《免疫学》（英文版） Chapter 05 Organization and Expression of Immunoglobulin Genes

8536d_ch05_105-1368/22/02 2: 47 EM Page 115 mac46 mac46: 1256_deN: 8536d: Goldsby et al./Immunology 5e Organization and Expression of Immunoglobulin Genes CHAPTER 5 115 both the coding joint and the signal joint(and intervening Another consequence of imprecise joining is that gene DNA)on the chromosome In the human K locus, about half segments may be joined out of phase, so that the triplet read- of the Vk gene segments are inverted with respect to Jk and ing frame for translation is not preserved. In such a nonpro- their joining is thus by inversion. ductive rearrangement, the resulting V] or VD] unit is likely IgGene Rearrangements May Be to contain numerous stop codons, which interrupt transla tion(Figure 5-9). When gene segments are joined in phase, Productive or Nonproduct the reading frame is maintained. In such a productive re- One of the striking features of gene-segment recombination arrangement, the resulting V] or VDJ unit can be translated is the diversity of the coding joints that are formed between in its entirety, yielding a complete antibody. If one allele rearranges nonproductively, a B cell may still breaks that initiate V-(D)-Jrearrangements are introduced be able to rearrange the other allele productively. If an in precisely at the junctions of signal sequences and coding se- phase rearranged heavy-chain and light-chain gene are not quences, the subsequent joining of the coding sequences is produced, the B cell dies by apoptosis. It is estimated that precise. Junctional diversity at the V-J and V-D-) coding only one in three attempts at VL-JL joining, and one in three joints is generated by a number of mechanisms: variation in subsequent attempts at VH-DHhH joining, are productive. As cutting of the hairpin to generate P-nucleotides, variation in a result, less than 1/9(11%)of the early-stage pre-B cells in trimming of the coding sequences, variation in N-nucleotide the bone marrow progress to maturity and leave the bone addition, and flexibility in joining the coding sequences. The marrow as mature immunocompetent B cells introduction of randomness in the joining process helps gen- Allelic Exclusion Ensures a Single erate antibody diversity by contributing to the hypervariabil- AntigenIc Specificity ity of the antigen-binding site. (This phenomenon is covered in more detail below in the section on generation of antibody B cells, like all somatic cells, are diploid and contain both ma- diversity. ternal and paternal chromosomes. Even though a B cell is ACTGTG GTGGACTAG □ GAGGATGCTCC CACAGTG Gene rearrangement Productive Glu Asp Ala Thr Arg Maternal H chain ① GAGGATGCGACTAGG Maternal GAGGATGGG AGG lu Asp Trp Thr Arg Maternal ③ GAGGATTGGACTAGG H chain Nonproductive arrangements ④ GAGGATGCGGACTAGG GAGGTGGACTAGG of allelic exclusion, the immunoglobulin avy.and light-chain genes of only one parental chromosome FIGURE 5.9 Junctional flexibility in the joining of immunoglobulin expressed per cell. This process ensures that B cells possess a single gene segments is illustrated with V and Jx. In-phase joining(arrows antigenic specificity. The allele selected for rearrangement is chosen 1, 2, and 3)generates a productive rearrangement, which can be randomly. Thus the expressed immunoglobulin may contain one ma- translated into protein. Out-of-phase joining(arrows 4 and 5) leads ternal and one paternal chain or both chains may derive from only to a nonproductive rearrangement that contains stop codons and is one parent. Only B cells and T cells exhibit allelic exclusion. Asterisks not translated into protein ()indicate the expressed allele

both the coding joint and the signal joint (and intervening DNA) on the chromosome. In the human  locus, about half of the V gene segments are inverted with respect to J and their joining is thus by inversion. Ig-Gene Rearrangements May Be Productive or Nonproductive One of the striking features of gene-segment recombination is the diversity of the coding joints that are formed between any two gene segments. Although the double-strand DNA breaks that initiate V-(D)-J rearrangements are introduced precisely at the junctions of signal sequences and coding se￾quences, the subsequent joining of the coding sequences is imprecise. Junctional diversity at the V-J and V-D-J coding joints is generated by a number of mechanisms: variation in cutting of the hairpin to generate P-nucleotides, variation in trimming of the coding sequences, variation in N-nucleotide addition, and flexibility in joining the coding sequences. The introduction of randomness in the joining process helps gen￾erate antibody diversity by contributing to the hypervariabil￾ity of the antigen-binding site. (This phenomenon is covered in more detail below in the section on generation of antibody diversity.) Another consequence of imprecise joining is that gene segments