chapter 4: dna and chromosomes. the structure and function of dna scientists intrigued by chemical...

Chapter 4: DNA and Chapter 4: DNA and

ChromosomesChromosomes

The Structure and Function of DNAThe Structure and Function of DNA

Scientists intrigued by chemical structure of genesScientists intrigued by chemical structure of genes► How can chemical info in genes copied and transmitted to daughter cells How can chemical info in genes copied and transmitted to daughter cells

millions of times, unchanged?millions of times, unchanged?► How are they able to direct the development of an organism and the daily life How are they able to direct the development of an organism and the daily life

of cell?of cell?► What kind of instructions does genetic info contain?What kind of instructions does genetic info contain?► How is genetic info organized in tiny space of cell?How is genetic info organized in tiny space of cell?

The Structure and Function of The Structure and Function of DNADNA

Landmark DiscoveriesLandmark Discoveries1940’s Studies with Fungi reveal Genetic info = instructions for making proteins1940’s Studies with Fungi reveal Genetic info = instructions for making proteins

1940’s Discovery of DNA is genetic info1940’s Discovery of DNA is genetic info

1953 Structure of DNA Resolved1953 Structure of DNA Resolved

1964 Genetic Code revealed1964 Genetic Code revealed


Structural Features of DNAStructural Features of DNA ► 2 Complementary chains of nucleotides2 Complementary chains of nucleotides► A, T, G, Cs held by H-bondsA, T, G, Cs held by H-bonds► Nucelotides linked by sugar-phosphate bondsNucelotides linked by sugar-phosphate bonds► Deoxyribose sugarDeoxyribose sugar► Polar 5’ phosphate linked to 3’ OHPolar 5’ phosphate linked to 3’ OH► Complementary base pairingComplementary base pairing► AntiparallelAntiparallel


Structure of DNA Provides Mechanism for Heredity:Structure of DNA Provides Mechanism for Heredity:

Genes carry biological info that must be copied accurately for transmissionGenes carry biological info that must be copied accurately for transmissionto next generation ea time cell dividesto next generation ea time cell divides

DNA encodes info through order or sequence of nucleotidesDNA encodes info through order or sequence of nucleotides

Organisms differ because of respective DNA which encodes differentOrganisms differ because of respective DNA which encodes differentbiological messagesbiological messages


From DNA to ProteinsFrom DNA to Proteins► Properties of proteins governed by 3d structureProperties of proteins governed by 3d structure► Sequence of nucleotides in gene must determine Sequence of nucleotides in gene must determine

sequence of aa in proteinsequence of aa in protein► Genetic code not obvious from DNA structureGenetic code not obvious from DNA structure


GenomeGenome► Complete set of info in organisms DNAComplete set of info in organisms DNA► Human genome: ~50,000 distinct Human genome: ~50,000 distinct

proteinsproteins


How is the Genome copied at ea cell division?How is the Genome copied at ea cell division? ea DNA strand can act as a templateea DNA strand can act as a template template used as mold for synthesis of new complementary strandtemplate used as mold for synthesis of new complementary strand


DNA is housed in the nucleusDNA is housed in the nucleus► comprises 10% cellcomprises 10% cell► Delimited by nuclear envelope with poresDelimited by nuclear envelope with pores► Envelope contiguous w/EREnvelope contiguous w/ER► Mechanically supported by intermediate filaments of nuclear lamina and Mechanically supported by intermediate filaments of nuclear lamina and

network of intermediate filaments surrounding outer membranenetwork of intermediate filaments surrounding outer membrane► Protects and compartmentalizesProtects and compartmentalizes

Packaging of DNA into Packaging of DNA into ChromosomesChromosomes

Challenges of Packaging DNAChallenges of Packaging DNA► How to get 2 meters of DNA into nucleus of 6 umHow to get 2 meters of DNA into nucleus of 6 um► Packaging accomplished w/ help of proteinsPackaging accomplished w/ help of proteins► Must be compacted in manner that still allows for it to be accessed by Must be compacted in manner that still allows for it to be accessed by

enzymes that govern replication, transcription, and repairenzymes that govern replication, transcription, and repair


Eucaryotic DNA is packaged into a set of Eucaryotic DNA is packaged into a set of chromosomeschromosomes

► DNA divided into set of chromosomesDNA divided into set of chromosomes► Chromosome= single DNA molecule and proteins associated with itChromosome= single DNA molecule and proteins associated with it


