DNA and Chromosomes Review

Historical Milestones in Genetic Research

• 1859 - Charles Darwin: Proposed the theory of evolution by natural selection.

• 1865 - Gregor Mendel: Established that heredity is transmitted in discrete units.

• 1869 - Frederick Miescher: Isolated DNA from cells for the first time, originally naming the substance "nuclein."

• 1928 - Frederick Griffith: Discovered the phenomenon of transformation in bacteria.

  • Experiment: Used Streptococcus pneumoniae, which exists in two forms: pathogenic S strain (lethal, encased in a slimy polysaccharide capsule) and harmless R strain (lacks the protective coat).

  • Finding: Heat-killed infectious (S) bacteria can transform live harmless (R) bacteria into live pathogens.

• 1944 - Oswald Avery, Colin MacLeod, and Maclyn McCarty: Provided the first evidence that DNA is the genetic material.

  • Experiment: Worked with the S strain of pneumococci to show that the "transforming principle" changing R strain into S strain was DNA.

• 1952 - Alfred Hershey and Martha Chase ("The Waring Blender Experiment"): Definitively showed that genes are made of DNA.

  • Method: Labeled viral DNA with radioactive phosphorous ($^{32}P$) and viral proteins with radioactive sulfur ($^{35}S$).

  • Result: Only the radioactive DNA entered the bacteria to direct the production of more viruses.

• 1953 - Rosalind Franklin and Maurice Wilkins: Utilized X-ray crystallography to determine the double-helix structure of DNA, identifying that the phosphate backbone is on the exterior.

• 1953 - James Watson, Francis Crick, and Maurice Wilkins: Constructed a model of DNA showing the complementary base pairing and the antiparallel nature of the two strands.

• 2003: Completion of the Human Genome Sequencing project.

The Molecular Structure of DNA

• Component Chains: A DNA molecule consists of two complementary chains of nucleotides.

• Backbone: Two sugar-phosphate backbones run in an antiparallel orientation.

• Nitrogenous Bases: Project inward from the backbones.

  • Adenine (A) and Thymine (T): Form two hydrogen bonds ($A=T$).

  • Guanine (G) and Cytosine (C): Form three hydrogen bonds ($G \equiv C$).

• Structural Features:

  • Major and Minor Grooves: These grooves are lined by potential hydrogen bond donors and acceptors (highlighted as yellow), which provide specific interaction sites for proteins and other molecules.

  • Physical Dimensions: The distance between each base pair in the double-strand helix is approximately $0.34\,nm$.

Genomic Composition and Human DNA

• Human Genome Size: Approximately $3200 \times 10^6$ (3.2 billion) nucleotide pairs per single strand genome ($3 \times 10^9$ base pairs).

• Functional Distribution:

  • Protein-Coding/Functional RNA: Only about $1.5\%$ of human DNA encodes proteins and functional RNAs.

  • Regulatory and Introns: The majority consists of regulatory sequences that control gene expression and non-coding introns.

  • Mobile DNA Elements: Approximately $45\%$ of human DNA is derived from mobile elements that have contributed to genome evolution.

Eukaryotic Chromosome Structure and Karyotyping

• Packaging: DNA is packaged into multiple chromosomes. In a single human cell, the total length of DNA measures approximately $2\,m$ but is contained within a nucleus with a diameter less than $10\,\mu m$.

• Spectral Karyotyping: Multi-color fluorescence in situ hybridization (FISH) is used to "paint" chromosomes. DNA is treated to partially separate strands (denature), allowing single-stranded labeled DNA probes to base-pair with target sequences.

• Chromosomal Abnormalities:

  • Reciprocal Translocation: A segment of one chromosome is swapped with a segment from another chromosome.

  • Implications: While some occur naturally during evolution, they are often linked to aneuploidy, infertility, or cancer (e.g., translocation $t(4;6)(q22;q15)$ is a recurrent alteration in prostate cancer).

• Genomic Divergence Example: Closely related species can have different chromosome numbers without significant differences in gene count.

  • Indian Muntjac: $2n = 7$ (males) / $6$ (females).

  • Chinese Muntjac: $2n = 23$.

The Cell Cycle and Chromosome Dynamics

• Interphase:

  • The DNA replicates at specialized "origins of replication."

