Genes, Chromosomes & Molecular Biology

• The goal is determining the functions of all genes and proteins.

Approaches to the goal

• Sequence many genomes for Structural Genomics, because DNA sequence data predicts sequence/function of products.
• Functional Genomics studies the functions of genetic information within genomes.
• Proteomics characterizes all genome-encoded proteins, is a structural biology element, and determines structure, functional domains, interactions, and expression levels.

Gene Expression

• A gene is a DNA nucleotide sequence on a chromosome, encoding a protein, tRNA, or rRNA molecule that regulates transcription of a sequence.
• It recognizes how genetic material (DNA) transfers to the final product (protein/polypeptide), and involves how these genes can be regulated for a desired product.

Gene Concept Overview

• Genes were known via Mendel's work as hereditary factors associated with specific characters that control traits.
• Experiments with labeled bacteriophages showed DNA as hereditary material.
• Known that genes are on chromosomes and the chromosomes are made of DNA
• Grifiths & co. postulated DNA as the genetic material
• Genes are connected with specific traits/characters

DNA Knowledge

• One-Gene-One Enzyme Theory postulated that genes control protein structure.
• The outcome: proteins are made with one of the two basic functions: structural contribution to properties of cell organizations or enzymes that catalyze cell chemical reactions.
• Genes don't work in isolation because their action is affected by the environment.
• Genetic information carried by a gene exists in just one of the two helix strands
• Thus, polynucleotide acts as a template for the complementary RNA molecule synthesis.
• Transcription = the first stage of gene expression
• The amount of gene information is unlimited.
• Not all genes would be used: rules that limit the number of sequence/sense.
• The number of genes varies in different organisms
• Bacteria cells may have ~4 million DNA bp or 4000Kbp
• If a single gene = ~1.1Kbp in length, a bacterium MAY have ~3500 genes if all DNA code for protein
• But not all DNA codes for proteins, therefore there may be 3000 or less coding genes, of which ONLY ~1000-2000 may be expressed at any particular time.
• Genes are discrete segments of DNA molecules and may be separated from each other by intergenic DNA
• They are arranged in different ways in different types of animals.
• Viruses have closely packed genes with very little intergenic DNA between them.
• In other organisms, they are spread out and have long intergenic spaces
• In higher organisms, genes are spread out and separated by very long intergenic regions.
• In humans, genes make up 30% of total DNA in the cell.
• Majority is spaced out, more or less randomly, but in some cases are grouped into distinct clusters.
• Individual genes in a cluster are unrelated however the clusters are made up of genes that contain related units of bio information (e.g. operon and multigene family)
• Biologic info is split into distinct units separated by DNA segments
• Segments containing bio information = EXONS
• Intervening segments = INTRONS
• These gene types are discontinuous/split/mosaic genes.
• Introns are common in higher organisms, many viruses and bacteria.
• Human genes for the cystic fibrosis transmembrane regulator = 24 exons and 23 introns.
• Causes cystic fibrosis when it does not function correctly.
• The cystic fibrosis transmembrane regulator gene split into 24exons and 23 introns.
• Exons scattered the gene length and are separated by introns ranging in size 2-35bp.
• Average exon length: 277bp, which is 2.4% of the gene.
• Genes that are very close together on the same chromosome tend to be inherited together

Biological Information Transfer

• Experiments show that in gene expression, RNA is an intermediate between DNA and protein.
• For each gene, RNA is transcribed for only one of the DNA strands (the template).
• Transcription and Translation = the two main stages in gene expression.
• DNA info can be copied into more DNA during replication or be translated into protein.
• Gene Expression/how genes are expressed (transcription, translation, etc.)
• Genotype + environment -> phenotype

Stages of Information Transfer:

• Replication
• Transcription
• Translation
• GENE EXPRESSION makes biological information in a gene available to the cell.
• Two steps: transcription and translation. Transcription: mRNA synthesis by reading the DNA (changing the code to mRNA).
• This occurs in the nucleus of eukaryotes Translation: polypeptide synthesis using genetic code (mRNA molecule)
• Changes the mRNA into using tRNA to interpret mRNA.
• Net outcome: protein is made up with one of the two basic functions.
• Proteins may be contributing structurally to cell property (muscle and hair protein), or May be an enzymes that catalyze one of the chemical reactions of the cells by coding two important factor-Biological structures and Biological functions
• F. Crick proposed biological information in gene DNA is transferred to RNA then protein
• This information flow was unidirectional, and proteins cannot direct DNA and RNA synth.
• These ideas created the Central Dogma of Genetics.
• Howard Temin and David Baltimore disproved the second part of Crickís proposition because certain viruses undergo reverse transcription (Reverse transcriptase).
• Eukaryotic DNA includes regions of coding and noncoding DNA
• Regions where DNA codes for proteins are exons, while regions that do not code are introns.
• Following mitosis/meiosis in eukaryotes, DNA recoils but regions remain relaxed for transcription, and these relaxed DNA areas are called euchromatin

