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1、Ig gene rearrangement in antibody producing cells -Southern blot analysis non lymphoid cell:V,C in the distant fragments mature B cell:V,C in the same fragments V region of light chain:V gene segment (VL protein)(VL gene)J gene segment (JL gene)V region of heavy chain:V gene segment (VH protein)(VH
2、gene)D gene segment (DH gene)J gene segment (JH gene)第1页/共26页第2页/共26页第3页/共26页-The numbers of functional gene segments for the V region of human heavy and light chains pseudogenes :not functional protein-The germline organization of the Ig genes gene family 第4页/共26页第5页/共26页第6页/共26页Rearrangement of V,
3、D,J gene segments -DNA rearrangements are guided by conserved (recombination signal sequence,RSS)heptamer:contiguous with the coding sequence space :12 or 23 nucleotides long nonamer :second conserved sequence -coding sequence-heptamer-space-nonamer (12/23)-12/23 rule -exception:D-D joining(fusion)i
4、n human 5%looping out and deletion:more common no deletion 第7页/共26页第8页/共26页第9页/共26页第10页/共26页第11页/共26页Enzymatic steps in the gene rearrangement -RAG(Recombination Activating Genes RAG-1,RAG-2)protein complexes bind to RSSs make single-strand DNA breaks at sites 5 of each RSS -RAG are only expressed i
5、n developing lymphocytes -Other ubiquitously expressed DNA-modifying proteins DNA ligase IV:joins the processed ends together with XRCC4 DNA-dependent protein kinase(DNA-PK)Ku:a heterodimer Ku70:Ku80 Ku+DNA-PK+Artemis complex :a protein involved in the opening of covalently closed DNA hairpin struct
6、ures -terminal deoxynucleotidyl transferase(TdT)randomly adding nucleotides -RAG genes,DNA-PK,TdT gene deficiency Severe combined immune deficiency(SCID)mouseSignal joint and coding joint第12页/共26页第13页/共26页第14页/共26页第15页/共26页 The diversity of the Ig repertoire:four main process -combinatorial diversit
7、y multiple gene segments different combination different combination of H,L chain -Junctional diversity -Somatic hypermutationCombinatorial diversity kappa chain 40 5=200 lambda chain 30 4=120 H chain 40 25 6=6000 -Combinatorial diversity is likely to be less from the theoretical calculation320106第1
8、6页/共26页Junctional diversity -The diversity of CDR3 in H,L chains is significantly increased by the addition and -The added nucleotide are known as P-nucleotides and N-nucleotides P-nucleotides:palindromic sequences at the end of the gene segment N-nucleotides:non template-encoded -Addition of P,N nu
9、cleotide is a random process Productive or nonproductive rearrangement第17页/共26页第18页/共26页第19页/共26页第20页/共26页TCR gene segments and rearrangement -The same enzyme for Ig gene rearrangement can be used for TCR gene rearrangement -The numbers of human TCR gene segment TCR gene rearrangement第21页/共26页第22页/共26页第23页/共26页第24页/共26页第25页/共26页感谢您的观看！第26页/共26页