Human ChromosomesHuman Chromosomes► Human DNA 3.2 x 10Human DNA 3.2 x 1099 bases distributed bases distributed

over 24 chromosomesover 24 chromosomes► Each human cell contains 46 Each human cell contains 46

chromosomeschromosomes

22 homologous chromosomes22 homologous chromosomes

2 sex chromosomes (XX in females;2 sex chromosomes (XX in females;

XY in males)XY in males)


Bacteria have single circular DNA chromosomeBacteria have single circular DNA chromosome Proteins pack DNA into compact structureProteins pack DNA into compact structure Proteins associated w/ processes of transcription, repair, Proteins associated w/ processes of transcription, repair,

replicationreplication


Chromosomes Contain Long Strings of Chromosomes Contain Long Strings of GenesGenes

► Most impt function of chromosomes= carry Most impt function of chromosomes= carry genesgenes

► Gene= segment of DNA containing info for Gene= segment of DNA containing info for making protein (not true for RNA molec that making protein (not true for RNA molec that perform diverse functions)perform diverse functions)

► Correlation btwn complexity of organism and Correlation btwn complexity of organism and gene numbergene number

► Generally, more complex organisms have larger Generally, more complex organisms have larger genomes, but not alwaysgenomes, but not always

► No relationship btwn chromosome number and No relationship btwn chromosome number and complexity or genome sizecomplexity or genome size

Packing of DNA into ChromosomesPacking of DNA into Chromosomes

Most Striking Features of Human GenomeMost Striking Features of Human Genome► ~30,000 gene sequences~30,000 gene sequences► Vast amount of non-coding DNAVast amount of non-coding DNA► Lg number of transposable elementsLg number of transposable elements► Lg average gene size 27,000 bp when only 1300 required to Lg average gene size 27,000 bp when only 1300 required to

encode avg size protein encode avg size protein ► Critical info seems to be in alarming state of disarray!Critical info seems to be in alarming state of disarray!


Comparing DNA from Related OrganismsComparing DNA from Related Organisms► Gene Prediction difficult with all the “junk DNA”Gene Prediction difficult with all the “junk DNA”► Comparative DNA sequencing studies highlights important Comparative DNA sequencing studies highlights important

regions in genomeregions in genome► Conserved synteny= when gene order and content is Conserved synteny= when gene order and content is

conservedconserved

Packing of DNA into Packing of DNA into ChromosomesChromosomes

Chromosomes Exist in Different States Throughout Life of CellChromosomes Exist in Different States Throughout Life of Cell► Mitotic chromosomes= highly condensed chromosomes of dividing cellMitotic chromosomes= highly condensed chromosomes of dividing cell► During interphase chromosomes present as extended thin threads in During interphase chromosomes present as extended thin threads in

nucleus, not readily visiblenucleus, not readily visible


Essential Features of a Linear Chromosome:Essential Features of a Linear Chromosome:► CentromereCentromere► (2) Telomeres(2) Telomeres► Replication OriginsReplication Origins


DNA is highly condensed in chromosomesDNA is highly condensed in chromosomes► Compression of DNA performed by proteinsCompression of DNA performed by proteins

human chromosome 22 human chromosome 22

48 million bp48 million bp

extended 1.5 cm; packed in space 0.2 um extended 1.5 cm; packed in space 0.2 um

10,000X compaction ratio10,000X compaction ratio► DNA of interphase chromo less dense but overall compaction DNA of interphase chromo less dense but overall compaction

ratio still ~1000Xratio still ~1000X► Chromosome structure is dynamicChromosome structure is dynamic


Chromatin= nuclear DNA plus all the proteins bound to itChromatin= nuclear DNA plus all the proteins bound to it

Two classes of proteins bind to DNA to form chromosomeTwo classes of proteins bind to DNA to form chromosome

1.1. histoneshistones

2.2. nonhistonenonhistone


5 levels of Chromosomal Packaging5 levels of Chromosomal Packaging


Nucleosomes as Basic Unit of Eucaryotic Chromosome Nucleosomes as Basic Unit of Eucaryotic Chromosome StructureStructure

