The guanosine triphosphatases (GTPases) of the immunity-associated protein (GIMAP) family of putative GTPases has been implicated in the regulation of T-lymphocyte development and survival. A mouse conditional knockout allele was generated for the immune GTPase gene

To date, there has been no systematic study of the process of affinity maturation of human antibodies. We therefore sequenced the variable region genes (V genes) of 14 human monoclonal antibodies specific for the erythrocyte Rh(D) alloantigen and determined the germline gene segments of origin and extent of somatic hypermutation. These data were correlated with determinations of antibody affinity. The four IgM antibodies (low affinity) appear to be derived from two germline heavy chain variable region gene segments and one or two germline light chain variable region gene segments and were not extensively mutated. The 10 IgG antibodies (higher affinity) appear to be derived from somatic hypermutation of these V gene segments and by use of new V gene segments or V gene segment combinations (repertoire shift). Affinity generally increased with increasing somatic hypermutation; on average, there were 8.

SUMMARYIntegrin aEb7 is expressed almost exclusively by mucosal T cells and mucosal dendritic antigenpresenting cells (APCs) and is thought to be induced locally by transforming growth factor-b (TGF-b). In mice, mRNA for the aE subunit was found to be abundant in mucosal T cells but absent from other tissues. Exposure of a T-cell line to TGF-b strongly up-regulated aE mRNA levels within 30 min, and nuclear run-on experiments established that regulation occurred at the level of transcription. The organization of the human aE gene and a very closely linked novel gene, ELG, was determined. The aE promoter was tested in T cells and ®broblasts and functioned equally well in both cell types and did not confer TGF-b responsiveness. Regions of the promoter providing enhancer activity and phorbol 12-myristate 13-acetate (PMA) responsiveness were identi®ed by deletion studies. DNAse 1 hypersensitivity analysis of 36 kb of the aE gene revealed one hypersensitive site, found only in aE + cells, located near the transcription start points. These results show that, unlike the situation with other integrins, lineage speci®city and cytokine responsiveness of aE transcription are not conferred by the proximal promoter. Speci®city may depend on distant control elements that have not yet been identi®ed.

Reports suggest that two members of the novel immune-associated nucleotide (Ian) GTPase family, Ian1 and Ian5, play roles in T cell development. We performed real-time PCR analysis of the expression of Ian genes of the rat during T cell maturation, in macrophages and in cell lines. We found that all of the genes were expressed at relatively low levels at the early double-negative thymocyte stage but were expressed more strongly at later cell stages. Our study also revealed the fact that the previously reported Ian9, Ian10 and Ian11 genes are, instead, parts of a single gene for which we retain the name Ian9, potentially encoding a GTPase with a highly unusual triplicated structure. Antisera were developed against both Ian1 and Ian9. We established that Ian9 is produced as an approximately 75-kDa protein in both T cells and thymocytes. We observed that levels of both Ian1 and Ian9 proteins are profoundly reduced in T cells from lymphopenic rats as compared with wild-type rats. It was demonstrated that thymocytes and B cells from lymphopenic rats (Ian5 null) did not show enhanced sensitivity to gamma-irradiation-induced apoptosis.

A mutation in the rat GIMAP5 gene predisposes for autoimmunity, most famously in the BB rat model of autoimmune type 1 diabetes mellitus. This mutation is associated with severe peripheral T lymphopenia, as is mutation of the same gene in mice, but the mechanism by which GIMAP5 normally protects T cells from death is unknown. GIMAP5 is a putative small GTPase, a class of proteins which often fulfil their functions in the vicinity of cellular membranes. The objective of this study was to determine the normal intracellular location of GIMAP5 in lymphoid cells. Combining studies in rat, mouse and human systems, novel monoclonal antibodies (mAbs) were used to examine the localization of GIMAP5 and the closely-related protein, GIMAP1, in lymphoid cells by means of confocal microscopy and sub-cellular fractionation combined with immunoblotting. Additionally, human Jurkat T cells that inducibly express epitope-tagged GIMAP5 were established and used in electron microscopy (EM). Endogenous GIMAP5 was found to be located in a membraneous compartment/s which was also detected by established markers of lysosomes. GIMAP1, by contrast, was found to be located in the Golgi apparatus. EM studies of the inducible Jurkat T cells also found GIMAP5 in lysosomes and, in addition, in multivesicular bodies. This study establishes that the endogenous location of GIMAP5 is in lysosomes and related compartments and provides a clearer context for hypotheses about its mechanism of action. Key words: GIMAP, GTPase, golgi apparatus, lysosome has shown that the mammalian GIMAPs are members of the AIG1 class of GTPases and have relatives in higher plants and non-mammalian vertebrates but not in many convenient experimental organisms such as E. coli, C. elegans or D. melanogaster. The autoimmunity-related GIMAP5 GTPase is a lysosome-associated protein