may be joined out of phase, so that the triplet read￾ing frame for translation is not preserved. In such a nonpro￾ductive rearrangement, the resulting VJ or VDJ unit is likely to contain numerous stop codons, which interrupt transla￾tion (Figure 5-9). When gene segments are joined in phase, the reading frame is maintained. In such a productive re￾arrangement, the resulting VJ or VDJ unit can be translated in its entirety, yielding a complete antibody. If one allele rearranges nonproductively, a B cell may still be able to rearrange the other allele productively. If an in￾phase rearranged heavy-chain and light-chain gene are not produced, the B cell dies by apoptosis. It is estimated that only one in three attempts at VL-JL joining, and one in three subsequent attempts at VH-DHJH joining, are productive. As a result, less than 1/9 (11%) of the early-stage pre-B cells in the bone marrow progress to maturity and leave the bone marrow as mature immunocompetent B cells. Allelic Exclusion Ensures a Single Antigenic Specificity B cells, like all somatic cells, are diploid and contain both ma￾ternal and paternal chromosomes. Even though a B cell is Organization and Expression of Immunoglobulin Genes CHAPTER 5 115 Jκ Vκ CACTGTG GTGGACTAGG GAGGATGCTCC CACAGTG RSS RSS 2 3 4 5 GAGGATGCGACTAGG Glu Asp Ala Thr Arg 1 GAGGATGGGACTAGG Glu Asp Gly Thr Arg GAGGATTGGACTAGG Glu Asp Trp Thr Arg Productive rearrangements 2 3 GAGGATGCGGAC TAG G Glu Asp Ala Asp Stop GAGGTGGAC TAG G Glu Val Asp Stop Nonproductive rearrangements 4 5 1 Joining flexibility FIGURE 5-9 Junctional flexibility in the joining of immunoglobulin gene segments is illustrated with V and J. In-phase joining (arrows 1, 2, and 3) generates a productive rearrangement, which can be translated into protein. Out-of-phase joining (arrows 4 and 5) leads to a nonproductive rearrangement that contains stop codons and is not translated into protein. κκ λλ HH κ λ H * * * * κ λ H Paternal chromosomes Gene rearrangement Maternal chromosomes Maternal H chain Maternal κ chain Maternal H chain Paternal λ chain FIGURE 5-10 Because of allelic exclusion, the immunoglobulin heavy- and light-chain genes of only one parental chromosome are expressed per cell. This process ensures that B cells possess a single antigenic specificity. The allele selected for rearrangement is chosen randomly. Thus the expressed immunoglobulin may contain one ma￾ternal and one paternal chain or both chains may derive from only one parent. Only B cells and T cells exhibit allelic exclusion. Asterisks (∗) indicate the expressed alleles. 8536d_ch05_105-136 8/22/02 2:47 PM Page 115 mac46 mac46:1256_des:8536d:Goldsby et al. / Immunology 5e:

8536d_ch05_105-1368/22/02 2: 47 EM Page 116 mac46 mac46: 1256_deN: 8536d: Goldsby et al./Immunology 5e 116 PART I Generation of B-Cell and T-Cell Response diploid, it expresses the rearranged heavy-chain genes from maturing B cell to turn off rearrangement of the other only one chromosome and the rearranged light-chain genes heavy-chain allele and to turn on rearrangement of the k from only one chromosome. The process by which this is ac- light-chain genes. If a productive k rearrangement occurs, complished, called allelic exclusion, ensures that functional light chains are produced and then pair with u heavy chains B cells never contain more than one VHDHH and one Vul to form a complete antibody molecule. The presence of this unit(Figure 5-10). This is, of course, essential for the antibody then turns off further light-chain rearrangement. antigenic specificity of the B cell, because the expression of If K rearrangement is nonproductive for both k alleles, re- both alleles would render the B cell multispecific. The phe- arrangement of the A-chain genes begins. If neither A allele nomenon of allelic exclusion suggests that once a productive rearranges productively, the B cell presumably ceases to ma VH-DH-JH rearrangement and a productive V1-JL rearrange- ture and soon dies by apoptosis. ment have occurred, the recombination machinery is turned Two studies with transgenic mice have supported the hy- off, so that the heavy- and light-chain genes on the homolo- pothesis that the protein products encoded by rearranged gous chromosomes are not expressed heavy-and light-chain genes regulate rearrangement of the G. D Yancopoulos and E W Alt have proposed a model to remaining alleles. In one study, transgenic mice carrying a ccount for allelic exclusion(Figure 5-11). They suggest that rearranged u heavy-chain transgene were prepared. The u once a productive rearrangement is attained, its encoded transgene product was expressed by a large percentage of the protein is expressed and the presence of this protein acts as B cells, and rearrangement of the endogenous immunoglob a signal to prevent further gene rearrangement. According ulin heavy-chain genes was blocked. Similarly, cells from a to their model, the presence