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L Hood

Publications and source records attributed to L Hood.

At least 145 records · Page 8Linked to original sources
PubMed1994

The expression of mouse T-cell receptor TCRDV genes in BALB/c spleen.

The rearrangement and expression of six different mouse T-cell receptor TCRDV (V delta) gene subfamilies have been studied in BALB/c mouse spleen. The results show that all the TCRDV gene segments studied can be rearranged and expressed with both the TCRA (alpha) and the TCRD (delta) chain. The apparently restricted and separated V gene repertoire for the TCRD and TCRAT-cell receptor may result from thymus and peripheral selection rather than from selective rearrangement in the DNA level. Several J gene segments show much higher concentration in spleen with TCRDV gene segments, however, these J gene segments do not tend to be located at either end of the J gene cluster.

Amino Acid Sequence↗
PubMed1994

Striking sequence similarity over almost 100 kilobases of human and mouse T-cell receptor DNA.

We report here the comparative DNA sequence analysis of nearly 100 kilobases of contiguous DNA in the C delta to C alpha region of the alpha/delta T cell receptor loci (TCRAC/TCRDC) of mouse and man. This analysis--the largest genomic sequence comparison so far--provides new insights into the functions of the T cell receptor genes as well as the surrounding chromosome structure through the identification of actively conserved DNA sequences. In this comparison we have identified a very high level of organizational and noncoding sequence similarity (approximately 71%) in contrast to previous findings in the beta-globin gene cluster. This observation begins to question the notion that much of the chromosomal non-coding sequence is junk.

Amino Acid Sequence↗
PubMed1994

T-cell receptor repertoire in tumor-infiltrating lymphocytes. Analysis of melanoma-specific long-term lines.

Cytotoxic T-lymphocytes (CTLs) can be isolated from human melanoma biopsies that specifically lyse autologous melanoma in vitro and can be effective therapeutic agents for patients with advanced disease. Recent evidence indicates that HLA-A2-restricted, melanoma-specific tumor-infiltrating lymphocytes (TILs) recognize melanomas obtained from different HLA-A2+ patients, suggesting the presence of one or more common melanoma antigens. Furthermore, T-cell receptor (TCR) repertoire analysis by other groups of TILs from fresh melanoma biopsies suggests that there is limited TCR V gene usage in TILs. One serious limitation in analyzing the TCR repertoire in fresh tumors has been the inability to correlate TCR usage with immune function. Therefore, the TCR repertoire was determined in long-term TIL cultures that specifically lysed autologous melanoma in vitro and in many cases mediated in vivo regression of metastatic cancer in patients with advanced disease. The TCR repertoire in cultured melanoma-specific TILs was diverse, with each TIL containing an average of 9.5 +/- 5.7 of the 23 V alpha and 11.2 +/- 5.9 of the 23 V beta subfamilies. Despite the large diversity observed, several V alpha and V beta genes (V alpha 1, V alpha 2, V alpha 22, V beta 13, V beta 14, and V beta 18) are very commonly found in melanoma-specific TILs. No statistically significant associations were observed between the presence of a TCR V gene subfamily in TILs and clinical response, HLA haplotype, or age of the culture. Even though the results in this study suggest that certain TCR V gene segments may be involved in immune responses to human melanoma, we were unable to demonstrate functionally that a particular T-cell clonotype recognizes melanoma tumor-associated antigens. Only the analysis of melanoma-specific CTL clones can determine which clonotypes are important in lysis of human melanoma.

Base Sequence↗
PubMed1994

Restoration of brain stem auditory-evoked potentials by gene transfer in shiverer mice.

We studied the shiverer mouse as a model for correcting hearing disorders resulting from genetic abnormalities of the central nervous system (CNS). Shiverer mice are homozygous for an autosomal recessive mutation (deletion) in the gene for myelin basic protein (MBP), a major protein component of the myelin sheath in the CNS. Under electron microscopic observation of the cochlear nerve, the CNS portion in shiverer mice showed hypomyelination, but the peripheral portion, including spiral ganglion cells, was normal. We produced MBP-transgenic mice by microinjection of an MBP cosmid clone into the pronucleus of fertilized eggs from shiverer mice. The transgenic mice were found to recover MBP levels up to 25% of normal. A greater number of axons in the transgenic mice were myelinated than in the shiverer mice, but the myelin sheath was not as thick as in normal controls. Every interpeak latency of brain stem auditory-evoked potentials was prolonged in the shiverer mice and improved in the transgenic mice. This study provides an example of gene therapy for hearing disorders caused by a CNS abnormality. We discuss some strategies for researching genetic hearing impairment or deafness in both animals and humans.

