TY - JOUR
T1 - Ig heavy chain expression and class switching in vitro from an allele lacking the 3' enhancers DNase I-hypersensitive hs3A and hs1,2
AU - Saleque, Shireen
AU - Singh, Mallika
AU - Birshtein, Barbara K.
PY - 1999/3/1
Y1 - 1999/3/1
N2 - The murine Ig heavy chain (IgH) 3' regulatory region contains four enhancers: hs3A, hs1,2, hs3B, and hs4. Various studies have suggested a role for these enhancers in regulating IgH expression and class switching. Here we assess the role of hs3A and hs1,2 in these processes by exploiting a naturally occurring deletion of these enhancers from the expressed, C57BL/6 allele of the F1 pre-B cell line, 70Z/3. Equivalent μ expression in 70Z/3 and 18-81 (which has an intact 3' region) indicated that hs3A and hs1,2 were not essential for μ expression at the pre-B cell stage. To further examine the role of hs3A and hs1,2 in IgH function at the plasma cell stage, we fused 70Z/3 with the plasmacytoma NSO. Electromobility shift assay analysis of the 70Z/3-NSO hybrids revealed a transcription factor complement conducive to the activation of the 3' enhancers. Despite the lack of enhancers, hs3A and hs1,2, the level of μ RNA and protein in the 70Z/3-NSO fusion hybrids was substantially elevated relative to its pre-B parent and comparable with that observed in a number of μ-producing spleen cell hybridomas. Additionally, ELISAspot assays showed that the 70Z/3-NSO hybrid underwent spontaneous class switching in culture to IgG1 at a frequency comparable with that of most hybridomas. These results indicate that hs3A and hs1,2 are not essential for high levels of IgH expression or for spontaneous class switching in a plasma cell line.
AB - The murine Ig heavy chain (IgH) 3' regulatory region contains four enhancers: hs3A, hs1,2, hs3B, and hs4. Various studies have suggested a role for these enhancers in regulating IgH expression and class switching. Here we assess the role of hs3A and hs1,2 in these processes by exploiting a naturally occurring deletion of these enhancers from the expressed, C57BL/6 allele of the F1 pre-B cell line, 70Z/3. Equivalent μ expression in 70Z/3 and 18-81 (which has an intact 3' region) indicated that hs3A and hs1,2 were not essential for μ expression at the pre-B cell stage. To further examine the role of hs3A and hs1,2 in IgH function at the plasma cell stage, we fused 70Z/3 with the plasmacytoma NSO. Electromobility shift assay analysis of the 70Z/3-NSO hybrids revealed a transcription factor complement conducive to the activation of the 3' enhancers. Despite the lack of enhancers, hs3A and hs1,2, the level of μ RNA and protein in the 70Z/3-NSO fusion hybrids was substantially elevated relative to its pre-B parent and comparable with that observed in a number of μ-producing spleen cell hybridomas. Additionally, ELISAspot assays showed that the 70Z/3-NSO hybrid underwent spontaneous class switching in culture to IgG1 at a frequency comparable with that of most hybridomas. These results indicate that hs3A and hs1,2 are not essential for high levels of IgH expression or for spontaneous class switching in a plasma cell line.
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M3 - Article
C2 - 10072526
AN - SCOPUS:0033104687
SN - 0022-1767
VL - 162
SP - 2791
EP - 2803
JO - Journal of Immunology
JF - Journal of Immunology
IS - 5
ER -