newstrand to eachone, creating two new DNA mol- ffi@ ecules,which are typically identical to the originalone. On rare Cellsare genetically identical to Cellscarry DNA of ancestors ancestors,except for an acquired aswell as DNA acquired occasions,however, genes can be passedfrom one bacteriumto pointmutation. by horizontalgene transfer. another,through a processknown as horizontal gene transfer.

5.2 MUTATTONS:CREATTNG VARtATtON 135 .t t1 i1 tl. 14 !.frl I r

Figure5.14 Pointmutations in humanprotein-coding genes can have phenotypic effects that rangefrom the benign(e.g., eye color) to the severe.Shown here are a varietyof striking(but rare)mutations. For example, a mutationin the FGFR3gene that replacesthe aminoacid proline with serineat position380 is responsiblefor albinism(A; Oetting and King 1993); a singleC-to-T transitionin the IMBRlgene leads to triphalangealthumb polydactyly (B; Wang et a1.2007);a missensemutation in exonXIV of theGl13 gene, replacing the amino acid proline with serine, lead: to Greig'scephalopolysyndactyly (C); a missensesubstitution in the K/f proto-oncogenereplacing theamino acid arginine with glycine at position795 leads to piebaldism(D; Sdnchez-Martin et al. 2003);a mutationin theACVR1 gene substituting the amino acid arginine with histidine at positio' 206results in fibrodysplasiaossificans progressiva (E; Shore et a1.2006);and a pointmutation in the LaminA genecauses Hutchinson-Gilford progeria syndrome (F; Eriksson et al.2OO3).

Cis-actingelements: Stretches of Mutations can also have important effectswithout altering the product of a gt'n' DNAlocated near a gene-either Instead, they can simply change how much of a protein is made, or they can chan,. immediatelyupstream (adjacent to the timing or location of its production. These changes in levels o/gene expres\r,, the promoterregion), downstream, or can alter the behavior of cells or tissues and can have profound consequences 1, insidean intron-that influence the evolution as an additional component of heritable variation. Mutations can alter ger . expressionof that gene.Cis regions expressionby affecting where, when, or how much a gene is transcribed. For exanrpl. oftencode for bindingsites for oneor mutations may cause a transcription factor to bind more strongly than it did befor moretransposable factors. Or they may prevent a specific transcription activator protein from binding, so tl).. the gene no longer can be expressedin a particular kind of tissue. Transcription factors, hormones, and other regulatory molecules are themseh' Trans-actingelements: Sequences of encoded in genes,which means mutations that alter their genes ultimately can a11,. DNAthat arelocated away from the the genes they regulate. As a result of these interactions, a gene can be affecteil : focalgene (e.g., on anotherchromo- a mutation that is far away from the gene itself. (Nearby elements that affect gt.: some).These stretches of DNAgener- expression are cis-acting elements; faraway ones are trans-acting elements.) allycode for a protein,microRNA, or Mutations, as we'll see in later chapters, are required for evolution to occur. I.] otherdiffusible molecule that then it's important to bear in mind that they're extremely rare. To measure just how olt, \nfluencesexpress\on of \he foca\ gene mutations occur, scientists can erther run experiments onpopu\ations o{ ce\\s or ma,

136 cHAprERFrvE RAw MATERtaL:HERITaBLE vARlarroN AMoNG rNDrvrDUALs Table5.3 Themany ways mutations caninfluence expression of a gene. Location of Mutation Type of Mutation Consequencefor Gene Action

CodingRegion Substitution,insertion, Altersthe product of the gene, deletion,duplication. andthus its functionor activity. cis-Regulatory Substitution,insertion, Altersthe timing, location, or Regions deletion,duplication that levelof expressionof the gene. altersthe bindingaffnity Altersthe developmentalor of promoters,activators, environmentalcontext in which repressors,erc. the geneis expressed.

trans-Regulatory Mutationto codingregions Altersthe bindingaffnity and Regions of trans-actingfactor. thus the activityof a promoter, activator,repressor, etc.

Mutationto cis- or trans- Alterswhere, when, or to what regulatoryregions of extentinhibitory, activating, or trans-actingfactors. othertrans-acting regulatory fac- tors areexpressed.

