Sylvie Huck

New subgroups in the human T cell rearranging V gamma gene locus.

Two new V gamma genes in humans are described from rearrangement in T cell lines, which constitute single members of new V gene subgroups of the T‐cell rearranging gamma (TRG gamma) locus. These two genes (herein designated as belonging to V gamma III and V gamma IV subgroups) are located between V gamma I/V gamma II subgroups and the constant (C) gamma genes. The existence of these new genes brings the number of different, potentially useable, human TRG V gamma genes to eight (excluding at least five pseudo V gamma genes) and the number of distinct subgroups to four. Polymorphism in the sequence of the V gamma II subgroup gene is also described and rearranged fragment sizes which make possible an unequivocal assignment of a V gamma rearrangement are given. These results extend previous conclusions of the inherited diversity of the human TRG V gamma locus.

Variable region genes in the human T-cell rearranging gamma (TRG) locus: V-J junction and homology with the mouse genes.

The locus of the human T cell rearranging gamma (TRG) or T cell receptor gamma chain genes comprises at least 14 variable genes (TRGV) belonging to four subgroups, five joining segments (TRGJ) and two constant region genes (TRGC). Nine V gamma genes belong to subgroup I, whereas subgroups II, III and IV each consists of a single gene respectively designated V9, V10 and V11. T cells expressing the gamma chain (TRG+) and recognized by the anti‐Ti gamma A monoclonal antibody have been shown to rearrange the V9 gene. In order to assess the N diversity at the V‐J junction in the TRG+ cells, the germline sequences of the segments involved in the V‐J rearrangements must be known. In this paper, we report the sequences of the germline V9 and V10 genes. Comparison of the V‐J junction and N region from transcripts or rearranged TRG genes belonging to the different subgroups shows no evidence of D segments in the human TRG locus. Sequences of the rearranged V11 gene from the JM cell line and those of the VA and VB pseudogenes, located upstream of V9 and V11 respectively, are given. Our results bring the number of human V gamma genes whose sequence is known to 13 and reveal unexpected homology with the mouse V gamma genes.

Rearrangements to the JP1, JP and JP2 segments in the human T-cell rearranging gamma gene (TRGγ) locus

In the human T‐cell rearranging gamma (TRGγ) locus, five joining (J) segments have been identified: J1, J2 and three additional segments JP, JP1 and JP2. We report the sequence of the germline JP1 segment and compare it with the other human and mouse Jγ segments. We also demonstrate that rearrangements to the three additional Jγ segments can be identified by hybridization of the KpnI digests to the Jγ1 probe pH60. Since rearrangements to J1 or J2 can be assigned, using the same pH60 probe, to one of the nine variable (V) γ genes known to rearrange [(1987) EMBO J. 6, 1945–1950], our results show that a unique probe can detect all the TRGγ rearrangements and be particularly useful for assessing the preferential usage of Vγ and Jγ segments in the TRG γ‐expressing cells.

Exon duplication and triplication in the human T-cell receptor gamma constant region genes and RFLP in french, lebanese, tunisian, and black african populations

The human T-cell receptor gamma (TCRG) locus comprises 14 variable genes (TRGV), five joining segments (TRGJ), and two constant region genes (TRGC). The constant gamma 1 gene, TRGC1, consists of three exons, whereas the TRGC2 gene contains four or five exons due to the duplication or triplication of exon 2 and spans 9.5 kb or 12 kb, respectively. In this paper, we define the alleles of the T-cell receptor gamma J2 and C2 genes, and we show that two Hind III allelic fragments, 5.4 kb and 8 kb, characterize unambiguously the C2 gene with duplication or triplication of exon 2. We show also that the cDNA of the HPB-MLT cell line results from the transcription of an allelic TRGC2 gene with duplicated exon 2. We propose a model involving unequal crossing-overs to explain the organization and the evolution of the TRGC locus. Moreover, we analyze the TCRG haplotypes in four different populations (French, Lebanese, Tunisian, and Black African) to underline their interest for population genetics.

