Genomes and the Resulting Proteomes
Focus:
This CREATE module was developed for an upper level seminar course focused on Genomics and Proteomics. The goal is to learn how we know what we know about genomes and the resulting proteomes. Many recent papers in genomics are composed of more discovery-based research that simply analyzes a wealth of data obtained through some high-throughput method. As I would like students to learn a range of techniques going from gene to protein, this module will focus on how we have learned more about neuropeptides, largely focusing on one particular gene, proopiomelanocortin (POMC).
Overview:
The POMC gene is well-known for its role in producing many bioactive peptides; some of these bioactive peptides play an important role in the hypothalamic-pituitary-adrenal (HPA) axis and others function as opioid peptides. Hence the biological relevance of POMC is immediately apparent. But how did we discover this gene? And how do we know that these peptides come from this gene? Even if we have DNA sequence for the gene, it is not immediately apparent that the resulting protein is processed into peptides. Hence the necessity for proteomics. The papers studied in this module focus on some of the highlights of POMC biology, from initial efforts to identify peptides by antibody-based antibodies, to the identification of the relevant genes in the genome of a popular model organism, to the use of exon sequencing to identify variants associated with obesity, and finally the investigation of brain peptides using mass spectrometry methods. Through this survey of one gene and its functions, an overview of a number of genomic/proteomic technologies can be addressed.
Applicable for Courses:
Courses focusing on Genomics and ProteomicsEducational Level:
Upper-levelRoadmap Objectives:
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- Article: Met- and Leu-enkephalin immunoreactivity in separate neurones. Larsson LI, Childers S, Snyder SH. (1979) Nature 282:407-10.
- Content area/major concepts: An introduction to how proteins and peptides have historically been identified and studied using antibodies. The purpose of this paper is to show students how things used to be done.
- Methods or technology used to obtain data: Radioimmunoassays, immunocytochemistry
- How the CREATE strategy was used:
- Biggest teaching challenge: Students will need to grasp how antibodies work (protein structure, binding), as well as the historical use of radioactivity in molecular biology.
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- Article: Identification of two proopiomelanocortin genes in zebrafish (Danio rerio). Gonzalez-Nunez V, Gonzalez-Sarmiento R, RodrÃguez RE. (2003) Molecular Brain Research 120:1-8.
- Content area/major concepts: An analysis of the POMC genes identified in the zebrafish, a popular model organism, including details on how they were discovered and annotated.
- Methods or technology used to obtain data: cDNA synthesis, PCR, gene cloning, DNA sequencing, basic bioinformatics
- How the CREATE strategy was used:
- Biggest teaching challenge: Concepts to understand include: homology and gene duplication, the usefulness of model organisms, and cloning techniques. This paper is just on the cusp of the genomics era, when it was still a little harder to identify and annotate a gene and its products.
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- Article: Identification of POMC exonic variants associated with substance dependence and body mass index. Wang F, Gelernter J, Kranzler HR, Zhang H. (2012) PLoS ONE 7(9):e45300. doi:10.1371/journal.pone.0045300
- Content area/major concepts: Sequencing of all POMC exons in a large cohort of subjects was performed and identified gene variants correlated with substance dependence and BMI. An example of how genomic sequencing may be used towards medical analysis and treatment.
- Methods or technology used to obtain data: Sanger DNA sequencing, statistical and bioinformatics analyses
- How the CREATE strategy was used:
- Biggest teaching challenge: The statistical analyses used in this paper are fairly complex.
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- Article: Quantitative peptidomics reveal brain peptide signatures of behavior. Brockmann A, Annangudi SP, Richmond TA, Ament SA, Xie F, Southey BR, Rodriguez-Zas SR, Robinson GE, Sweedler JV. (2009) Proc Natl Acad Sci U S A. 106(7):2383-8.
- Content area/major concepts: While this paper does not specifically identify POMC peptides in the honey bee (does POMC exist in the honey bee?), it does show the power of mass spectrometry to identify the wealth of information found in the proteome of an organism, things that cannot be known by simply studying the genome.
- Methods or technology used to obtain data: Peptide extraction, mass spectrometry
- How the CREATE strategy was used:
- Biggest teaching challenge: Mass spectrometry is a complex technique, but becoming more prominent in everyday science. It may take some time for students to understand this technique.