Animals↗
PubMed1993

Identification of an embryonic isoform of myelin basic protein that is expressed widely in the mouse embryo.

We have identified a myelin basic protein (MBP) isoform in mouse embryos that includes an exon upstream of the usual transcription initiation site. This isoform, embryonic-neonatal MBP (E-MBP), is expressed at the protein level in the embryonic nervous system at a time when other MBP isoforms are not detected. In addition to the central and peripheral nervous systems of the embryo and neonate, the thymus, spleen, and testes also express E-MBP at the protein level. The expression of E-MBP in cell types distinct from the nervous system strongly suggests that this MBP isoform has a role apart from the formation of myelin.

Amino Acid Sequence↗
PubMed1993

Cyclic AMP has a differentiative effect on an immortalized oligodendrocyte cell line.

We investigated the effects of increasing the concentration of intracellular cyclic adenosine monophosphate (cAMP) on genes associated with oligodendrocyte differentiation in an immortalized glial cell line, 6E12, derived from the spinal cord of an MBP-SV40 large T-antigen transgenic mouse. Raising intracellular levels of cAMP induced expression of oligodendrocyte differentiation antigens recognized by O4 and anti-galactocerebroside antibodies, up-regulated expression of the proteolipid protein (PLP) gene, and down-regulated glial fibrillary acidic protein (GFAP) expression. There was no treatment effect on myelin-associated glycoprotein (MAG) expression. These phenotypic changes are consistent with oligodendrocyte differentiation. Treatment of 6E12 cells with dibutyryl cyclic AMP (DBC) down-regulated myelin basic protein (MBP) gene expression, perhaps, because it also up-regulated expression of a putative MBP repressor SCIP/Tst-1. Moreover, the 6E12 cells expressed high levels of MBP mRNA but no MBP translation products were detected in the presence or absence of DBC. This immortalized glial cell line is proposed as a CNS model for cAMP-modulated myelin gene expression and for post-transcriptional regulation of MBP.

Animals↗
PubMed1993

DNA sequence determination by hybridization: a strategy for efficient large-scale sequencing.

The concept of sequencing by hybridization (SBH) makes use of an array of all possible n-nucleotide oligomers (n-mers) to identify n-mers present in an unknown DNA sequence. Computational approaches can then be used to assemble the complete sequence. As a validation of this concept, the sequences of three DNA fragments, 343 base pairs in length, were determined with octamer oligonucleotides. Possible applications of SBH include physical mapping (ordering) of overlapping DNA clones, sequence checking, DNA fingerprinting comparisons of normal and disease-causing genes, and the identification of DNA fragments with particular sequence motifs in complementary DNA and genomic libraries. The SBH techniques may accelerate the mapping and sequencing phases of the human genome project.

Animals↗
PubMed1993

Transgenic mouse model for neurocristopathy: Schwannomas and facial bone tumors.

We have characterized a strain of double transgenic mice with simian virus 40 large tumor antigen and prokaryotic lacZ under the control of the myelin basic protein promoter that develops spindle-cell sarcomas and osteogenic sarcomas at 5-7 months of age. Although poorly differentiated, the spindle-cell sarcomas were characterized as malignant Schwannomas based on their neural association, the presence of basal lamina, and expression of Schwann cell-specific genes. The osteogenic sarcomas were often multiple and appeared predominantly in the facial bones, less frequently in the ribs and vertebral column, and only rarely in the appendicular skeleton. Benign osteoblastic lesions were often observed adjacent to these sarcomas. Both the osteoblastic cells in the facial skeleton and Schwann cells are regarded as neural crest derivatives. The biological properties and anatomical location of these tumors suggest that they may share a common origin from the neural crest or its derivatives. R.P. Bolande [Hum. Pathol. (1974) 5, 409-429] introduced the term neurocristopathy as a unifying concept to describe such lesions arising from the neural crest or its derivatives. Cell lines established from both bone and Schwann cell tumors arising in these transgenic mice express simian virus 40 large tumor antigen mRNA as well as functional large tumor antigen. Such cell lines are potentially valuable in the search for markers that identify mammalian neural crest derivatives.

