Survey
* Your assessment is very important for improving the workof artificial intelligence, which forms the content of this project
* Your assessment is very important for improving the workof artificial intelligence, which forms the content of this project
2012-05-09 Genetic recombination Although genetic stability is essential for survival of individuals, genetic variation is essential for long term survival in a changing environment Genetic recombination by breaking and joining of DNA sequences Genetic recombination occurs upon formation of germ cells, during DNA repair and transposition and integration of DNA sequences Two main variants * General (homologous) recombination occurs between homologous sequences, e.g. during meiosis, DNA repair RecA dependent or similar protein Driven by DNA homology * Site-specific recombination occurs between non-homologous sequences of limitited homology, specific 15 bp sequences recognized by specific enzymes, e.g. integrase Driven by DNA-protein interactions Transposition requires special sequence on the donor DNA and transposase Illegitimate recombination (very rare) no specific sequences or enzymes required 1 2012-05-09 General recombination An important event ocurring in viruses, bacteria and eukaryotes Exchange can occur anywhere in the homologous region Strand exchange resulting in a cross-over No nucleotide sequence loss/gain at the site of cross-over. Exact! Recombination events occur at heteroduplex joints At the joint some mismatches can happen Sequences are not necessarily exactly the same on either side of the joint 2 2012-05-09 Recombination is initiated by a nick/break event RecBCD protein of E.coli, chemical agents and irradiation can produce nicks A random search for basepairing allows for rapid zippering and formation of a double helix slow 3 2012-05-09 Enzymes required for homologous recombination RecA (RAD51) forms triple strands by pairing ssDNA with homologous dsDNA RecBCD unwinds and rewinds DNA at specific recognition sequence RUV protein forms the four-stranded Holliday structure DNA polymerase copies DNA SSB keeps ssDNA straight DNA ligase seals DNA 1. In E. coli RecA protein binds to single-stranded DNA and can hold a single strand and a double helix together 4 2012-05-09 2. RecA directs branch migration in one direction Cross-strand exchange A. Nick and exchange of one strand B, C. Molecules are rotated (isomerized) D. Cut Robin Holliday, 1964 5 2012-05-09 Gene conversion can result in an unequal inheritance of gene parts Gene conversion is caused by mismatch repair on heteroduplex during recombination 6 2012-05-09 Mismatch proof-reading can prevent promiscuous recombination Site-specific recombination Guided by recombination enzyme No need for base-pairing (homology) between molecules Heteroduplex joint is only a few basepairs Integration and excision of bacteriophage lambda in the E. coli chromosome by the enzyme lambda integrase Transposition 7 2012-05-09 Site-specific recombination by λ phage in E.coli continued 8 2012-05-09 attP attB Int protein for integration into bacterial genome Xis protein for excision from bacterial genome Gene rearrangements in immunoglobulins and T-cell receptors accounts for high diversity of antibodies 300 V 4J numerous D 8C Somatic hypermutation due to Deaminations 3000 L x 5000 H 15 million comb. 9 2012-05-09 Transposable genetic elements (transposons) insert randomly in genome by recombination independent of RecA One contributing factor for spread of antibiotic resistance Transposition mechanism 10 2012-05-09 Effects of transposition • Target site (3-12 basepairs) is duplicated upon insertion of the transposon • Can induce homologous recombination between duplicated sequences causing restructuring of chromosome • Can inactivate genes upon insertion • Can cause deletions and inversion • Can activate neighbouring genes • Can transfer antibiotic resistance by transferring antibiotic resistance passenger gene • Retroviruses have similarities to transposons Alu sequences (300bp) are similar to transposable elements and common in the mammalian genome • • • • • Retrotransposon Do not encode protein 10 % of the human genome In non-coding regions Associated to some diseases Human chromosomes stained with Alu-probe (green) 11 2012-05-09 Position of Alu sequences is conserved in mammals and are found outside coding regions 12