What information does the gene possess? © 2016 Paul Billiet ODWS

The genetic approach Before the discovery of the importance of DNA, geneticists managed to determine what information is represented in the gene www.laskerfoundation.org/.../beadletatum.jpg © 2016 Paul Billiet ODWS

Beedle and Tatum (1940) The red bread mould Neurospora crassa A haploid organism, all the genes are expressed Different individuals can be separated because the spores form in long thin “pods” called ascs Its requirements are simple It grows quickly and produces results in a few days Neurospora crassa © 2016 Paul Billiet ODWS

Synthesis of enzymes Genes ? Wild type feeds on minimal medium (nitrates, sulphate, phosphate other inorganic salts, sugar and biotin) Synthesis of enzymes Genes ? 20 amino acids 9 vitamins other organic compounds © 2016 Paul Billiet ODWS

Producing mutant strains Alter the genes by mutation and you will alter the dietary requirements Wild type mould Mutagens (ionising radiation) Separate spores Which ones are mutant moulds? © 2016 Paul Billiet ODWS

Grow up on complete medium Finding the mutants Mutant moulds? Grow up on complete medium = Minimal medium PLUS Select for deficient strains which will grow on minimal medium plus ONE supplementary nutrient © 2016 Paul Billiet ODWS

Cross breeding strains Deficient strain (n) X Wild type (n) Fusion Zygospore (2n) Meiosis I and II 4 haploid nuclei Mitosis 8 haploid spores Separate spores and grow on complete medium Screen for nutrient requirements © 2016 Paul Billiet ODWS

Neurospora ascs containing haploid spores Mutant alleles are fluorescing © 2016 Paul Billiet ODWS

Results Four of the daughter spores gave rise to colonies that are like the deficient parent and four give colonies that are like the wild type parent The condition is hereditary It is caused by the mutation of a single gene © 2016 Paul Billiet ODWS

Therefore ONE GENE corresponds to ONE ENZYME Conclusion Enzymes are needed by this mould to synthesise vitamins and amino acids The production of these compounds is affected by mutation (damaged genetic material) The inheritance pattern of these defective enzymes is the same as that of single genes Therefore ONE GENE corresponds to ONE ENZYME

Therefore ONE GENE corresponds to ONE PROTEIN Generalising Enzymes are all proteins…. but not all proteins are enzymes Proteins are all affected by mutations in the same way Therefore ONE GENE corresponds to ONE PROTEIN © 2016 Paul Billiet ODWS

Therefore ONE GENE corresponds to ONE POLYPEPTIDE More precisely…. Some proteins are made of subunits (quaternary structure) e.g. Haemoglobin = 4 subunits (2 x α and 2 x β) Each subunit is independently affected by mutations Therefore ONE GENE corresponds to ONE POLYPEPTIDE © 2016 Paul Billiet ODWS

So a gene does not necessarily correspond to a polypeptide at all That’s not all… The hypothesis was established by 1962 there was still another refinement when gene expression was worked out Genes are transcribed to make RNA molecules mRNA molecules are translated into polypeptides but… not all RNA is mRNA, genes are also transcribed into tRNA molecules and rRNA molecules tRNA and rRNA are not translated (though they are used in the translation process) So a gene does not necessarily correspond to a polypeptide at all

It gets better (or worse!)… Immunoglobulins (antibodies) are made of a polypeptide with two domains (constant C and variable V) Two genes are needed to construct the polypeptide of an immunoglobulin, one for each domain The gene for the variable part is transcribed as pre-RNA This pre-RNA is then spliced in different places by spliceosomes to take out introns Different introns can be cut out to make different immunoglobulins from the same pre-RNA (alternative splicing) So a single gene can code for more than one protein © 2016 Paul Billiet ODWS

So what is a gene? It contains information That may code for a protein Or part of protein Or more than one protein Or RNA that does not translate into a protein The smallest physical unit of heredity encoding a molecular cell product (Penguin Dictionary of Biology). © 2016 Paul Billiet ODWS

The genome

The proteome Total set of proteins expressed by a cell, tissue or organ Information to construct a protein lies in genes Proteome ≠ genome Proteome > total genes in genome © 2016 Paul Billiet ODWS

From genome to proteome

Proteomics Study of the structure and function of proteins Human Proteome Project (HPP) Proteome and cancer research © 2016 Paul Billiet ODWS