of u heavy chains signals the transgenic mouse carrying a k light-chain transgene did not heavy chain inhibits μ+ K chains inhibit arrangement of u allele =2 arrangement of K allele = λ rearrangement μ+ chains inhibit Productive Productiv ◎=◎ allele=2 allele=1 allele#1 Progenitor allele #2 Productive allele=1 Nonproductive J, Productive allele #2 allele #2 ductive Nonproductive allele #2 allele #2 Cell death Cell death FICURE 5-11 Model to account for allelic exclusion. Heavy-chain either K or A rearrangement can proceed once a productive heavy enes rearrange first, and once a productive heavy-chain gene chain rearrangement has occurred. Formation of a complete rearrangement occurs, the u protein product prevents rearrange. immunoglobulin inhibits further light-chain gene rearrangement. If ment of the other heavy-chain allele and initiates light-chain gene a nonproductive rearrangement occurs for one allele, then the cell arrangement. In the mouse, rearrangement of K light-chain genes attempts rearrangement of the other allele. [Adapted from G. D precedes rearrangement of the A genes, as shown here. In humans, Yancopoulos and F. w. Alt, 1986, Annu. Rev. Immunol. 4: 339

diploid, it expresses the rearranged heavy-chain genes from only one chromosome and the rearranged light-chain genes from only one chromosome. The process by which this is ac￾complished, called allelic exclusion, ensures that functional B cells never contain more than one VHDHJH and one VLJL unit (Figure 5-10). This is, of course, essential for the antigenic specificity of the B cell, because the expression of both alleles would render the B cell multispecific. The phe￾nomenon of allelic exclusion suggests that once a productive VH-DH-JH rearrangement and a productive VL-JL rearrange￾ment have occurred, the recombination machinery is turned off, so that the heavy- and light-chain genes on the homolo￾gous chromosomes are not expressed. G. D. Yancopoulos and F. W. Alt have proposed a model to account for allelic exclusion (Figure 5-11). They suggest that once a productive rearrangement is attained, its encoded protein is expressed and the presence of this protein acts as a signal to prevent further gene rearrangement. According to their model, the presence of heavy chains signals the 116 PART II Generation of B-Cell and T-Cell Responses FIGURE 5-11 Model to account for allelic exclusion. Heavy-chain genes rearrange first, and once a productive heavy-chain gene rearrangement occurs, the protein product prevents rearrange￾ment of the other heavy-chain allele and initiates light-chain gene rearrangement. In the mouse, rearrangement of  light-chain genes precedes rearrangement of the  genes, as shown here. In humans, Vκ Jκ µ Productive D allele #1 H JH VH DH JH VH DH JH VH DH JH VH DH JH Productive allele #2 Cell death Nonproductive allele #1 Vκ Jκ Vλ Jλ Vλ Jλ Productive allele #1 Productive allele #2 Productive allele #1 Productive allele #2 Nonproductive allele #1 Nonproductive allele #2 Nonproductive allele #2 Cell death Nonproductive allele #2 Nonproductive allele #1 µ + λ chains inhibit rearrangement of λ allele #2 µ + κ chains inhibit λ rearrangement µ + κ chains inhibit rearrangement of κ allele #2 and λ rearrangement µ heavy chain inhibits rearrangement of µ allele #2 and induces κ rearrangement Progenitor B cell Ig µ µ + κ µ µ + κ µ + λ µ µ + λ µ µ maturing B cell to turn off rearrangement of the other heavy-chain allele and to turn on rearrangement of the  light-chain genes. If a productive  rearrangement occurs,  light chains are produced and then pair with heavy chains to form a complete antibody molecule. The presence of this antibody then turns off further light-chain rearrangement. If  rearrangement is nonproductive for both  alleles, re￾arrangement of the -chain genes begins. If neither  allele rearranges productively, the B cell presumably ceases to ma￾ture and soon dies by apoptosis. Two studies with transgenic mice have supported the hy￾pothesis that the protein products encoded by rearranged heavy- and light-chain genes regulate rearrangement of the remaining alleles. In one study, transgenic mice carrying a rearranged heavy-chain transgene were prepared. The transgene product was expressed by a large percentage of the B cells, and rearrangement of the endogenous immunoglob￾ulin heavy-chain genes was blocked. Similarly, cells from a transgenic mouse carrying a  light-chain transgene did not either  or  rearrangement can proceed once a productive heavy￾chain rearrangement has occurred. Formation of a complete immunoglobulin inhibits further light-chain gene rearrangement. If a nonproductive rearrangement occurs for one allele, then the cell attempts rearrangement of the other allele. [Adapted from G. D. Yancopoulos and F. W. Alt, 1986, Annu. Rev. Immunol. 4:339.] 8536d_ch05_105-136 8/22/02 2:47 PM Page 116 mac46 mac46:1256_des:8536d:Goldsby et al. / Immunology 5e: