The Linker for Activation of T Cells (LAT) Signaling Hub: From Signaling Complexes to Microclusters

Since the cloning of the critical adapter, LAT (linker for activation of T cells), more than 15 years ago, a combination of multiple scientific approaches and techniques continues to provide valuable insights into the formation, composition, regulation, dynamics, and function of LAT-based signaling complexes. In this review, we will summarize current views on the assembly of signaling complexes nucleated by LAT. LAT forms numerous interactions with other signaling molecules, leading to cooperativity in the system. Furthermore, oligomerization of LAT by adapter complexes enhances intracellular signaling and is physiologically relevant. These results will be related to data from super-resolution microscopy studies that have revealed the smallest LAT-based signaling units and nanostructure.     

Chromosomal localization and genomic organization for the galactose/ N-acetylgalactosamine/N-acetylglucosamine 6-O-sulfotransferase gene family

The galactose/N-acetylgalactosamine/N-acetylglucosamine 6-O-sulfotransferases (GSTs) are a family of Golgi-resident enzymes that transfer sulfate from 3'phosphoadenosine 5'phospho-sulfate to the 6-hydroxyl group of galactose, N-acetylgalactosamine, or N-acetylglucosamine in nascent glycoproteins. These sulfation modifications are functionally important in settings as diverse as cartilage structure and lymphocyte homing. To date six members of this gene family have been described in human and in mouse. We have determined the chromosomal localization of these genes as well as their genomic organization. While the broadly expressed enzymes implicated in proteoglycan biosynthesis are located on different chromosomes, the highly tissue specific enzymes GST-3 and 4 are encoded by genes located both in band q23.1--23.2 on chromosome 16. In the mouse, both genes reside in the syntenic region 8E1 on chromosome 8. This cross-species conserved clustering is suggestive of related functional roles for these genes. The human GST4 locus actually contains two highly similar open reading frames (ORF) that are 50 kb apart and encode two highly similar enzyme isoforms termed GST-4 alpha and GST-4 beta. All genes except GST0 (chondroitin 6-O-sulfotransferase) contain intron-less ORFs. With one exception these are fused directly to sequences encoding the 3' untranslated regions (UTR) of the respective mature mRNAs. The 5' UTRs of these mRNAs are usually encoded by a number of short exons 5' of the respective ORF. 5'UTRs of the same enzyme expressed in different cell types are sometimes derived from different exons located upstream of the ORF. The genomic organization of the GSTs resembles that of certain glycosyltransferase gene families.     

The Us3 Protein of Herpes Simplex Virus 1 Inhibits T Cell Signaling by Confining Linker for Activation of T Cells (LAT) Activation via TRAF6 Protein

Herpes simplex virus 1 (HSV-1) is the most prevalent human virus and causes global morbidity because the virus is able to infect multiple cell types. Remarkably, HSV infection switches between lytic and latent cycles, where T cells play a critical role. However, the precise way of virus-host interactions is incompletely understood. Here we report that HSV-1 productively infected Jurkat T-cells and inhibited antigen-induced T cell receptor activation. We discovered that HSV-1-encoded Us3 protein interrupted TCR signaling and interleukin-2 production by inactivation of the linker for activation of T cells. This study unveils a mechanism by which HSV-1 intrudes into early events of TCR-mediated cell signaling and may provide novel insights into HSV infection, during which the virus escapes from host immune surveillance.     

Chromosomal localization of the human hexabrachion (tenascin) gene and evidence for recent reduplication within the gene

Using analysis of rodent-human somatic cell hybrids as well as in situ hybridization of hexabrachion cDNA probes to normal human metaphase chromosomes, we have localized the human hexabrachion gene to chromosome 9, bands q32-q34. We also put forward the hypothesis that there has been a recent reduplication of a small segment of the human hexabrachion gene. We support this hypothesis by comparison of codon usage in this segment of the gene to codon usage in the remainder of the gene. This hypothesis is also supported by comparison of the sequence of human hexabrachion to that of the chicken hexabrachion. In addition, the latter comparison shows that the reduplication most likely occurred after the divergence of mammalian and avian species.     

Chromosomal localization and molecular organization of human genomic fragment containing TNF/LT locus in transgenic mice

Molecular organization, copy number and chromosomal localization of human TNF/LT locus fragment were determined in genomes of two transgenic mouse lines. Genome of the first one contains two copies, organized in head-to-tail manner and determined on eighth chromosome by karyotyping; single transgene copy of the second line is observed on the fifth chromosome. These mice could serve as valuable model for studying both human tumor necrosis factor and lymphotoxin physiological functions.     

Chromosomal localization and molecular organization of the human genomic fragment containing the TNF/LT locus in transgenic mice

The molecular organization, copy number, and chromosome location of the human TNF/LT transgenes were studied in the genomes of two transgenic mouse strains. One strain proved to carry two transgene copies arranged head-to-tail and detected on chromosome 8 by karyotyping. The other strain had one transgene copy observed on chromosome 5. The strains provide a model for studying the physiological functions of the tumor necrosis factor and lymphotoxin.     

Chromosomal localization, genomic characterization, and mapping to the Noonan syndrome critical region of the human Deltex (DTX1) gene

The human Deltex (DTX1) gene encodes a cytoplasmic protein that functions as a positive regulator of the Notch signaling pathway. We have determined the genomic organization and map location of the human gene. DTX1 encodes a 2.5-kb cDNA that is composed of nine exons. The DTX1 gene maps to chromosomal region 12q24 in the vicinity of the Noonan syndrome critical region. We have fine-mapped DTX1 to within this critical region and evaluate it as a candidate gene for this disorder.     

Chromosomal localization of the human elastin gene.

mRNA isolated from fetal human aorta was used to synthesize cDNA that was cloned into the PstI site of pBR322. The recombinant clones were screened with an authentic sheep elastin cDNA, and one human clone that hybridized strongly was isolated and characterized. The 421-base pair (bp) insert of this human clone was sequenced by the dideoxy method, and the DNA sequence showed strong homology to the nontranslated portion of the sheep elastin cDNA. This result unequivocally identified the human clone, designated pcHEL1, as an elastin clone. Plasmid pcHEL1 labeled with [3H] nucleotides was used in in situ hybridization experiments utilizing normal metaphase chromosomes and also with cells carrying a balanced translocation between chromosomes 1 and 2: 46,XY,t(1;2)(p36;q31). The results strongly suggest that the elastin gene is localized to the q31----qter region of chromosome 2.     

Chromosomal localization of the human gene for palmitoyl-CoA ligase (FACL1).

We have localized the gene (FACL1) encoding human long chain fatty acid-coenzyme A ligase, E.C.6.2.1.3, also known as palmitoyl-CoA ligase. Using in-situ hybridization, we have mapped this gene to chromosome region 3q13 in the human karyotype.     

The Linker for Activation of B Cells (LAB)/Non-T Cell Activation Linker (NTAL) Regulates Triggering Receptor Expressed on Myeloid Cells (TREM)-2 Signaling and Macrophage Inflammatory Responses Independently of the Linker for Activation of T Cells

Triggering receptor expressed on myeloid cells-2 (TREM-2) is rapidly emerging as a key regulator of the innate immune response via its regulation of macrophage inflammatory responses. Here we demonstrate that proximal TREM-2 signaling parallels other DAP12-based receptor systems in its use of Syk and Src-family kinases. However, we find that the linker for activation of T cells (LAT) is severely reduced as monocytes differentiate into macrophages and that TREM-2 exclusively uses the linker for activation of B cells (LAB encoded by the gene Lat2^−/−) to mediate downstream signaling. LAB is required for TREM-2-mediated activation of Erk1/2 and dampens proximal TREM-2 signals through a novel LAT-independent mechanism resulting in macrophages with proinflammatory properties. Thus, Lat2^−/− macrophages have increased TREM-2-induced proximal phosphorylation, and lipopolysaccharide stimulation of these cells leads to increased interleukin-10 (IL-10) and decreased IL-12p40 production relative to wild type cells. Together these data identify LAB as a critical, LAT-independent regulator of TREM-2 signaling and macrophage development capable of controlling subsequent inflammatory responses.     

Chromosomal localization of the human interleukin 1 alpha (IL-1 alpha) gene

The human interleukin 1 alpha gene was assigned to chromosome 2 using Southern transfer analysis of human-rodent somatic cell hybrid DNAs. The gene was regionally localized to 2q12-21 using in situ hybridization to metaphase chromosomes. These results indicate that the IL-1 alpha gene maps to the same general region on the long arm of chromosome 2 as the IL-1 beta gene, which has been previously assigned.     

Nucleotide sequence and genomic organization of a human T lymphocyte serine protease gene

We have determined the nucleotide sequence of 4508 base pairs of human genomic DNA which contain the human serine esterase gene from cytotoxic T lymphocytes (SECT) (equivalent to the 1-3E cDNA clone) and include 879 bp of 5' flanking DNA and 393 bp of 3' flanking DNA. The gene consists of five exons of 88, 148, 136, 261, and 257 nucleotides separated by four introns of 1043, 455, 205, and 643 nucleotides. The location of introns with respect to protein coding sequences in the SECT gene is identical to that of the human cathepsin G and murine granzyme B genes. Comparison of SECT gene exonic sequences to murine granzyme B-F cDNA sequences indicates similarities of 75 and 72% for granzymes B and C and 61, 59, and 61% for granzymes D, E, and F, respectively. The 5' flanking sequence of the SECT gene showed similarity only to the 5' flanking sequence of the murine granzyme B gene, indicating that these genes are homologous. Comparison of the SECT gene sequence to the human cathepsin G sequence indicated no similarity in the 5' flanking DNA although the exonic sequences show 64% sequence similarity overall and 45% sequence similarity in the respective 3' untranslated regions. These similarities suggest that the SECT and cathepsin G genes are members of the same family of serine protease genes. Evidence from high and low stringency Southern transfer analysis of human genomic DNA indicates the presence of another gene of at least 85% sequence similarity to the SECT gene.     

Chromosomal localization of the human alpha-L-iduronidase gene (IDUA) to 4p16.3.

The lysosomal hydrolase alpha-L-iduronidase (IDUA) is one of the enzymes in the metabolic pathway responsible for the degradation of the glycosaminoglycans heparan sulfate and dermatan sulfate. In humans a deficiency of IDUA leads to the accumulation of glycosaminoglycans, resulting in the lysosomal storage disorder mucopolysaccharidosis type I. A genomic subclone and a cDNA clone encoding human IDUA were used to localize IDUA to chromosome 4p16.3 by in situ hybridization and this was confirmed by Southern blot analysis. This localization is different from that of a previous report mapping IDUA to chromosome 22 and places the gene for IDUA in the same region of chromosome 4 as the Huntington disease gene. Measurement of expressed human IDUA activity in human-mouse hybrid cell lines confirmed that IDUA is on chromosome 4.     

Chromosomal localization of ARSB,the gene for human N-acetylgalactosamine-4-sulphatase

Summary A deficiency of N-acetylgalactosamine-4-sulphatase (G4S, gene symbol ARSB), results in the accumulation of undegraded substrate and the lysosomal storage disorder, Maroteaux-Lamy syndrome (mucopolysaccharidosis type VI). In situ hybridization using an ³H-labelled human G4S genomic DNA fragment to human metaphase chromosomes localized ARSB to chromosome 5q13–5q14. This location is consistent with, an refines, previous chromosomal assignments based on the expression of human G4S in somatic cell hybrids.