BIO 121 – Molecular Cell Biology

Lecture Section I

A. Fundamental Cell Theory and Taxonomy

B. The Age of Genomic Taxonomy

C. Introduction to Multicellularity

D. Regulation of Transcription

E. Regulation of RNA and Protein Processing

F. Mutation and Repair

A. Fundamental Cell Theory and Taxonomy

1. How do we define ‘alive’?

2. How do we classify living organisms?

3. What are the universal features of cells?

1. What is Life? = What are Cells?

“The basic unit of life on Earth”

“All life is cellular”

“No entity not composed of cells is alive”

OK – So What is Life?(Our current definition of life is descriptive at best)

• Homeostasis: Regulation of the internal environment.

• Organization: Being structurally composed of one or more cells.

• Metabolism: Transformation of energy by converting chemicals and energy into cellular components (anabolism) and decomposing organic matter (catabolism).

• Growth: Maintenance of a higher rate of anabolism than catabolism.

• Adaptation: The ability to change over a period of time in response to the environment.

• Response to stimuli: from simple to complex.

• Reproduction: The ability to produce new individual organisms.

Fig. 16-3

Bacterial cell

Phage head

Tail sheath

Tail fiber

DNA

100

nm

1. What is Life?

• Viruses are most often considered replicators rather than forms of life.

• They have been described as "organisms at the edge of life", since they possess genes, evolve by natural selection, and replicate by creating multiple copies of themselves through self-assembly.

• However, viruses do not metabolize and require a host cell to make new products.

• Virus self-assembly within host cells has implications for the study of the origin of life, as it may support the hypothesis that life could have started as self-assembling organic molecules. (Wikipedia, 2010)

1. What is Life?• A prion is an infectious agent that causes bovine

spongiform encephalopathy and Creutzfeldt–Jakob disease.

• Prions are mis-folded proteins that propagate by entering a healthy organism and inducing normal forms of the protein to convert into the rogue form.

• Since the new prions can then go on to convert more proteins themselves, this triggers a chain reaction that produces large amounts of the prion form.

• Evolutionarily, prion replication has been shown to be subject to mutation and natural selection just like other forms of replication. (Wikipedia, 2010)

2. How do we classify living organisms?

• Domains: Archaea, Bacteria, Eukarya

• Old Version: Prokarya and Eukarya

(Kingdoms in Prokarya: Bacteria, Archaea)

• Kingdoms in Eukarya: Animalia, Plantae, Fungi, Protista

Figure 1-21 Molecular Biology of the Cell, Fifth Edition (© Garland Science 2008)

How do we classify living organisms?

• Phyla (Animalia):

  Chordata, Echinodermata, Arthropoda, Annelida, Mollusca, Nematoda, Platyhelminthes, Cnidaria, Porifera

• Sub-Phyla (Chordata): Vertebrata, Urochordata, Cephalochordata 

• Class (Vertebrata): Mammalia, Amphibia, Reptilia, Osteicthyes, Aves

• Order (Mammalia): Primates, Rodentia, Artiodactyla, Perissodactyla......

• Genus/Species (Primates):

Homo sapiens Pan troglodytes Macaca mulatta........

The closer together the taxa, the more similarities in the cells...

How many cellular species are there?

• Estimates range from 10-100 million species

• Only ~1.8 million have been identified and named

• Vertebrates 62,305

• Invertebrates 1,305,250

• Plants 321,212

• Fungi 100,000

• Estimated 5–10 million bacteria

3. The universal features of cells

a. Basic features of all cells

Plasma membrane: Selectively permeable lipid bilayer

Cytosol: Variably viscous internal fluid

Double-stranded DNA, RNA and proteins

Require an external source of energy

Metabolism: Build-up and break-down of molecules

Intracellular homeostasis

Ability to sense and respond to the environment

Copyright © 2008 Pearson Education, Inc., publishing as Pearson Benjamin Cummings

• Eukaryotic cells are generally much larger than prokaryotic cells (a few m, 15X larger, 1000X greater in volume)

• Eukaryotic cells are also characterized by having stuff prokaryotes don’t have:

– Membrane-bound organelles

– Compartmentalized function

– Multicellular organisms

– *Extracellular homeostasis

– *Cytoskeleton

Copyright © 2008 Pearson Education, Inc., publishing as Pearson Benjamin Cummings

b. Differences Between Euks and Proks

Prokaryoticexamples ofthese havebegun to blurthe lines!

Nucleoid

Plasma membrane

Cell wallBacterialchromosome

Prokaryotes: Cytoplasm bound by plasma membrane, no organelles No nucleus, DNA in an unbound region called the nucleoid

The plasma membrane is a selective barrier that allows sufficient passage of oxygen, nutrients, and waste to service the volume of every cell

Smooth ERRough ER

CYTOSKELETON:

Microfilaments

Intermediatefilaments

Microtubules

MitochondrionLysosome

Golgiapparatus

Plasma membrane

Nuclearenvelope

Typicalanimal

cell

Fig. 6-9b

Nuclear envelope Rough endoplasmic reticulum

Smooth endoplasmic reticulum

Central vacuole

Microfilaments

Intermediate filaments

Microtubules

CYTO-SKELETON

Chloroplast

PlasmodesmataWall of adjacent cell

Cell wall

Plasma membrane

Mitochondrion

Golgiapparatus

Plant and animal cells have most of the same organelles

Same for fungiand protists.....