Physiological Mutationsalter where, Altersthe timing, location, or Pathways(e.g., when,or howmuch an en- levelof expressionof the gene. docrinesignal is produced. hormones) Altersthe developmentalor environmentalcontext in which the geneis expressed.

surveysof living populations.In 2008,for example,Michael Lynch and his colleagues at Indiana University rearedcolonies ofyeast (Lynchet al. 2008).From a singleances- tor, Lynch and his colleaguesreared hundreds of geneticallyidentical populations of yeast.They then allowed theselines to reproducefor 4800 generations.After select- ing someof the descendants,the scientistssequenced all 12 million basepairs of DNA in eachcell's genome. Each time a yeast cell divides, the scientistsfound, each site in its DNA has a 0.00000003percent chance of undergoinga point mutation. This probability is so low that a typical yeastcell may not acquirea single point mutation in its whole genome. But in a population of millions of yeastcells, point mutations will arisein thousands of individuals in eachnew generation. Somaticmutations: Mutations that Lynch'sexperiments also showed that different kinds of mutations occur at dif- affectcells in the body("soma") of ferent rates.The investigatorsfound that each gene has a roughly one-in-a-million anorganism. These mutations affect chanceof being lost or duplicatedeach time a cell divides.Duplications and deletions allthe daughtercells produced by are rare, in other words, but they're also about a thousand times more likely than the affectedcell andcan affectthe point mutations. phenotypeof the individual.In animals, Estimatingmutation ratesin multicellular organismslike humans is a more com- somaticmutations are not oassed plicated matter, for severalreasons. First, all of our genes are present in duplicate downto offspring.ln plants,somatic tdiploidy),and this meansthat many mutations that arisein one of the chromosomes mutationscan be passeddown during rvill be hidden or maskedby a functional copy of the same gene on the sister chro- vegetativereproduction. mosome.Second, not all mutations that arise in our bodies are transmitted to our offspring.Any individual cell in our body has a chanceof mutating asit divides.If it's Germ-linemutations: Mutations that a skin cell,the skin cellsthat descendfrom it will continue to carry that mutation. But affectthe gametes(eggs, sperm) of an this lineageof cells will come to an end when we die. Such mutations are known as individualand can be transmitted from somatic mutations becausethey occur in the "soma," or body. parentsto offspring.Because they can If, on the other hand, a mutation arisesin the line of cells that gives rise to sperm be passedon, germ-line mutations cre- or egg cells, it may be passedon to ofispring. And those offspring, in turn, may pass atethe heritablegenetic variation that the mutation down to their own descendants.These mutations are known as germ- is relevantto evolution.

5.2 MUTATToNs:cREATTNG vARrATroN 137 line mutations. Even though somatic mutations sometimes drastically reduce the performance and fitness of an individual (e.g.,many cancers,as we seein Chapter 18, are the result of somatic mutations), they are not heritable. Heritable variation within populations arises becauseof the gradual accumulation of germ-line mutations. Until recently scientists could make only very indirect estimatesof the germ-line mutation rate in humans. One common method was to study the rate of diseasesthat are caused by a mutation of a single gene. But improvements in DNA sequencing have made it possible for scientists to make far more precise estimates.In 2010,Leroy Hood of the Institute for SystemsBiology in Seattleand his colleaguessequenced the entire of tvvo human siblings and their parents. They searchedthe genomes of the children for mutations that their parents did not carry. All told, they identified 70 new mutations in eachchild {Roachet al. 2010).

KeyConcepts Changesin levelsof geneexpression can have profound consequences forevolution by adding anothercomponent to heritablegenetic variation.

Geneticchanges in geneexpression arise when mutationsoutside of the codingregions affect where, when,or how mucha geneis transcribed. i 5.3 Heredity Once new mutations arise,organisms can passthem down, along with their unmu- tated DNA, to their offspring. Bacteria and archaeareproduce by dividing and making a new copy of their genomefor eachdaughter cell (Box 5.1).In sexuallyreproducing, multicellular eukaryotes,on the other hand, heredity is more complex.For one thing, only germJine cells can pass on genes to offspring. For another, the development of cells introduces new genetic variation, so that sexually reproducing organisms producegenetically unique offspring insteadof clones. One reasonthat parentsdo not producefamilies of identical children is that each parent'sown paired chromosomesare not identical.One chromosomemay have one :One of severalalternative version of a gene (an allele) while the other chromosomehas an allelewith a slightly formsof the DNAsequence of the different DNA sequence.Differences between the chromosomesin eachpair generate samelocus. variation among the cells that are produced, so that no tvvo sperm or egg cells are identical. Gametesof sexuallyreproducing organismsare produced through a distinctive Meiosis:A form of celldivision that kind of cell division known as meiosis (Figure5.'15). Meiosis can generatea stunning occursonly in eukaryotes,in whichthe amount of geneticvariation. During the final stageof meiosis,each pair of chromo' numberof chromosomesiscut in half. somes separates,so that only a single copy ends up in each daughter cell. Which copy' Meiosisgives rise to gametesor endsup in eachcell occursrandomly, and it occursrandomly for eachof the different sporesand is essential for sexual chromosomal pairs. This means that a single sperm cell may inherit the maternal reproduction. copy of one chromosome,but the paternal copy of another.Humans have 23 pairs of chromosomes,and the independentassortment of eachseparate chromosome can result in many different combinationsof maternally inherited and paternally inher ited genes. In addition, each pair of chromosomes may cross over during meiosis and Geneticrecombination: The exchange exchange segments of DNA in a process known as . By the ofgeneticmaterial between paired time these chromosomes are packed into gamete cells, many of them have alreadr' chromosomesduring meiosis. Recom- been rearranged so that they differ from either of the parental chromosomes.Recom binationcan form newcombinations bination also can creategenetic variation among the different gametesproduced br of allelesand is an importantsource of an individual. Consequently,the processesof independent assortment and recombi heritablevariation. nation can, through the rearrangement of , generate a staggering number of possiblegamete genotypes. This rich mixing of alleles occurs in the formation of gametes of both the male and the female. Sexualreproduction brings the chromosomalforms of each parenr

