DNA: The Genetic Material PDF

CH 5.2 Genetics– BIO310 DNA: The Genetic Material Structure of Eukaryotic Chromosomes in Nondividing Cells Definition of chromatin The structures of nucleosomes, the 30-nm fiber, and radial loop domains Noll’s results and how they support the beads-on-a-string model Description of a chrom...

CH 5.2 Genetics– BIO310 DNA: The Genetic Material Structure of Eukaryotic Chromosomes in Nondividing Cells Definition of chromatin The structures of nucleosomes, the 30-nm fiber, and radial loop domains Noll’s results and how they support the beads-on-a-string model Description of a chromosome territory Difference between euchromatin and heterochromatin and between constitutive heterochromatin and facultative heterochromatin 2  Eukaryotic Chromatin Compaction If stretched end to end, a single set of human chromosomes would be over 1 meter long! ◦ Yet the cell’s nucleus is only 2 to 4 mm in diameter Therefore, DNA must be tightly compacted to fit The compaction of linear DNA in eukaryotic chromosomes involves interactions between DNA and various proteins ◦ In eukaryotes, this DNA-protein complex is known as chromatin Changes in the proteins affect the degree of chromatin compaction during the life of the cell 3  Nucleosomes The repeating structural unit within eukaryotic chromatin is the nucleosome A nucleosome is composed of double-stranded DNA wrapped around an octamer of histone proteins ◦ An octamer is composed two copies each of four different histones ◦ H2A, H2B, H3, and H4 (core histone proteins) ◦ 146 or 147 bp of DNA make 1.65 negative superhelical turns around the octamer ◦ Linker region between nucleosomes 20 to 100 bp ◦ Structure of connected nucleosomes like “beads on a string” ◦ This shortens the DNA length about seven-fold 4 5 Histone proteins Histone proteins contain many positively-charged amino acids - lysine and arginine ◦ These bind to the negatively charged phosphates along the DNA ◦ Histone proteins have a globular domain and a flexible, charged amino terminus or ‘tail’ 6  Histone proteins2 There are five types of histones ◦ H2A, H2B, H3 and H4 are the core histones Two of each make up the octamer ◦ H1 is the linker histone Binds to linker DNA Also binds to nucleosomes ◦ But not as tightly as are the core histones Nonhistone proteins also bind the linker region, aid in chromosome compaction and affect gene expression 7 ©Laguna Design/SPL/Science Source 8 Play a role in the organization and compaction of the chromosome 9  The Structure of the Nucleosome The model of nucleosome structure was proposed in 1974 by Roger Kornberg DNA double helix wrapped around an octamer of core histone proteins Kornberg based his proposal on various observations about chromatin ◦ Biochemical experiments ◦ X-ray diffraction studies ◦ Electron microscopy images 10 The Structure of the Nucleosome 2 Markus Noll tested Kornberg’s model The Hypothesis ◦ If the Kornberg model was correct, digestion of the linker region of chromatin should result in DNA fragments that are only 200 bp long ◦ The rationale ◦ Linker DNA is more accessible than the “core DNA” to the enzyme DNase I ◦ DNase I cuts should occur in the linker DNA but not the DNA wrapped around the core histones 11 12 The Data At low concentrations, DNase I did not cut all the linker DNA At high concentrations of DNase I, all chromosomal DNA digested into fragments that are ~ 200 bp in length Source: Adapted from Noll, M. (1974), Subunit Structure of Chromatin, Nature, vol. 251, 249–251. Results strongly support nucleosome model for chromosome structure 13  Nucleosomes Join to Form a 30-nm Fiber Nucleosomes associate with each other to form a more compact structure – the 30-nm fiber Shortens the total length of DNA 7-fold Structure has been difficult to determine ◦ Zigzag model proposed ©Jerome Rattner/University of Calgary 14  Nucleosomes zigzag back and forth with a straight linker region ©Jerome Rattner/University of Calgary 15 Further Compaction of the Chromosome Together the nucleosomes and 30-nm fiber shorten the DNA about 50-fold A third level of compaction involves formation of loops (called loops domains) CCCTC binding factor (CTCF) binds to 3 regularly spaced repeats of the sequence CCCTC Two different CTCFs bind to the DNA to form a loop SMC proteins can also form a DNA loop SMC proteins often found in the region of CTCFs 16 17 Chromosome Territories Each chromosome in the nucleus is located in a distinct chromosome territory These are visible when chromosomes