A gamma 3 hinge region probe: first specific human immunoglobulin subclass probe

We report the first specific human immunoglobulin subclass probe which was obtained by subcloning the gamma 3 hinge region. This specific γ3 probe allowed us to identify with certainty the Cγ3 gene on Southern genomic blots to describe the first Cγ3 restriction fragment length polymorphism (EZZ γ3 RF) and to show that an IgG3 selective deficiency previously described serologically was not due to a deletion of the C3 gene. Such a probe should be particularly useful for screening libraries from individuals with IgG3 immunodeficiencies or presenting unusual Cγ3 genes and consequently for studying the Cγ gene evolution. Immunoglobulin IgCH gene (Human) Gm allotype Restriction fragment length polymorphism Immunoglobulin G3 deficiency Hinge

S-phase progression stimulates both the mutagenic KU-independent pathway and mutagenic processing of KU-dependent intermediates, for nonhomologous end joining

We used intrachromosomal substrates to directly monitor the effect of the cell cycle on the efficiency and the accuracy of nonhomologous end joining (NHEJ) in mammalian cells. We show that both KU and KU-independent (KU-alt) pathways are efficient when maintaining cells in G1/S, in G2/M or during dynamic progression through S phase. In addition, the accuracy of NHEJ is barely altered when the cells are blocked in G1/S or in G2/M. However, progression through S phase increases the frequency of deletions, which is a hallmark of the KU-alt pathway. Moreover, we show that the intermediates that are generated by the KU-dependent end joining of non-fully complementary ends, and which contain mismatches, nicks or gap intermediates, are less accurately processed when the cells progress through S phase. In conclusion, both KU and KU-alt processes are active throughout the cell cycle, but the repair is more error prone during S phase, both by increasing the mutagenic KU-alt pathway and decreasing the accuracy of the repair of the intermediates generated by the KU-dependent pathway.

The IGHG3 gene shows a structural polymorphism characterized by different hinge lengths: sequence of a new 2-exon hinge gene

Abstract Four of the five human IGHG genes (G1, GP, G2, and G4) display a hinge region consisting of a unique exon. In contrast, IGHG3 exhibits a different structure in which the hinge is constituted by four or, less frequently, three exons. We report here the nucleotide sequence of a new 2-exon hinge G3 gene found in a Mandenka individual from Eastern Senegal. A comparison of this sequence with that of 4-exon and 3-exon hinge G3 genes suggests that the 3-exon and 2-exon hinge forms arose independently by deletion events in a 4-exon hinge gene.

The human T-cell receptor gamma variable pseudogeneV10 is a distinctive marker of human speciation

The V10 variable gene of the human T-cell receptor gamma locus (TCRG-V10), the only member of the subgroup III, has a structural defect which inhibits the splicing of the leader intron. We show that there is a single point mutation in the V10 leader donor splice site responsible for this situation and that this mutation is found in the different populations tested, indicating that V10 corresponds to a pseudogene in humans. We restored the splice site by mutagenesis and obtained correct splicing in vitro. Analysis of the V10 germline gene in different primates reveals functional splice sites in the closest human apes, the chimpanzee and the gorilla. The splice competence of TCRG-V10 in higher primates was addressed in peripheral blood lymphocytes from chimpanzee by specific cDNA amplification, and correct splicing of the TCRG-V10 leader intron was found as well as a majority of in frame rearrangements involving only the TCRG-J1 or J2 segments. These results suggest that V10(+)gamma /delta T cells may represent an important subset in the non-human higher primates, contrary to the situation observed in the human.

Deletion, insertion, and restriction site polymorphism of the T-cell receptor gamma variable locus in French, Lebanese, Tunisian, and black African populations.

The human T-cell receptor gamma region spans 160 kb of genomic DNA and is densely populated by coding sequences. Restriction fragment length polymorphisms have been previously documented for the constant region genes, the joining segments, and the variable genes belonging to subgroups I and IV. Here were further define the polymorphism of theV gamma I subgroup genes, based on complete mapping of theEco RI andTaq I allelic restriction fragments. We describe seven haplotypes; five result from polymorphic restriction sites, the sixth corresponds to a deletion of about 10 kb encompassingV4 andV5, and the seventh results from an insertion of an additional gene,V3P, betweenV3 andV4. As a consequence of the deletion or insertion polymorphism, the number ofV gamma I subgroup genes vary from seven in haplotypeTRGVI*3 to ten in haplotypeTRGVI*4, whereas the most common haplotype,TRGVI*1, has nineV genes, five of them being functional. Frequencies of the differentTRGVI haplotypes in French, Lebanese, Tunisian, and Black African populations are given.