138 cHAprERFrvE RAw MATERTaL:HERITaBLE vaRtarloN AMoNG lNDtvtDUALS Father

Chromosomes Chromosomes areduplicated areduplicated.

Chromosomescross over andexchange segments of DNA.

Segregationof homologous chromosomes

Segregationof sisterchromatids

Haploid gametes

f \

gametereceives only one copy from Figure5.15 Amongsexually reproducing organisms, like humans, exchangesegments of DNA.Each As a result,each child carries a unique malesand females combine their gametes to reproduce.During the eachoair of chromosomes. parents. productionof gametes,each pair of chromosomescross over ancl combinationof the DNAof hisor her

5.3 HEREDTTY 139 Figure5.16 Independentassortment together, creating yet another combi- occursduring meiosis in eukaryotic nation of alleles.Consequently, when lrlr Maternalcopies organismsand produces gametes a human sperm cell fertilizes an egg, with a mixtureof maternaIand the chromosomescombine to produce paternalchromosomes. Along with a new set of 23 pairs. But this new set chromosomalcrossover, independent is drawn from a rich pool of genetic assortmentincreases genetic diversity variants reflecting millions of possible by producingnovel combinations of combinations of allelesinherited from alleles. both the father and the mother. The particular combination that fuses to form the new diploid offspring indi' vidual is literally one in a million, and it is not likely to happen twice. This is why sibling offspring from the same parents alwaysdiffer in their inherited characteristics.(An exception to that rule, of course,is identical who developfrom a single fertilized egg.) The best estimate of the differ- ences between our paired chromo. somes comes from Craig Venter, a genome-sequencingpioneer. In 2007 he and his colleaguespublished the complete sequenceof his own genome (Levy et aL.2007).They comparedeach pair of chromosomes,tallying up the @@ @)@ differences.The researchersidentified Novelcombinations of alleles 3.2 million placeswhere a single nucle otide in one chromosomedid not match the correspondingnucleotide in its partner. The scientistsalso found about a million segmentsof DNA on one chromosomethat were missing from its partner,or that had been inserted.

KeyConcept Becauseof theindependent assortment of chromosomesand genetic recombination, meiosis can generateextraordinary genetic diversity among gametes.

Genotype:The genetic makeup of an 5.4 TheLink between Most Phenotypes individual.Although a genotypein- andGenotypes ls Complex cludesall the alleles of allthe genes in Scientistsdraw a distinction between the genetic material in an organism and the that individual,the term isoften used traits that the geneticmaterial encodes.The geneticmakeup of an organism is knor.r'n to referto the specificalleles carried by as its genotype, and the manifestationof the genotypeis known as the phenotype. anindividual for anyparticular gene. Organismsdo not inherit a phenotype;they inherit genes,which together constitute Phenotype:An observable,measur- a genotype,which givesrise to a phenotype. ablecharacteristic of anorganism. A Understandinghow phenotypesemerge from genotypesis no easytask. A trait phenotypemay be a morphological does not come with a label on it, detailing all the genesthat helped to build it and structure(e.g., antlers, muscles), a the specificrole played by each of the genes.Instead, scientists rely on a variety of developmentalprocess (e.g., learning), methods to explore how genesand gene expressioncontribute to the formation oi a physiologicalprocess or performance organismalphenotypes. These methods range from controlledbreeding experiment' trait(e.g., running speed), or a behavior to detailed genetic mapping studies-even to perturbations of expressionof focal (e.g.,mating display). Phenotypes can developmentalgenes (Chapter 10 describesmany of thesemethods in greaterdetailr evenbe the moleculesproduced by The traits that Mendel studied in his peas (Box5.2) have relatively simple, dis genes(e.9., hemoglobin). crete, alternative phenotypic states.The peas were either wrinkled or smooth, for

f4O cHAprERFrvE RAw TTATERTAL:HERITABTE vARtATtoN At oNG tNDtvtDUALs