are fluorescently labeled in interphase cells 18 Courtesy of Felix Habermann and Irina Solovei, University of Munich (LMU, Biocenter) 19 Heterochromatin versus Euchromatin Euchromatin Less condensed regions of chromosomes Transcriptionally active Heterochromatin Tightly compacted regions of chromosomes Transcriptionally inactive (in general) 20  Constitutive versus facultative heterochromatin Constitutive heterochromatin Chromosomal regions that are heterochromatic in all cell types Highly repetitive sequences Centromeres & telomeres Facultative heterochromatin Differs among different cells of the body Occurs where genes are located Can convert to euchromatin as a way to regulate gene expression 21 22  Structure of Eukaryotic Chromosomes During Cell Division The levels of compaction that lead to a metaphase chromosome 23  Metaphase Chromosomes As cells enter M phase, the level of compaction changes dramatically to become metaphase chromosomes ◦ Nucleosomes ◦ Zigzag structure to form a 30-nm fiber ◦ 30-nm fibers form loop domains ◦ Closer association of loop domains with each other and closer association between adjacent nucleosomes ◦ Chromosome has a diameter of 700 nm; two chromatids have a diameter of 1400 nm but much shorter length than interphase chromosome ◦ These highly condensed metaphase chromosomes undergo little gene transcription 24 (a) ©Dr. Barbara A. Hamkalo; (b) ©Jerome Rattner/University of Calgary 25  Compaction level in euchromatin Compaction level in heterochromatin (c) ©Dr. James Paulson, Ph.D.; (d) Peter Engelhardt/Department of Virology, Haartman Institute Access the text alternative for slide images. 26 Metaphase Chromosomes Controversy over whether or not nonhistone proteins form a scaffold for organizing the shape of metaphase chromosomes When a metaphase chromosome is treated with high salt to remove histones, the highly compact configuration is lost Bottoms of loops remain attached to a scaffold made of nonhistone proteins Does this scaffold organize metaphase chromosomes or is it a remnant of treatment with high salt? 27 (a) Courtesy of Peter Engelhardt/Department of Virology, Haartman Institute; (b) ©Don W. Fawcett/Science Source 28 The Chromosome Composition of Humans BSIP/Phototake 29  Microscopic Examination of Eukaryotic Chromosomes The characteristics that are used to classify and identify chromosomes 30  Genetic Variation Genetic variation refers to differences in alleles and chromosomes, either among members of the same species or among different species ◦ Allelic variations are variations in specific genes ◦ Variations in chromosome structure and number ◦ Typically affect more than one gene ◦ Important in evolution ◦ Can cause disease ◦ Important for new strains of crops 31 Variations in Chromosomes Can be Seen by Light Microscopy Structure and number of chromosomes typically studied by light microscopy ◦ Cytogeneticist – Scientist who studies chromosomes under the microscope Different species can be distinguished from each other based on the number and size of chromosomes 32 a: (1) Scott Camazine/Science Source; (2) Michael Abbey/Science Source; (3) Carlos R. Carvalho/Universidade Federal de Viçosa; c: PTP/Phototake 33 Chromosome Classification Different chromosomes of the same species can be also distinguished from each other Chromosomes can be classified by ◦ Size ◦ Position of centromere ◦ Banding pattern ◦ Staining reveals bands 34 Centromere position Metacentric – centromere near the middle Submetacentric – slightly off center Acrocentric – more off center Telocentric – centromere at the end 35 36  Karyotype A karyotype is a micrograph of metaphase chromosomes from a cell arranged in standard fashion Chromosomes are arranged from largest to smallest 37 38 Giemsa staining Different chromosomes have similar sizes and centromeric locations Staining is used to identify specific chromosomes Example: Giemsa stain – G bands Next slide figure- shows conventional numbering system used to designate G bands along a set of human chromosomes 39 40 Why is the banding pattern useful? ◦ Distinguish between different chromosomes ◦ Detect changes in chromosome structure ◦ Used to assess evolutionary relationships between species 41 Changes in Chromosome Structure: An Overview The four types of changes in chromosome structure 42  Mutations Can Alter Chromosome Structure Primary ways chromosome structure can be altered: ◦ Deletion (also called Deficiency) ◦ Portion of the chromosome is missing ◦ Duplication ◦ Portion of the chromosome is repeated ◦ Inversion ◦ A change in the direction of part of the genetic material along a single chromosome ◦ Translocation ◦ Segment of one chromosome becomes attached to a non homologous chromosome 43 Translocations Simple translocations ◦ Single piece of chromosome is attached to another chromosome Reciprocal translocations ◦ Two different chromosomes exchange pieces 44 Human chromosome 1 Human chromosome 21 45  Deletions The phenotypic consequences of deletions depends on ◦ Size of the deletion ◦ Chromosomal material deleted ◦ Are the lost genes vital to the organism? 