B. The Age of Genomic Taxonomy

1. The Existing Genomes in the World Today

2. Mechanisms of Genomic Change

3. Gene Families

All cells store their hereditary info in double-stranded DNA, use RNA as an intermediate and protein as the principle functional molecules.

genome = a species’ DNA sequence

genotype = an individual within a species DNA sequence

• Nearly all of the cells in an individual have exactly the same DNA, only the sex cells are different by being randomly assorted haploids

traditional genetic phenotype = variations in visible ‘characters’

molecular phenotype = variations in protein sequence and function

cellular phenotype = cell specialization in multicellular organisms resulting from expression of a subset of the inherited genotype

1. The Existing Genomes in the World

a. The number of bases and the complexity of their organization vary far more than the number of genes

b. The conservation of critical functions and the base sequence of the genes that code for them show that all cells are related

c. These close structure and function relationships allow us to gain information about ourselves from a wide variety of organisms

a. Life’s genetic complexity is less than you think.

• The smallest genome: Mycoplasma genitalia has 477 genes (580,070 bases)

• The largest animal genome: The waterflea, Daphnia pulex, has 31,000 genes (200M bases)

• Arguably, the most complex: Homo sapiens has ~24,000 genes and ~3 billion bases in our 46 chromosomes.

• There is a fundamental ‘core’ of genes shared by ALL organisms of about 60 genes

Figure 1-37 Molecular Biology of the Cell, Fifth Edition (© Garland Science 2008)

• 477 genes compared to 31,000 = ~65X

• 580K bases compared to 3B = ~5,000X

• We have only a 40 fold increase in gene number over Mycoplasma genitalia!

• The big difference between eukarya and prokarya is in non-coding sequence

Figure 1-14a Molecular Biology of the Cell, Fifth Edition (© Garland Science 2008)

Mycoplasma genitalia

Mycoplasma genitalium: 477 genes, 580,070 basepairs

37 code for non-messenger RNAs

297 of the 440 that code for protein are ‘known’

153 processing of DNA, RNA, protein 71 involved in metabolism 33 involved in nutrient transport 29 involved in surface membrane 11 involved in cell division

143 remain unidentified

Prokaryotic cells

DNA is a single, circular double helix containing 106-107 base pairs and 1,000-6,000 genes

Eukaryotic cells

Chromosomes are linear DNA molecules (fruit flies have 10, we have 46, dogs 78, etc.)

Smallest chromosome in humans is 2 million bases, total DNA is 3.2 x 109 base pairs

As animals become more complex, not just more DNA in the nucleus, embedded controls become more complex

Our chromosomes are 50% protein!!

Plants have greater tendency to have [DNA] change

Eukaryotes also have genes from their mitochondria and/or chloroplasts

• Our mitochondria have a genome of 16,569 base pairs which codes for:

– 13 proteins

– 2 ribosomal RNA components

– 22 transfer RNAs

– The rest of the functional DNA components for mitochondrial function reside in the nucleus

Human Mitochondrial Genome

Several neuromusculardiseases are associatedwith mitochondrial mutations

Chloroplast Genome

Usually 110-120 genes

Some as high as 200

b. Gene Conservation

1. If the function is essential and unchanged, the structure (sequence) must be unchanged - because you’d die if you lost an essential function.

2. Example 1: All cells from bacteria to humans have extremely high sequence homology (structure) in the small ribosomal sub-unit because they use the same mechanism to express DNA (function).

3. Example 2: MADS-box family transcription factors have been found in every eukaryotic cell type on Earth.1. MEF-2, agrafens, deficiens, SRF

Figure 1-22 Molecular Biology of the Cell, Fifth Edition (© Garland Science 2008)

Conservation of sequence of the small ribosomal subunit

c. Model Organisms

1. Prokaryotic bacteria: Escherichia coli

2. Eukaryotic yeast: Saccharomyces cerivisiae

3. Eukaryotic protists: Tetrahymena pyriformis

4. Complex plants: Lycopersicon esculentum and Arabidopsis thaliana

5. Nematode worms and fruitflies: Caenorhabditis elegans and Drosophila melanogaster

6. Frogs, fish and birds: Xenopus laevis, Danio rerio, Gallus gallus, Coturnix coturnix

7. Rats and mouse: Rattus norvegicus and Mus musculus

8. Chimpanzees and monkeys: Pan troglodytes, Macaca mulatta (rhesus)

9. Humans: Homo sapiens

Figure 4-83 Molecular Biology of the Cell (© Garland Science 2008)

Figure 1-42a Molecular Biology of the Cell, Fifth Edition (© Garland Science 2008)

