Proceedings of the XLV Italian Society of Agricultural Genetics - SIGA Annual Congress
Salsomaggiore Terme, Italy - 26/29 September, 2001
EXON-INTRON ORGANIZATION OF TRGC GENES IN RUMINANT ANIMALS
MICCOLI M.C., ANTONACCI R., VACCARELLI G., MASSARI S., CICCARESE S.
Dipartimento di Anatomia Patologica e di Genetica - Università di Bari, Via Amendola 165/A, 70126 BARI
T cell receptor, TRGC genes, Sheep, Cow, Exon-intron organization
Recent studies in ruminants have revealed that the rule of alfa/beta and gamma/delta TcR does not apply to all mammals since a large and sometimes predominant fraction of T cells in sheep and bovine expresses the gamma/delta TcR. Moreover, in addition to this striking difference between the relative usage of the two types of TcR in these divergent species, the gamma chain complex differs at the structural level. In fact, sheep and bovine contain five functional TRGC genes, isolated as cDNA, all of which are related to each other by sequence, and, at a lower level, to their human and mouse homologues. On the contrary humans contain two TRGC genes and mouse three TRGC genes respectively. The ovine and bovine TcR gamma chains contain the longest hinge region of any TcR gamma chain described so far. In addition the sheep hinge regions are notably heterogeneous ranging in length from 24 to 75 amino acids. Some residues are conserved among all sequences, including the four most membrane-proximal ones (SAYY) and a cystein residue. In the extreme distal region of the connecting peptide, each chain, Cgamma1, Cgamma2 and Cgamma4, contains two additional cystein residues at conserved positions. Thus although the essential structure of the gamma chain appears to be well conserved through evolution, the marker of heterogeneity, evident in the hinge region of the chain both within and between species, may be of structural and functional importance.
A series of genomic clones derived from a sheep library were used to determine the germ line configuration and the exon-intron organization of TRGC2, TRGC3 and TRGC4 genes. Based on the outcomes of molecular analysis we have compared and aligned the genomic sequences with all the known complete cDNA sequences of cow and sheep and we have deduced the exon-intron organization of all TRGC genes in these ruminant animals. The EX1, corresponding to the disulfide-linked constant domain and the EX3, corresponding to the transmembrane and cytoplasmatic domains are similar in length in all genes. Conversely, the hinge-encoding EX2A, EX2B and EX2C exons differ in number and length between genes and EX2A contains the TTKPP motif irrespective of whether it occurs in single or quadruplicate form. In addition molecular data indicate that at least two TRGC2 regions are present in sheep and cow. The complexity of the TRG locus in ruminant animals is emphasized by chromosomal mapping results which indicate that in sheep TRG2 maps at bands 4q1.5-2.2 whereas TRG1 maps at 4q3.1. The same split of TRG locus was observed in the homologous chromosome 4 in cow. Our recent results show that sheep TRG4 maps at the same position as TRG2, while the chromosomal mapping of the TRG3 overlaps TRG1 position.
The phylogenetic analysis grouped the ruminant TRGC genes in two clusters that could emerge from two ancestral forms that underwent a series of duplications giving rise to the new sequences that were selected and then fixed in the ruminant lineages. A correlation between cluster distribution in the phylogenetic tree of TRGC genes and their expression during fetal development has been hypothesized.