Reassessment of germline heavy chain transcripts from two murine VH families.
While expression of functional heavy chain immunoglobulin mRNA requires rearrangement of variable (VH), diversity (D) and (JH) gene segments, these individual gene segments can be transcribed prior to their rearrangement. It has been proposed that the resulting germline, or sterile, transcripts play an important role in the rearrangement process because strong correlations between rearrangement frequency and sterile transcript levels have been observed in some studies. Murine VH genes have been grouped into families on the basis of coding sequence homology. VH families rearrange in a developmentally regulated manner, so that rearrangements of genes from several VH families are detected earlier than rearrangements of J558 family genes. Paradoxically, the only VH family for which sterile transcripts have been documented is the J558 family. We used RT-PCR analyses to ask whether sterile transcripts from other VH families could be detected in fetal liver samples prior to their rearrangement. While J558 family germline transcripts were easily detected, no sterile transcripts were observed from the S107 family. Our studies also revealed the ability of small quantities of degraded genomic DNA to nonspecifically prime cDNA synthesis, emphasizing the need for caution in interpreting RT-PCR data in which family-specific oligos are used for cDNA production. These results cast doubt on the idea that sterile transcripts are required for V(H)DJ(H) rearrangement.
Webb
C F
CF
Department of Immunobiology/Cancer, Oklahoma Medical Research Foundation, Oklahoma City 73104, USA.
Dou
S
S
Buchanan
K L
KL
Resta
R
R
Smithson
G
G
Smith
E A
EA
D000818:Animals; D002874:Chromosome Mapping; D004247:DNA / Q000235:genetics / Q000378:metabolism; D017931:DNA Primers / Q000235:genetics; D018076:DNA, Complementary / Q000096:biosynthesis; D005333:Fetus / Q000276:immunology / Q000378:metabolism; D015321:Gene Rearrangement; D005803:Genes, Immunoglobulin; D007143:Immunoglobulin Heavy Chains / Q000096:biosynthesis* / Q000235:genetics* / Q000276:immunology; D007075:Immunoglobulin M / Q000235:genetics / Q000378:metabolism; D007135:Immunoglobulin Variable Region / Q000096:biosynthesis* / Q000235:genetics* / Q000276:immunology; D008099:Liver / Q000196:embryology / Q000276:immunology / Q000378:metabolism; D051379:Mice; D016133:Polymerase Chain Reaction; D012313:RNA / Q000235:genetics; D014158:Transcription, Genetic
D017931:DNA Primers; D018076:DNA, Complementary; D007143:Immunoglobulin Heavy Chains; D007075:Immunoglobulin M; D007135:Immunoglobulin Variable Region; D012313:RNA; D004247:DNA