Animals↗
PubMed1993

Structural studies of the scrapie prion protein using mass spectrometry and amino acid sequencing.

The only component of the infectious scrapie prion identified to date is a protein designated PrPSc. A posttranslational process converts the cellular PrP isoform (PrPC) into PrPSc. Denatured PrPSc was digested with endoproteases, and the resulting fragments were isolated by HPLC. By both mass spectrometry and Edman sequencing, the primary structure of PrPSc was found to be the same as that deduced from the PrP gene sequence, arguing that neither RNA editing nor protein splicing feature in the synthesis of PrPSc. Mass spectrometry also was used to search for posttranslational chemical modifications other than the glycosylinositol phospholipid anchor attached to the C-terminus and two Asn-linked oligosaccharides already known to occur on both PrPSc and PrPC. These results contend that PrPSc molecules do not differ from PrPC at the level of an amino acid substitution or a posttranslational chemical modification; however, we cannot eliminate the possibility that a small fraction of PrPSc is modified by an as yet unidentified posttranslational process or that PrPC carries a modification that is removed in the formation of PrPSc. It seems likely that PrPSc differs from PrPC in its secondary and tertiary structure, but the possibility of a tightly bound, disease-specific molecule which purifies with PrPSc must also be considered.

Amino Acid Sequence↗
PubMed1993

Transgenic mice that express a myelin basic protein-specific T cell receptor develop spontaneous autoimmunity.

We constructed a transgenic mouse model that mimics the human autoimmune disease multiple sclerosis in its spontaneous induction and pathology. Transgenic mice were constructed expressing genes encoding a rearranged T cell receptor specific for myelin basic protein (MBP). T cell tolerance was not induced in the periphery, and functional, autoreactive T cells were found in the spleen and lymph nodes of these mice. Transgenic mice developed experimental allergic encephalomyelitis (EAE) following immunization with MBP and adjuvant plus pertussis toxin as well as with administration of pertussis toxin alone. Spontaneous EAE can develop in transgenic mice housed in a non-sterile facility but not in those maintained in a sterile, specific pathogen-free facility. This model system affords a unique opportunity to dissect the genetic and environmental variables that may contribute to the development of spontaneous autoimmune disease.

Animals↗
PubMed1993

Distinct hypomyelinated phenotypes in MBP-SV40 large T transgenic mice.

To study the effect of SV40 large T-antigen expression in myelin-forming cells of both the central and peripheral nervous system, a series of transgenic mice were generated expressing the SV40 large T-antigen under control of the myelin basic protein (MBP) promoter. Two neurologic phenotypes, designated A and B, appeared among individual transgenic founders and their progeny. The A mice developed a severe action tremor at about 10 days of age that progressed into periods of convulsions and early death by three to four weeks of age. In contrast, the B mice exhibited a progressive hindlimb ataxia and had a more normal lifespan. The A mice displayed hypomyelinating lesions in the central nervous system (CNS), whereas the B mice had lesions in either the peripheral nervous system (PNS) alone or in both the PNS and CNS. Immunohistochemical staining of spinal cord sections of a type A mouse showed a substantial depletion in MBP. Moreover, T-antigen-positive cells appeared predominantly in white matter tracts as randomly distributed single cells. Double labeling immunocytochemistry demonstrated that some of these T-antigen-positive cells were positive for oligodendrocyte differentiation markers MBP and O4. Thus, T-antigen expression appeared to coincide with a terminal stage of oligodendrocyte differentiation.

Animals↗
PubMed1993

Identification and physical mapping of a polymorphic human T cell receptor V beta gene with a frequent null allele.

