Stable RNA/DNA hybrids in the mammalian genome: inducible intermediates in immunoglobulin class switch recombination.
Although it is well established that mammalian class switch recombination is responsible for altering the class of immunoglobulins, the mechanistic details of the process have remained unclear. Here, we show that stable RNA/DNA hybrids form at class switch sequences in the mouse genome upon cytokine-specific stimulation of class switch in primary splenic B cells. The RNA hybridized to the switch DNA is transcribed in the physiological orientation. Mice that constitutively express an Escherichia coli ribonuclease H transgene show a marked reduction in RNA/DNA hybrid formation, an impaired ability to generate serum immunoglobulin G antibodies, and significant inhibition of class switch recombination in their splenic B cells. These data provide evidence that stable RNA/DNA hybrids exist in the mammalian nuclear genome, can serve as intermediates for physiologic processes, and are mechanistically important for efficient class switching in vivo.
Tracy
R B
RB
Department of Pathology, University of Southern California Keck School of Medicine, Los Angeles, CA 90089-9176, USA.
Hsieh
C L
CL
Lieber
M R
MR
D000818:Animals; D001402:B-Lymphocytes / Q000276:immunology*; D015152:Blotting, Northern; D004247:DNA / Q000378:metabolism*; D017578:Immunoglobulin Class Switching; D007134:Immunoglobulin Switch Region; D007136:Immunoglobulins / Q000096:biosynthesis; D016753:Interleukin-10 / Q000276:immunology; D015847:Interleukin-4 / Q000276:immunology; D008070:Lipopolysaccharides / Q000494:pharmacology; D008213:Lymphocyte Activation; D051379:Mice; D008810:Mice, Inbred C57BL; D008822:Mice, Transgenic; D008957:Models, Genetic; D009693:Nucleic Acid Hybridization; D012313:RNA / Q000378:metabolism*; D012326:RNA Splicing; D011995:Recombination, Genetic; D020133:Reverse Transcriptase Polymerase Chain Reaction; D016914:Ribonuclease H / Q000378:metabolism; D012259:Ribonuclease, Pancreatic / Q000378:metabolism; D014158:Transcription, Genetic
D007136:Immunoglobulins; D008070:Lipopolysaccharides; D016753:Interleukin-10; D015847:Interleukin-4; D012313:RNA; D004247:DNA; D016914:Ribonuclease H; D012259:Ribonuclease, Pancreatic