► Discovered 1974Discovered 1974► DNA wrapped around octameric core of histone proteinsDNA wrapped around octameric core of histone proteins► Repeat every 200 bpRepeat every 200 bp► Linker DNA btwn nucleosomes ~80 bpLinker DNA btwn nucleosomes ~80 bp► 3x level of compaction 3x level of compaction

human DNA 6.4 x 10human DNA 6.4 x 1099 bp w/ 30 million nucleosomes bp w/ 30 million nucleosomes


Characteristics of Histone ProteinsCharacteristics of Histone Proteins► 60 million/cell60 million/cell► Histone mass of chromosome = DNA massHistone mass of chromosome = DNA mass► Responsible for packaging DNA into nucleosomesResponsible for packaging DNA into nucleosomes► 4 different types: H2A, H2B, H3, H44 different types: H2A, H2B, H3, H4► Highly conserved, 102-135 aaHighly conserved, 102-135 aa► Rich in lys and argRich in lys and arg► Long N-terminal tail subjected to covalent Long N-terminal tail subjected to covalent

modificationmodification► Specialized variant histonesSpecialized variant histones


Histones as basic unit of nucleosomesHistones as basic unit of nucleosomes► Interface btwn DNA and histone extensive; ~142 H-bondsInterface btwn DNA and histone extensive; ~142 H-bonds► Hydrophobic and salt linkages also involved in DNA:histoneHydrophobic and salt linkages also involved in DNA:histone► Covalent modifications of N-terminal tail of histones controls aspects of Covalent modifications of N-terminal tail of histones controls aspects of

chromatin structurechromatin structure► Positioning of nucleosomes determined by DNA flexibility and other DNA Positioning of nucleosomes determined by DNA flexibility and other DNA

bound proteinsbound proteinsshort AT rich regions impart flexibility to DNAshort AT rich regions impart flexibility to DNAbound proteins can facilitate formation of nucleosomes or present obstaclebound proteins can facilitate formation of nucleosomes or present obstacle

► Arrangement of nucleosomes dynamicArrangement of nucleosomes dynamic


30 nm Chromatin Fiber30 nm Chromatin Fiber► Nucleosomes packed into 30 nm fiber Nucleosomes packed into 30 nm fiber ► Fiber formation facilitated by H1 histone (histone larger than Fiber formation facilitated by H1 histone (histone larger than

core histones) thot to link core histones togethercore histones) thot to link core histones together► Tails of core histones may attach one nucleosomes to anotherTails of core histones may attach one nucleosomes to another► Humans 1 cm in length spans nucleus 100XHumans 1 cm in length spans nucleus 100X


ATP Driven Remodeling ComplexesATP Driven Remodeling Complexes► Temporarily change nucleosome structureTemporarily change nucleosome structure► Complex of > 10 subunitsComplex of > 10 subunits► Some provide acessSome provide acess► Others reform nucleosomes or even transfer nucleosomesOthers reform nucleosomes or even transfer nucleosomes► Regulation of complexes may be afforded by Regulation of complexes may be afforded by

phosphorylation/dephosphorylationphosphorylation/dephosphorylation


Covalent Modification of Histone TailsCovalent Modification of Histone Tails► Affect stability of nucleosomeAffect stability of nucleosome► Attract specific proteinsAttract specific proteins► HATs histone acetyl transferases add acetyl HATs histone acetyl transferases add acetyl

grps to lysgrps to lys► HDACs histone deacetylases take acetyl HDACs histone deacetylases take acetyl

grps offgrps off► Methylation of lysMethylation of lys► Phosphorylation of serPhosphorylation of ser


Loops and CoilsLoops and Coils► Lg loops emanate from linear chromo axis► Loops always contains same DNA► Genes in loops are actively expressed► DNA not in loops condensed in

chromomeres on axis► Fiber converted to loop by histone

modifying enzymes, remodeling complexes and other proteins required for gene expression

► Ea loop is independent functional domain of chromatin structure


Lampbrush Chromosomes

Meiotically paired chromo growing amph. oocytes


Polytene Chromosomes of Drosophlia

►1024 identical DNA strands line up (10 cycles of DNA syn w/out cell division

►Alternating bands (95%) and interbands (5%); dynamic

►Interbands represent genes being actively transcribed


Features of Interphase Features of Interphase ChromosomesChromosomes

► 30 nm fiber = default structure30 nm fiber = default structure► Histone modifying enzymes, remodeling Histone modifying enzymes, remodeling

complexes, and other proteins resp. for complexes, and other proteins resp. for converting fiber into extended loop form converting fiber into extended loop form when gene expressedwhen gene expressed