Differences in gene expression

• Prokaryotes: no membrane bound organelles, are more primitive, contain only one circular chromosome, and are bacteria.
• Eukaryotes contain membrane bound organelles, whose chromosomes are paired not circular, and includes protists, fungi, plants, and animals.
• Prokaryote transcription and translation occur in the cytoplasm.
• Two step sequence eukaryotic transcription occurs inside the nucleus with premRNA containing exons and introns of gene.
• mRNA is only coding portions
• Translation occurs in the cytoplasm at ribosomes.

RNA

• Ribonucleic Acid (RNA) found all over the cell: nucleus, mitochondria, chloroplasts, ribosomes, and the soluble part of the cytoplasm.
• A single polynucleotide strand which may be looped/coiled.
• Contains ribose (not deoxyribose)
• Uses bases: adenine, guanine, cytosine, and uracil (not thymine).
• Types: Messenger RNA (mRNA) <5%, Ribosomal RNA (rRNA) Up to 80%, Transfer RNA (tRNA) About 15%
• In eukaryotes, includes small nuclear ribonucleoproteins (snRNP).
• Messenger RNA (mRNA) is a long molecule 1 million Daltons, ephemeral, difficult to isolate, and provides the plan for polypeptide chain
• Transfer RNA (tRNA) is short at 25,000 Daltons being soluble
• Consists of 61 different forms and has a specific anticodon as part of its structure.
• rRNA provides platform for protein synthesis
• tRNA translates the mRNA message into a polypeptide chain.
• Information must be transcribed from DNA for functions
• Polymerization is catalyzed by RNA polymerase and can initiate synthesis, uses rNTPS, requires a template, and unwinds/rewinds DNA
• Consisting of 5 subunits, 449 kd, with Core enzyme of 2 α, and 1 β holding enzyme together, and β’ binds to templates
• Holoenzyme= Core + sigma.

Transcription

• Begins at a promoter sequence and ends at a termination signal.
• Proceeds in to 5' to 3' direction.
• Forms a temporary DNA:RNA hybrid.
• Has complete processivity
• Regions that signal initiation are called promoters and there are two types: 35 promoters and the Pribnow box, whose numbers refer to distance from the start point
• The Polymerase surrounds the first base that is transcribed into RNA, start point.
• Sequences prior to the start point: upstream
• Sequences after the start point: downstream.
• Numbers of nucleotides increase going downstream
• A base before the start point: numbered -1
• Numbers negative increasing going downstream
• RNA polymerase recognizes a recognition sequence
• Transcriptions start on the antisense strand in the 3¹ to 5¹whilst RNA is formed in the opposite (5¹ to 3¹) direction.
• Melting is essential to expose the base of the template in order to direct transcript synthesis.
• Factors dissociate from the enzyme when the complex is formed
• Core enzymes bind initiation site and unwind the double helix.
• Core Enzymes that bind initially is called PROMOTER.
• Transcription moves process of transcription.
• Ribonucleotides are then based paired to template strand positions +1 and +2 which is catalyzed by enzyme

Transcript Elongation:

• RNA is synthesized in 5-3 direction using nucleotide triphosphate as an acting substrate
• Step involves one ribonucleotide to growing chain using a triphosphate
• The traverses the entire gene until sequences signal fall off.
• In termination, strands unwind unlike DNA strands
• A small region of the double helix is unwound causing a transcription BUBBLE where the elongation begin.
• The polymerized protein the duplex of the RNA dissociates from strand
• Rate of elongation isn't constant, and terminators signal locations of DNA side reaching
• Such sequences are important to prokaryote closeness and include Rho-independent and Rho-dependent termination.
• RNA transcribes forming hairpin loops rich in G-C
• Termination is dependent
• Binding of p creates Rho Utilization. It pulls RNA
• Unlike prokaryotes, the eukaryotic transcript isn't free to associate during completion.

Explore the intricate relationship between genes, chromosomes, and the nucleus within a cell. Understand the central dogma of molecular biology, genomics, and the roles of proteins. Learn about the influence of environmental factors on gene action.