47 A chromosomal deletion occurs when a chromosome breaks and a fragment is lost breaks and a fragment is lost 48 Detrimental effects of deletions When deletions have a phenotypic effect, they are usually detrimental ◦ Example: cri-du-chat syndrome in humans Caused by a deletion in the short arm of chromosome 5 Abnormalities include mental deficiencies, unique facial abnormalities, unusual catlike cry 49 Duplications Duplications result in extra genetic material May be caused by abnormal crossing over Crossover may occur at misaligned sites on homologs during meiosis Can occur when a chromosome contains two or more homolgous segments of DNA with identical or similar sequences – repetitive sequences Called nonallelic homologous recombination – sites are homologous but are not alleles of the same gene Resulting chromosome with the extra genetic material carries a gene duplication – number of copies of a gene increases from 1 to 2 51 52 Duplications 2 Like deletions, the phenotypic consequences of duplications tend to be correlated with size; more likely to have phenotypic effects if they involve a large piece of the chromosome Duplications tend to have less harmful effects than deletions of comparable size ◦ In humans, relatively few well-defined syndromes are caused by small chromosomal duplications ◦ Example: Charcot-Marie-Tooth disease 53  Inversions A chromosomal inversion is a segment that has been flipped to the opposite orientation ◦ Total amount of genetic information stays the same ◦ Therefore, the great majority of inversions have no phenotypic consequences Pericentric inversion – the centromere is within the inverted region Paracentric inversion – the centromere is outside the inverted region 61  Centromere lies Centromere lies outside inverted within inverted region region 62  Consequences of Inversions In rare cases, inversions can alter the phenotype of an individual ◦ Breakpoints ◦ The breaks leading to the inversion occur in a vital gene ◦ Position effect ◦ A gene is repositioned in a way that alters its gene expression About 2% of the human population carry inversions that are detectable with a light microscope ◦ Most of these individuals are phenotypically normal ◦ However, in a few cases they can produce offspring with genetic abnormalities 63 Inversion Heterozygotes 1 Individuals with one copy of a normal chromosome and one copy of an inverted chromosome Such individuals may be phenotypically normal ◦ But they have a high probability of producing abnormal gametes Due to crossing-over in the inverted segment 64 Inversion Heterozygotes 2 During meiosis I, homologous chromosomes synapse with each other ◦ For the normal and inversion chromosome to synapse properly, an inversion loop must form ◦ If a crossover occurs within the inversion loop, highly abnormal chromosomes are produced ◦ Dicentric chromosome contains 2 centromeres connected by a dicentric bridge (occurs with a paracentric inversion) ◦ Acentric fragment has no centromere and is lost and degraded in subsequent cell divisions (occurs with a paracentric inversion) 65 66 Translocations involve exchanges between different chromosomes A chromosomal translocation occurs when a segment of one chromosome becomes attached to another Ends of eukaryotic chromosomes called telomeres prevent translocations from occurring Broken chromosomes ends lack telomeres and become reactive In reciprocal translocations two non-homologous chromosomes exchange genetic material 67 Telomeres prevent chromosomal DNA from sticking to each other 68 Reciprocal Translocations Reciprocal translocations arise from two different mechanisms 1. Chromosomal breakage and DNA repair 2. Non-homologous crossovers Reciprocal translocations lead to a rearrangement of the genetic material, not a change in the total amount; also called balanced translocations Reciprocal translocations, like inversions, are usually without