Saccharomycescerevisiae

Figure 1-46 Molecular Biology of the Cell, Fifth Edition (© Garland Science 2008)

Arabidopsisthaliana

Figure 1-47 Molecular Biology of the Cell, Fifth Edition (© Garland Science 2008)

Caenorhabditis elegans

Figure 1-48 Molecular Biology of the Cell, Fifth Edition (© Garland Science 2008)

Drosophilamelanogaster

Figure 1-50 Molecular Biology of the Cell, Fifth Edition (© Garland Science 2008)

Xenopustropicalis

Xenopuslaevis

Figure 1-53 Molecular Biology of the Cell, Fifth Edition (© Garland Science 2008)

2. Mechanisms of Genomic Change

a. Generation of new genes

b. Disruption or loss of existing genes

c. Size of a genome reflects the relative DNA addition and DNA loss

a. Generation of New Genes

1. New genes are generated from preexisting genes, no mechanism for new synthesis

2. Mutation - Accidents/mistakes followed by non-random survival

3. Gene Duplication - provides an important source of genetic novelty

4. DNA Shuffling - Reassortment during homologous recombination

5. Horizontal Transfer - Genes transferred between organisms, in the lab and in nature

6. Transposable elements

Figure 1-23 Molecular Biology of the Cell, Fifth Edition (© Garland Science 2008)

Intragenic Mutation

• Only 1 nucleotide pair per 1,000 is randomly changed in the germline per 1 million years

however.....

• This means that in a population of 10,000 every possible nucleotide substitution will have been tried out ~20 times in a million years

The evolution of sex has had a big impact on the next three.....

• Gene Duplication

• DNA Shuffling

• Horizontal Transfer

Gene Duplication

• The idea is that during meiosis in sexually reproducing organisms, crossover mutations can form multiple copies of an exon, a gene, a chromosome or the entire genome.

• The organism survived just fine with one copy so it only repairs damages to one copy, leaving the other to freely mutate.

• Once in a blue moon the mutated copy develops new, advantageous functions.

Figure 4-86 Molecular Biology of the Cell (© Garland Science 2008)

Gene Duplication

Figure 1-50 Molecular Biology of the Cell, Fifth Edition (© Garland Science 2008)

Xenopustropicalis

Xenopuslaevis

Wholegenome

duplication!

DNA Shuffling

• Pieces of different genes can be combined to form new genes with hybrid functions

• Incomplete or partial cross-over events

Figure 4-88 Molecular Biology of the Cell (© Garland Science 2008)

Exon Duplication

Horizontal Transfer

• Sexual reproduction is horizontal gene transfer within a species

• Viruses and plasmids can cause horizontal gene transfer across species

• Bacteria and yeast have multiple mechanisms

Transposable Elements

• “Parasitic” DNA sequences that colonize genomes and spread within them – many resemble viruses

• Can disrupt gene function, alter regulation

• Can create new genes by integration within and fusion with host gene segments

• Half of all human DNA has homology to known transposons

• 10% of currently occurring mouse mutations are transposon-driven

Figure 4-17 Molecular Biology of the Cell (© Garland Science 2008)

Table 5-3 Molecular Biology of the Cell (© Garland Science 2008)

b. Disruption or loss of existing genes

1. Mutation - Accidents/mistakes followed by non-random survival

2. DNA Shuffling - Reassortment during homologous recombination

3. Transposable elements

c. Size of a genome reflects the relative DNA addition and DNA loss

• Both loss and gain occur constantly

• The relative rates determine size

Figure 4-81 Molecular Biology of the Cell (© Garland Science 2008)

The Fugu – experienced a long period of low rates of DNA addition accompanied by normal rates of DNA loss

Figure 4-82 Molecular Biology of the Cell (© Garland Science 2008)

The huntington Gene

-High homology-Perfect exon alignment-Small introns-Little non-coding DNA

3. Gene Families

a. Gene duplications give rise to families of related genes in a single cell

b. More than 200 gene families are common to all three domains

c. The function of a gene can often be deduced from its sequence

Duplication and Divergence Give Rise to Related Genes – Very Common Events!

• 4873 protein-coding gene families have been identified in life on earth

– 264 are designated ‘ancient’ - in all lineages

– 63 are ubiquitous in all genomes analyzed

• Most of the shared ‘ancient’ families perform:

– replication and transcription

– translation and amino acid metabolism

Figure 1-24 Molecular Biology of the Cell, Fifth Edition (© Garland Science 2008)

Bacillus subtilis4014 Genes

47% in families

ABC Transporterfamily has 77 members in thissingle bacterium!

Expansion of gene families gives rise to function

• Transcription factors regulate gene expression and, thus, cell diversity and complexity in multicellular organisms

– bHLH TFs: 7 in yeast, 41 in worms, 84 in flies, 131 in humans

• Adhesion and signaling are far more critical in multicellular animals

– 2000 major plasmamembrane proteins in worms not present or in low numbers in yeast