Germline variation in genes that encode the human T cell receptors (TCRs) may have an important influence in shaping the immune T cell repertoire. In this report we describe a frequent null allele of the human V beta 18 gene, resulting from a nucleotide substitution that creates a stop codon (CGA<-->TGA). Approximately 11% of the population tested was homozygous for this null allele, indicating that this is a frequent "hole in the repertoire." We confirmed that there is a greatly reduced (undetectable) level of V beta 18 mRNA in peripheral blood lymphocytes from an individual homozygous for this null allele. In addition, all heterozygous individuals expressed detectable levels of only the functional V beta 18 allele in their peripheral blood lymphocytes. Two other DNA polymorphisms were identified in V beta 18, one of which would result in an amino acid substitution in an expressed V beta 18 gene. Genotypes for all three of these V beta 18 DNA polymorphisms were determined in a group of unrelated individuals. Statistical analyses of the associations between alleles of the V beta 18 polymorphisms and those of other DNA polymorphisms in the TCR beta locus suggested a close physical proximity between the V beta 18 gene and the 3' end of the C beta 2 region. This localization of human V beta 18 had been previously predicted by the sequence homology between human V beta 18 and mouse V beta 14, a V gene segment previously mapped to 3' of the mouse C beta genes. We confirmed this localization of the human V beta 18 gene by isolating a cosmid clone that contains both the V beta 18 and C beta 2 segments. Mapping by restriction enzyme digestion and by the polymerase chain reaction indicated that the V beta 18 gene segment is approximately 9 kb 3' of the C beta 2 gene, making this the only known human V beta gene 3' of the C beta region.

Alleles↗
PubMed1993

An experimentally derived data set constructed for testing large-scale DNA sequence assembly algorithms.

A data set consisting of DNA sequences from a large-scale shotgun DNA cloning and sequencing project has been collected and posted for public release. The purpose is to propose a standard genomic DNA sequencing data set by which various algorithms and implementations can be tested. This set of data is divided into two subsets, one containing raw DNA sequence data (1023 clones) and the other consisting of the corresponding partially refined or edited DNA sequence data (820 clones). Suggested criteria or guidelines for this data refinement are presented so that algorithms for preprocessing and screening raw sequences may be developed. Development of such preprocessing, screening, aligning, and assembling algorithms will expedite large-scale DNA sequencing projects so that the complete unambiguous consensus DNA sequences will be made available to the general research community in a quicker manner. Smaller scale routine DNA sequencing projects will also be greatly aided by such computational efforts.

Algorithms↗
PubMed1993

Organization of the murine T-cell receptor gamma locus.

Murine T cells carry either of two antigen receptors on their surface: alpha beta or gamma delta T-cell receptors (TCR). The function of gamma delta T cells is not well understood. In this population, subsets exist with tissue-specific localization, with invariant variable regions, and with specificity for self antigens. We have analyzed the murine TCR gamma locus by field inversion gel electrophoresis and cosmid cloning. The structure of the complete TCR gamma locus, the first complete physical map of any mouse TCR locus, is reported here. The locus spans 205 kb and consists of four clusters of genes and gene segments. Each cluster contains one or more variable elements, one joining element, and one constant element. We demonstrate that the C gamma 4 gene is distinguished from the other C gamma genes in that it has an additional hinge-encoding exon. New enhancer-like elements are identified in two of the clusters, downstream from the C gamma genes. The most upstream cluster has four V elements and resembles the human TCR gamma locus. The four clusters probably arose via duplications.

Animals↗
PubMed1993

Automated fluorescent DNA sequencing of polymerase chain reaction products.

The methods described in this chapter provide some useful approaches for DNA sequencing of templates produced by PCR. These procedures have been employed successfully for large-scale DNA sequencing of cosmid fragments subcloned in plasmid or M13 vectors, and for sequence analysis of cDNAs cloned in bacteriophage lambda vectors. In addition, the method describing direct sequencing from PEG-precipitated PCR product has been used successfully for analysis of Caenorhabditis elegans genomic and cDNA sequences. It is important to reiterate that for every combination of amplification primer pair and target DNA, there is an optimal method for PCR amplification; the ability to sequence the products of any PCR experiment directly will also vary. A coupled PCR/DNA sequencing method that works well for one experimental system may work quite poorly with others. Hence, a few days or hours spent optimizing PCR amplification conditions and selecting the best DNA sequencing method for the target DNA of interest will be time well spent.

Animals↗