Course Identification

An Overview of Molecular Cell Biology- Adapted for young in biology

Dan Michael

Part 1: The basic chemistry of nucleic acids

From atoms to covalent, hydrogen and ionic bonds; other intra- and inter-molecular forces.  The major macro-molecules in the cell. The nucleotides in the DNA, the chemistry of DNA, the basic structure of DNA. RNA is derived from DNA, the nucleotides in the RNA. RNA structure is sequence dependent. Preview: The central dogma in molecular biology.

Part 2: The basic chemistry of proteins, enzymes and antibodies

Proteins are translated from RNA. Properties of amino acids, the peptide bond. X-ray crystallography, Cryo-EM. Many proteins are enzymes but certain RNAs can be enzymes as well. From the primary to the quarterly structure of proteins. The dynamic nature of proteins structure. The role of covalent modifications. The role of ionization which is determined by pH. Ligands and ligand-gated channels, quarterly-enforced structures. Antibodies, from origin to physiological roles. Antibodies as tools to study molecular cell biology. Indirect immunocytochemistry.

Part 3: Cellular mechanisms underlying heredity

The hallmarks and complexity of life. Cell to cell inheritance of traits. From cells to organs to organisms. Evolution of species. Trans-generational heredity. Chromosome theory and chromosomes, nucleosomes. Heredity through cell division: ploidity, mitosis. Hereditary through sexual reproduction: meiosis. The extra combinatorial power of homologous recombination in offspring’s diversity.

Part 4: Establishing the central dogma in molecular biology

Historical millstones in genetics research: from traditional genetics to Mendel’s laws of modern genetics. Genes, genotype-phenotype relationships, dominant and recessive alleles. Homozygosity and heterozygosity. From biochemical millstones in macromolecules research to the experiment by Avery, McLeod and McCarthy. Deciphering the structure of DNA. DNA replication: from building blocks to semi-conservative mode of replication. From an apparent paradox to the role of RNA in the pathway from DNA to a protein. Definition of transcription. From nucleotides to the genetic code. Translation: the ribosome, the tRNAs, aminoacyl-tRNA synthases. The ribosome as a polymerase, the translation cycle at the A, P and E sites of the ribosome. Genetics disorders stemming from defects in translation. The energy requirements during translation and proof-reading during translation.

Part 5: Regulation of gene expression

From chromosomes to chromatin organization. Histones, nucleosomes. From nucleosomes to chromosome territories. From traits (sickle cell anemia) to a gene and its alleles. The b-globin gene. Regulation of transcription. Transcription factors and their binding sites. Promotors, enhancers, the pre-initiation complex (PIC). The mediator. The role of histone modifications. Rate limiting steps in the transcriptional cycle. RNA processing: Capping, splicing and polyadenylation. Regulation of translation. Heterogeneous ribosomes. Post translation modifications.

Part 6: Basic recombinant DNA

The restriction phenomenon between bacteria and bacteriophages. Restriction enzymes and DNA analysis in gels. DNA cloning in plasmids and bacteriophages. Modern biotechnology: human insulin produced in bacteria. Other transgenic organisms. DNA libraries. Basic properties of cancer cells and cloning an oncogene using DNA libraries. cDNA and DNA sequencing by the Sanger method. Open reading frames. Sequence comparisons lead to functional insights. The polymerase chain reaction (PCR). Using PCR to generate expression vectors. PCR in forensic medicine.

Part 7: Genomes, epigenomes, transcriptomes, proteomes and interactomes

Genome sequencing using deep sequencing can identify cancer causing mutations. Comparisons and the evolution of genomes. Sequence comparisons. Gene duplications enables genetic novelty. Paralogs and orthologues. Phylogenetic trees. Size of genomes and their functional annotation. The human ENCODE project. Mutations in the DNA that control gene expression. Sequencing hundreds of thousands of human genomes: the variation between genomes (“The Variome”). Most variants have weak effect on the phenotype. Epigenetics and epigenomes: Covalent modification on DNA and on histones. Writers and readers of histone modifications. The effect of chromatin structure on the contribution of the DNA to gene expression. Cell to cell epigenetic inheritance. The non-coding genome. Regulation of gene expression by non-coding RNA (ncRNA). Transcriptomes can be determined by high-throughput methods such as RNA-seq. Specialized cell types and cells under defined physiological conditions have defined transcriptomes. The entire collection of proteins (the proteome). The proteasome is a major a cellular protein degradation system that shapes the proteome. Proteins and their variants can be studied biochemically to appreciate the many steps from genomes to proteomes and interactomes. The generation of functional diversity: From the genome, transcriptomes, proteomes, interactomes to the phenotypes.