► Ea loop = independent functional domain of Ea loop = independent functional domain of chromatin structurechromatin structure

► Chromosomes occupy distinct place in Chromosomes occupy distinct place in nucleusnucleus


HeterochromatinHeterochromatin► diff types of condensed chromatin w/ distinct features and rolesdiff types of condensed chromatin w/ distinct features and roles► Found in variety of organismsFound in variety of organisms► Represents 10% of chromosomeRepresents 10% of chromosome► Highly organized, resistant to gene expressionHighly organized, resistant to gene expression► Resp for function of telomeres, centromeres, and may protect Resp for function of telomeres, centromeres, and may protect

genome from transposable elementsgenome from transposable elements


EuchromatinEuchromatin► Less condensedLess condensed► Composed of all types of chromatin Composed of all types of chromatin

structures- 30 nm fibers, loops, etcstructures- 30 nm fibers, loops, etc► 90% of chromatin90% of chromatin► transcribabletranscribable


Gene Silencing and Position EffectsGene Silencing and Position Effects► Genes relocated to region of Genes relocated to region of

heterochromatin are silencedheterochromatin are silenced► Position effects = when activity of gene Position effects = when activity of gene

depends on position along chromosomedepends on position along chromosome► Position effect variegation = molted Position effect variegation = molted

expressionexpressionpattern results from patches of cells in which a pattern results from patches of cells in which a silenced gene has become activated or when silenced gene has become activated or when selected regions randomly packaged to selected regions randomly packaged to heterochromatin and silenced== both then are heterochromatin and silenced== both then are stabily inheritedstabily inherited


Position Effect Variegation demonstratesPosition Effect Variegation demonstrates1.1. Heterochromatin is dynamic and can spread into region and later Heterochromatin is dynamic and can spread into region and later

retractretract2.2. Whether heterochromatin or euchromatin can be inheritedWhether heterochromatin or euchromatin can be inherited


Special Form of Heterochromatin at Chromosomal EndsSpecial Form of Heterochromatin at Chromosomal Ends► Chromatin region extending 5,000 bp from chromo ends resistant to gene Chromatin region extending 5,000 bp from chromo ends resistant to gene

expressionexpression► Many diff proteins required for special foldingMany diff proteins required for special folding► Mutations in silent information regulator proteins (SIR) prevent silencing of genes Mutations in silent information regulator proteins (SIR) prevent silencing of genes

near telomeresnear telomeres► Cooperative binding of SIRs occurs to facilitate modificationsCooperative binding of SIRs occurs to facilitate modifications


SIRsSIRs► SIRs typesSIRs types

1.1. recognizes underacetylated N-terminal tails of selected recognizes underacetylated N-terminal tails of selected histoneshistones2.2. SIR2 = conserved histone deacetylase- deacetylation of SIR2 = conserved histone deacetylase- deacetylation of histone histone tails thot to allow tighter packaging of nucleosomes tails thot to allow tighter packaging of nucleosomes that are less that are less susceptible to remodeling complexessusceptible to remodeling complexes

► SIR2 requires NAD+ cofactor; NAD+ levels fluctuate w/ nutritional SIR2 requires NAD+ cofactor; NAD+ levels fluctuate w/ nutritional status of cell increasing under nutritional deprivationstatus of cell increasing under nutritional deprivation


Mitotic Chromosomes= Chromatin in Most Condensed Mitotic Chromosomes= Chromatin in Most Condensed FormForm


Mitotic Chromosomes= Chromatin in Most Condensed FormMitotic Chromosomes= Chromatin in Most Condensed Form► 10x smaller than interphase chromosomes10x smaller than interphase chromosomes► Function Function

1.1. compaction protects fragile DNA from breakingcompaction protects fragile DNA from breaking2.2. allows sister chromatids to be easily separated by mitotic allows sister chromatids to be easily separated by mitotic apparatusapparatus

► Condensin proteins use ATP to drive coiling of interphase chromosomesCondensin proteins use ATP to drive coiling of interphase chromosomes► Characteristic pattern of condensed AT rich regions and less Characteristic pattern of condensed AT rich regions and less

condensed CG rich regions- geisma stainingcondensed CG rich regions- geisma staining

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