phenotypic consequences ◦ In a few cases, they can result in position effect 69 Unbalanced translocation Carriers of a balanced translocation are at risk of having offspring with an unbalanced translocation In unbalanced translocations, significant portions of genetic material are duplicated and/or deleted Unbalanced translocations are associated with phenotypic abnormalities or even lethality 70  Robertsonian translocation Example: the Robertsonian translocation ◦ Most common type of chromosomal rearrangement in humans ◦ Approximately one in 900 births ◦The majority of chromosome 21 is attached to chromosome 14 ◦This translocation occurs such that ◦ Breaks occur at the extreme ends of two non-homologous chromosomes ◦ The small acentric fragments are lost ◦ Larger fragments fuse at centromeric regions to form a single chromosome 71 72 Changes in Chromosome Number: An Overview Definition of euploid and aneuploid Polyploidy and aneuploidy 77 Variation in Chromosome Number Chromosome numbers can vary in two main ways ◦ Euploidy ◦ Variation in the number of complete sets of chromosomes ◦ Examples: triploid (3n), tetraploid (4n) ◦ Organisms with 3 or more sets of chromosomes are also called polyploid ◦ Aneuploidy ◦ Variation in the number of particular chromosomes within a set ◦ Regarded as abnormal ◦ Examples: trisomy (2n+1), monosomy (2n-1) 78 79 Aneuploidy Aneuploidy – Variation in the number of particular chromosomes within a set Aneuploidy commonly causes an abnormal phenotype ◦ It leads to an imbalance in the amount of gene products ◦ Three copies can lead to 150% production of the hundreds or even thousands of gene products from a particular chromosome 81 In most cases, these effects are detrimental They produce individuals that are less likely to survive than a euploid individual 82 Aneuploidy in Humans 1 Alterations in chromosome number occur frequently during gamete formation ◦ About 5 to 10% of embryos have an abnormal chromosome number ◦ Indeed, ~ 50% of spontaneous abortions are due to such abnormalities In relatively rare cases, an abnormality in chromosome number produces an offspring that can survive 83 Aneuploidy in Humans 2 Autosomal aneuploidies that are most compatible with survival are trisomies 13, 18 and 21 Sex chromosome aneuploidies generally have less severe effects ◦ Explained by X inactivation All but one X chromosome transcriptionally suppressed Phenotypes of X chromosome aneuploidies may be due to ◦ Expression of X-linked genes prior to X-inactivation ◦ Imbalance in the expression of pseudoautosomal genes 84 TABLE 8.1 Aneuploid Conditions in Humans Condition Frequency Syndrome Characteristics Autosomal Trisomy 13 1/15,000 Patau Mental and physical deficiencies, wide variety of defects in organs, large triangular nose, early death Trisomy 18 1/6000 Edward Mental and physical deficiencies, facial abnormalities, extreme muscle tone, early death Trisomy 21 1/800 Down Mental deficiencies, abnormal pattern of palm creases, slanted eyes, flattened face, short stature Sex Chromosomal XXY 1/1000 Klinefelter Sexual immaturity (no sperm), breast swelling (males) XYY 1/1000 Jacobs Tall and thin (males) XXX 1/1500 Triple X Tall and thin, menstrual irregularity (females) X0 1/5000 Turner Short stature, webbed neck, sexually undeveloped (females) 85 Influence of Age on Aneuploidies 1 Some human aneuploidies are influenced by parental age ◦ Older parents more likely to produce abnormal offspring ◦ Example: Down syndrome (Trisomy 21) Incidence rises with the age of either parent, especially mothers 86 Influence of Age on Aneuploidies 2 Down syndrome ◦ Failure of chromosome 21 to segregate properly due to chromosomal nondisjunction, usually in meiosis I in the oocyte Age of oocytes may play a role ◦ Primary oocytes are produced in the ovary of fetus prior to birth Oocytes arrested in prophase I until the time of ovulation Length of time that oocytes are arrested in prophase I may contribute to an increased frequency of nondisjunction 87 Euploidy 1 Euploidy – Variation in the number of complete sets of chromosomes Most species of animals are diploid In many cases, changes in euploidy are not tolerated ◦ Polyploidy in mammals is generally a lethal condition 89 Euploidy 2 Some euploidy variations are naturally occurring ◦ Example: Bees are haplodiploid ◦ Female bees are diploid ◦ Male bees (drones) are monoploid ◦ Contain a single set of chromosomes Many examples of vertebrate polyploid animals have been discovered ◦ Example: The frog Hyla 