  • The cell expresses many of its genes.

  • Chromosomes remain in non-overlapping territories within the nucleus.

  • Nucleolus: The most prominent interphase structure, containing over 400 copies of genes for ribosomal RNAs (rRNAs).

• M Phase (Mitosis):

  • Chromosomes condense into a highly compact form.

  • Two chromatids are tightly joined together (mitotic chromosome).

  • Chromosomes attach to the mitotic spindle at the centromere.

  • One complete set of chromosomes is pulled to each end of the cell.

  • A nuclear envelope forms around each set, and the cell divides into two daughters.

Nucleosomes and Levels of DNA Condensation

• Nucleosomes: The basic unit of eukaryotic chromosome structure.

  • Structure: DNA wrapped around a protein core called a histone octamer (composed of eight histone molecules).

  • Length: Approximately 147 nucleotide pairs wrap around the octamer nearly twice.

• Chromatin-Remodeling Complexes: Protein complexes that use ATP hydrolysis to loosen nucleosomal DNA, making specific sequences either accessible or hidden from DNA-binding proteins.

  • Example: A complex purified from yeast contains 15 subunits.

  • Note: Many of these are inactivated during mitosis to maintain compaction.

• Histone and Non-histone Proteins: Higher-order chromatin folding is organized by these proteins to manage the 280 Mb nucleotides per chromosome (human average).

Regulation of Chromosome Structure

• Chromatin Categories:

  • Euchromatin: Light-staining (whitish) areas; includes most transcriptionally active regions. Characterized by high levels of histone N-terminal tail acetylation.

  • Heterochromatin: Dark-staining areas; highly condensed. Includes centromeres, telomeres, and transcriptionally inactive genes.

• Histone Tail Modifications (Epigenetics): The pattern of modifications on histone tails (e.g., Histone H3) determines how chromatin is handled.

  • Acetyl groups (Ac): Added by Histone Acetyltransferases (HAT) using Acetyl CoA. This results in DNA exposure and recruitment of Transcriptional Activators (TA).

  • Methyl groups (M): Trimethylation at Lysine 9 (K9) recruits heterochromatin-specific proteins.

  • Phosphate groups (P).

• Transcriptional Repression: Transcriptional Repressor (TR) complexes interact with Histone Deacetylases (HDAC) to remove acetyl groups, strengthening electrostatic interactions between DNA and histones.

• Barriers: Acetylation of K9 blocks methylation at the same site, preventing the expansion of heterochromatin into euchromatin.

Health and Biological Implications

• Neurodevelopmental Disorders: Imbalances in histone acetylation and methylation can have detrimental effects.

• Rubinstein-Taybi Syndrome: Caused by a deficiency in a histone acetyltransferase (HAT) due to mutations in lysine acetyltransferase genes.

• Beta-Globin expression: The gene for beta-globin is near a heterochromatin region. If the barrier DNA is deleted, heterochromatin can invade the gene region, causing severe anemia.

• X-Chromosome Inactivation (Lyonization): Named after Mary F. Lyon (1961). In mammalian females, one of the two X chromosomes is randomly inactivated via heterochromatin formation in embryonic cells.

  • Example: The coat color of calico cats results from random X-inactivation in skin cells during development.

Questions & Discussion

• Where are viral protein and viral DNA found in the Hershey-Chase experiment? (Referring to Figure labels A and B): Viral DNA is found in the pellet (bacteria), and viral protein is found in the supernatant/extracellular space.

• What is not commonly found in heterochromatin? Answer: Chromosomal regions carrying genes that encode ribosomal proteins (these are typically in the nucleolus or euchromatin to allow high transcription).

• What stops heterochromatin spread? Answer: Encounters with a barrier like Histone H3 acetylation on lysine 9 (K9).

• Which modification always increases transcription? Answer: Histone acetyltransferase adding acetyl groups to histone tails (loosens chromatin).

• Total length of DNA calculation: $\text{Total Length} = (3 \times 10^9\text{ bp}) \times (0.34 \times 10^{-9}\text{ m/bp}) = 1.02\,m$ per haploid set, resulting in roughly $2\,m$ for a diploid human cell.