DNA Evolution of knowledge about gene. Contains information about RNAs and proteins. Polynucleotide chains; Double stranded molecule;

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1 Evolution of knowledge about gene G. Mendel Hereditary factors W.Johannsen, 1909 G.W.Beadle, E.L.Tatum, 1945 Ingram, 1957 Actual concepts The gene hereditary unit located in chromosomes Hypotheses One gene one enzyme Hypotheses One gene one polypeptide The gene a sequence of DNA responsible for synthesis of macromolecules DNA Double stranded molecule; Polynucleotide chains; Contains information about RNAs and proteins. 1

2 Genetic code - DNA: Letters: A, G, C, T Words (one codon one amino acid): AAG - lys AGC - ser GCA - ala TTC phe TAG stop Phrases: 5' AAGAGCGCATTCTAG 3' lys ser ala phe stop Gene expression DNA mrna Protein 5 -ATTGCAAGATTACCATGT-3 Coding strand (untranscribed) 3 -TAACGTTCTAATGGTACA-5 Template strand (transcribed) Transcription (RNA polymerase) 5 -AUUGCAAGAUUACCAUGU-3 mrna Translation (trna, ribosomes) Leu Ala Arg Leu Pro Cys polypeptide Definition: Gene a fragment of polynucleotide chain of DNA which contains information about synthesis of: one polypeptide or several polypeptides or a functional RNA (rrna, trna, snrna) 2

3 Gene expression DNA mrna Protein rrna trna Classification: 1 st class genes encode 5,8S, 18S and 28S rrna; 2 nd class genes = structural encode mrna proteins; 3 rd class genes encode trna, 5S rrna. Gene s localization: Genes are located in DNA molecules; Genes consist of unique or repeated sequences; The genes from one molecule of DNA are separated by non-coding sequences spacers; There are no morphological borders each gene has only functional frontiers; The length of genes is different. 3

4 - The dimensions of human genome 3,164 x 10 9 bp - 2% of human genome encode for proteins - Number of genes Chromosome 1 contains 3380 genes - Chromosome Y contains 397 genes - Known function 50% human studied genes - Average length of gene 3000 bp - gene for β-globin 1,5 kb - gene for insulin 1,7 kb - gene for catalase 34 kb - gene for dystrophin - 2,4 Mb 4

5 Distribution of human genes by length Length, kb % Up to 10 23, , , , ,7 over 500 1,2 General structure of the transcription unit Central region coding region; Regulatory regions: proximal PROMOTER distal TERMINATOR ± Modulation sequences Functions: Molecular level control of polypeptide s synthesis functional protein Cellular level production of a normal cellular structure, metabolic chain, signaling chain, etc. Tissue level realization of a specific function (respiration, digestion, contraction, etc.) Organism level a specific trait (character) 5

6 Each cell contains a complete set of genes ( pairs of genes in all 46 molecules of DNA) Expression only 10% of all genes Permanent expression rrna genes trna genes House keeping genes Temporary expression depending on: tissue; ontogenetic period; cell cycle period; environment factors No expression pseudogenes The 2 nd class genes = structural (25% of nuclear DNA) Encode one or several polypeptides; Form monocistronic transcription units; Have a mosaic structure (exon/intron); Could be transcribed: In all cells (house keeping genes) Specific, depending on type of cell, age, factors; Are transcribed by RNA-polymerase II in a primary transcript pro-mrna; Are numerous, usually unique and heterogeneous; May form repetitive or non-repetitive families of genes; Present individual polymorphisms. Types of structural genes House keeping genes that encode indispensable cell proteins, active in all cells, in all periods of life; Tissue-specific genes that encode for proteins require for tissue specialization; Regulatory of ontogenesis; Dependent on environment factors. 6

7 Gene families Repetitive gene s family: a family of identical genes Non-repetitive gene s family: a family of genes of related structure and usually related function Peculiarities of the 2 nd class genes structure Initiation of transcription of the 2 nd class genes TAFs TFIID TFIIA TFIIB RNA-polymerase II TFIIF TFIIE 7

8 Regions of structural genes Promoter: TATA box (-20, -30) CAAT box (-70, - 100) Tissue-specific boxes( - ) Coding region: site +1, leader sequence exon 1 /intron/exon 2 /intron/.../exon n Terminator Site of polyadenilation Enhancers and silencers Promoter of the 2 nd class genes Controls the initiation of transcription: Activation of gene; Fixing of TF and RNA-polymerase II; Identification of (+1) and transcribed strand; Directing of RNA-polymerase II. Is not transcribed; In different genes promoters contain different specific boxes; Mutations in promoter may induce gene inactivation. Conservative boxes in structure of eukaryotic promoters Structure Sequence Position Length of bound DNA Transcription factors TATA-box TATAAAA p.n. TBP CAAT-box GGCCAATCT p.n. CTF/NF1 GC-box GGGCGG p.n. SP1 Octamer ATTTGCAT 20 p.n. Oct1, Oct2 8

9 Structure of promoter in structural genes in eukaryotes (2 nd class genes) Structure of promoter in structural genes in prokaryotes Interaction promoter-enhancer Exons Sequences of structural genes that encode polypeptide sequences; Are found in pro-mrna and mrna; Are transcribed and translated; Each exon encodes a region of protein; During alternative splicing some exons may be removed. 9

10 Introns Non-coding sequences of structural genes that separate exons; Are found in pro-mrna but not in mrna; Are transcribed but not translated; During splicing all introns are removed; Structure of terminator 10

11 Structure of transcription unit which contains rrna genes in eukaryotes (1 st class genes) Promoter ( ) Gene 18S Gene 5,8S Gene 28S Terminator n Organization of the 3 rd class genes Promoter A box (+ 55) and B box (+ 80) Genes for trna / rrna 5S Terminator n Structure of 5S rrna genes 11

12 Structure of operon in prokaryotes Human mitochondrial genom 12

13 Mobile genetic elements = TRANSPOSONS Type Main gene Type of transposition Exemples DNA transposons Retrovirus like transposons Transposase Reverstranscriptase (revertase) Retrotransposons Reverstranscriptase (revertase) Transposition through excision or replication Transposition through RNA produced on the basis of promoters located in LTR Transposition through RNA produced on the basis of neighbor promoters Tn (bacterial) P (Drosophila) THE-1 (human) Ty (yeast) L1 - LINEs Biological role of transposons Site-specific recombination Evolution of genomes Individual polymorphism of DNA Genome instability fragile sites in DNA Insertional mutagenesis Medical importance of transposons Changes in structure / function of structural genes genetic diseases (hemophilia B, epilepsy, retinita pigmentosum, etc) Variability of pathogen agents resistance to antibiotics and immune system 13