90 ©A.B. Sheldon 91 Endopolyploidy In many animals, certain body tissues display normal variations in the number of sets of chromosomes Diploid animals sometimes produce tissues that are polyploid ◦ Termed endopolyploidy Liver cells can be triploid, tetraploid or even octaploid (8n) Polytene chromosomes of insects provide an unusual example of natural variation in ploidy 92 Polytene Chromosomes 1 Occur mainly in the salivary glands of Drosophila and a few other insects Polytene chromosomes have facilitated the study of the organization and functioning of interphase chromosomes ◦ Polytene chromosomes are easily seen with a microscope even in interphase ◦ Each has a characteristic banding pattern 93 Polytene Chromosomes 2 Chromosomes undergo repeated rounds of chromosome replication without cellular division ◦ In Drosophila, pairs of chromosomes double approximately nine times (29 = 512) These doublings produce a bundle of chromosomes that lie together in a parallel fashion ◦ This bundle is termed a polytene chromosome ◦ Central point where the chromosomes aggregate is the chromocenter 95 96  Polyploidy A condition in which the cells of an organism have more than two paired sets of chromosomes Common in plants ◦ 30 to 35% of ferns and flowering plants are polyploid ◦ Many fruits and grains are polyploids In many instances, polyploid strains of plants display outstanding agricultural characteristics ◦ They are often larger in size and more robust 97 Polyploids having an odd number of chromosome sets are usually sterile ◦ These plants produce highly aneuploid gametes Example: In a triploid organism there is an unequal separation of homologous chromosomes (three each) during anaphase I Each cell receives one copy of some chromosomes and two copies of other chromosomes 99 Sterility in Agriculture Although sterility is generally a detrimental trait it can be agriculturally desirable ◦ Seedless fruit Watermelons and bananas ◦ Triploid varieties ◦ Propagated by cuttings ◦ Seedless flowers Marigold flowering plants ◦ Triploid varieties ◦ Keep blooming 101 Mechanisms That Produce Variation in Chromosome Number How meiotic and mitotic nondisjunction occur and their possible phenotypic consequences Autopolyploidy, alloploidy, and allopolyploidy How colchicine is used to produce polyploid species 102 Chromosome Number Variation There are three natural mechanisms by which the chromosome number of a species can vary 1. Meiotic nondisjunction 2. Mitotic nondisjunction 3. Alloploidy (interspecies crosses) 103 Meiotic Nondisjunction Nondisjunction – Failure of chromosomes to segregate properly during anaphase Meiotic nondisjunction can produce cells that have too many or too few chromosomes ◦ If such a gamete participates in fertilization, the zygote will have an abnormal number of chromosomes ◦ Nondisjunction can occur in meiosis I ◦ Nonduisjunction can occur in meiosis II 104 All four gametes are abnormal 105  50 % 50 % Normal Abnormal gametes gametes 106 Complete Nondisjunction In rare cases, all the chromosomes can undergo nondisjunction and migrate to one daughter cell This is termed complete nondisjunction ◦ It results in one diploid cell and one without chromosomes ◦ The chromosome-less cell is nonviable ◦ The diploid cell can participate in fertilization with a normal gamete, yielding a triploid individual 107 Mitotic Nondisjunction Occurs after fertilization ◦ Mitotic nondisjunction Sister chromatids separate improperly ◦ Leads to trisomic and monosomic daughter cells ◦ Chromosome loss One of the sister chromatids does not migrate to a pole and is degraded if not included in reformed nucleus ◦ Leads to normal and monosomic daughter cells 108  This cell will be This cell will be monosomic trisomic This cell will be monosomic This cell will be Will be degraded if left outside of the nucleus normal when nuclear envelope reforms 110 Autopolyploidy Complete nondisjunction can produce an individual with one or more extra sets of chromosomes 112 Allopolyploidy A type of polyploidy that occurs when a polyploid offspring is derived from two distinct parental species An allotetraploid: Two complete sets of chromosomes from two different species 114 Experimental Treatments Can Promote Polyploidy Polyploid and allopolyploid plants often exhibit desirable traits Can be induced by abrupt temperature changes or drugs ◦ The drug colchicine is commonly used to promote polyploidy ◦ Binds to tubulin (a protein found in the spindle apparatus), promoting nondisjunction 116