Part 8: From recombinant DNA to reverse genetics

From a protein to a gene and its functions: The most studied gene, p53, as a case study for functional biology using reverse genetics. Overexpression of the gene product and its sequence characterization in human tumors provide the first functional clues. The need to knock-out and knock down the expression of a gene of interest to decipher gene function. The “acquired immune system” in bacteria provide the tools for efficient gene knockout as well as for genome editing by in-large: using the CRISPR system-derived tools for igniting efficient genome editing. Using CRISPR-derived tools to modulate gene expression. Knocking down expression requires RNA interference (RNAi) tools. From the effect of synthetic siRNA to the discovery miRs, their mode of endogenous production and their roles in regulation of gene expression. The role of a regulator of gene expression at the DNA level can be studied using ChIP-seq (Chromatin immunoprecipitation coupled to deep-sequencing). Detailed study of the DNA binding of p53 and other SSFs using experimental and computational aspects.

Part 9: Forward genetics

Defining modern forward genetics. Forward genetics to study cancer. Using miR overexpression in forward genetics to identify genes that control metastasis. CRISPRa (Cas9 fused to a transcription activator) as a tool in forward genetics to study cell movement and metastasis. From forward to reverse genetics and single cell analysis. CRISPR knock out-assisted forward genetics in viral infection studies and the corroboration of these studies by reverse genetics.

Part 10: Cells and their organelles under homeostasis and disease

A prototypic animal cell and a prototypic neuron. Biochemical fractionation, protein mass-spectrometry and immunohistochemistry to study cell structure and function. The plasma membrane.  The major lipid in a membrane bi-layer vary. Membranal proteins: single-pass and multi-pass. The neuronal membrane. Membranes are connected to the cytoskeleton. Function and dynamics of the cytoskeleton: Actin, myosin and microtubules. Intermediate filaments, cell polarity and coordination of the cytoskeleton. Cell junctions and the extracellular matrix. The nucleus and its membrane (the nuclear envelope). The other organelles in homeostasis and stress. The mitochondria allow energy conservation and metabolic compartmentation. Mitochondrial stress. Proteins move between organelles. Intracellular organization and protein sorting. The secretory pathway begins at the endoplasmic reticulum (ER). Translation on the ER and translocation of nascent proteins into the ER. The Golgi apparatus. Secretory vesicles. ER stress and pro-inflammatory responses. The lysosome and autophagy under homeostasis and stress.

Part 11: Cell signaling and molecular networks

Principles of cell signaling. Extracellular signaling molecules and their receptors. Endocrine, paracrine and autocrine signaling systems. Signals activate short or long intracellular pathways.  Specificity, robustness and amplification of signals. Signaling through G-Protein-coupled receptors. Some G proteins regulate the production of Cyclic AMP. The adrenaline-ignited pathway that orchestrates the flight of fight response. Receptor tyrosine kinases ignite intracellular signaling. The Ras-Raf-MAPK pathway. The signaling generated by ER stress. Down-regulation of cellular signaling. Cellular networks.

Part 12: From cell differentiation to development

Molecular genetic mechanisms that create and maintain specialized cell types. Combinatorial gene control create many different cell types. Combination of master transcription regulators specify cell types by controlling expression of many genes. Stable patterns of gene expression can be transmitted to daughter cells. Overview of development. The developmental potential of cells. Spatial patterning. Cell memory and complex tissue patterns. Lateral inhibition can generate complex tissue patterns. Developmental biology provides insights into disease and tissue maintenance. Morphogenesis and growth. The proliferation, death and size of cells determine organ and organism size. Hormones coordinate growth through the body. The dynamics of gene expression in embryogenesis at a single cell resolution. Methods and applications for single cell and spatial multi-omics. Stem cells and tissue homeostasis. Stem cells self-renew and produce differentiated cells. Self-renewal in the intestine and the body surface. Stem cell niche. Tissue aging.

Part 13: Frontiers in functional genomics and functional biology

Topics will vary according to most up-to-date literature.

Upon successful completion of this course, students aim to:

1. Discuss the chemistry and dynamics of cellular macromolecules as well as the modules defined in the central dogma of molecular biology.
2. Discuss the regulation of gene expression, recombinant DNA in research and biotechnology, genomes, genome editing, epigenomes, transcriptomes, proteomes, and interactomes.
3. Discuss cells and their organelles in homeostasis and disease, cell signaling, developmental biology, and frontiers in functional biology.
4. Appreciate an inventor’s mind that led to the emergence of a research breakthrough, often recognized by receiving a Nobel Prize, as well as the research methodologies used, and differentiate between methodologies that promote forward and reverse genetics.
5. Acquire the ability to raise hypotheses based on prior scientific knowledge in molecular cell biology.
6. Acquire skills to design experiments in molecular biology in order to address a hypothesis of interest, thus better understanding the process involved in generating relevant hypothesis-driven molecular data.
7. Interpret the data in recently published selective articles in molecular cell biology as well as critically interpret the data using original ideas to corroborate the research findings.
8. Given the student’s own discipline of origin, define problems of self-interest in molecular cell biology or medicine that may require a solution and then search for the most unexpected research directions to solve such challenges while emphasizing the need for newly acquired molecular data.