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1.  Lung cancer in HIV-infected patients 
Purpose
Several studies have shown that HIV patients are at higher risk of lung cancer. Our aim is to analyse the prevalence and features of lung cancer in HIV-infected patients.
Methods
The clinical charts of 4,721 HIV-infected patients seen in three hospitals of southeast Spain (study period 1992–2012) were reviewed, and all patients with a lung cancer were analysed.
Results
There were 61 lung cancers, giving a prevalence of 1.2%. There was a predominance of men (82.0%), and smokers (96.6%; mean pack-years 35.2), with a median age of 48.0 (41.7–52.9) years, and their distribution according to risk group for HIV was: intravenous drug use 58.3%, homosexual 20.0%, and heterosexual 16.7%. Thirty-four (56.7%) patients were Aids cases, and 29 (47.5%) had prior pulmonar events: tuberculosis 16, bacterial pneumonia 9, and P. jiroveci pneumonia 4. The median nadir CD4 count was 149/mm3 (42–232), the median CD4 count at the time of diagnosis of the lung cancer was 237/mm3 (85–397), and 66.1%<350/mm3. 66.7% were on ART, and 70% of them had undetectable HIV viral load. The most common histological types of lung cancer were adenocarcinoma and epidermoid, with 24 (40.0%) and 23 (38.3%) cases, respectively. There were 49 (80.3%) cases with advanced stages (III and IV) at diagnosis. The distribution of treatments was: only palliative 23 (39.7%), chemotherapy 14 (24.1%), surgery and chemotherapy 8 (13.8%), radiotherapy 7 (12.1%), surgery 4 (6.9%), and other combined treatments 2 (3.4%). Forty-six (76.7%) patients died, with a median survival time of 3 months. The Kaplan-Meier survival rate at 6 months was 42.7% (at 12 months 28.5%).
Conclusions
The prevalence of lung cancer in this cohort of HIV-patients is high. People affected are mainly men, smokers, with transmission of HIV by intravenous drug use, and around half of them with prior opportunistic pulmonary events. Most patients had low nadir CD4 count, and were immunosuppressed at the time of diagnosis. Adenocarcinoma is the most frequent histological type. The diagnosis is usually made at advanced stages of the neoplasm, and mortality is high.
doi:10.7448/IAS.15.6.18087
PMCID: PMC3512461
2.  Syphilis in HIV-infected patients: predictors for serological failure and serofast state 
Purpose
HIV-infected patients treated for syphilis may be at increased risk for serological failure and serofast state. Our aim was to analyse serological response to treatment in HIV-infected patients diagnosed with syphilis, and factors associated with serological cure and serofast state.
Methods
Open-label, no controlled study of a series of HIV-patients diagnosed with syphilis during 2004–2011. Patients were categorized by rapid plasma reagin titer (RPR) into success (4-fold decrease in RPR by 12 or 24 months after treatment of early or late syphilis), serofast (success with persistently stable reactive RPR), and failure/re-infection (failure to decrease 4-fold in RPR by 12 or 24 months after treatment or sustained 4-fold increase in RPR after treatment response).
Results
141 HIV-patients were diagnosed with syphilis during the study period (104 early syphilis, 36 late or indeterminate latent syphilis). The mean age was 36.3 years, 98.5% were male, and 87.2% homosexual men. In 46 (32.6%) cases, HIV and syphilis infection diagnosis were coincident (mean CD4 457/mm3 and HIV-VL 4.72 log10). Among patients with prior known HIV infection, 65 were on antiretroviral therapy (ART) at syphilis diagnosis (mean CD4 469/mm3, 76.9% undetectable HIV-VL). 116 patients satisfied criteria for serological response analysis (89 early, 24 late/indeterminate). At 12 months of early syphilis treatment (89.2% penicillin) there were 16 (18%) failures, and at 24 months of late/indeterminate syphilis (91.7% penicillin) there were 5 (18.5%) failures. Overall, 36 (31.0%) patients presented serofast state. Treatment failure was related with lower CD4 count (295 vs 510/mL; p=0.045) only in patients with coincident diagnosis. Serofast state was related with older age (41 vs 36 years; p=0.024), and lower CD4 count (391 vs 513/mm3; p=0.026).
Conclusions
In this series of HIV-infected patients, with many patients on ART and with good immunological and virological parameters, serological failure and serofast state were frequent. Immunological status, and age could influence on serological response to syphilis treatment in HIV-infected patients.
doi:10.7448/IAS.15.6.18106
PMCID: PMC3512468
3.  Expression of endogenous retroviruses is negatively regulated by the pluripotency marker Rex1/Zfp42 
Nucleic Acids Research  2012;40(18):8993-9007.
Rex1/Zfp42 is a Yy1-related zinc-finger protein whose expression is frequently used to identify pluripotent stem cells. We show that depletion of Rex1 levels notably affected self-renewal of mouse embryonic stem (ES) cells in clonal assays, in the absence of evident differences in expression of marker genes for pluripotency or differentiation. By contrast, marked differences in expression of several endogenous retroviral elements (ERVs) were evident upon Rex1 depletion. We demonstrate association of REX1 to specific elements in chromatin-immunoprecipitation assays, most strongly to muERV-L and to a lower extent to IAP and musD elements. Rex1 regulates muERV-L expression in vivo, as we show altered levels upon transient gain-and-loss of Rex1 function in pre-implantation embryos. We also find REX1 can associate with the lysine-demethylase LSD1/KDM1A, suggesting they act in concert. Similar to REX1 binding to retrotransposable elements (REs) in ES cells, we also detected binding of the REX1 related proteins YY1 and YY2 to REs, although the binding preferences of the two proteins were slightly different. Altogether, we show that Rex1 regulates ERV expression in mouse ES cells and during pre-implantation development and suggest that Rex1 and its relatives have evolved as regulators of endogenous retroviral transcription.
doi:10.1093/nar/gks686
PMCID: PMC3467079  PMID: 22844087
6.  β-Phase Morphology in Ordered Poly(9,9-dioctylfluorene) Nanopillars by Template Wetting Method 
An efficient method based in template wetting is applied for fabrication of ordered Poly(9,9-dioctylfluorene) (PFO) nanopillars with β-phase morphology. In this process, nanoporous alumina obtained by anodization process is used as template. PFO nanostructures are prepared under ambient conditions via infiltration of the polymeric solution into the pores of the alumina with an average pore diameter of 225 nm and a pore depth of 500 nm. The geometric features of the resulting structures are characterized with environmental scanning electron microscopy (ESEM), luminescence fluorimeter (PL) and micro μ-X-ray diffractometer (μ-XRD). The characterization demonstrates the β-phase of the PFO in the nanopillars fabricated. Furthermore, the PFO nanopillars are characterized by Raman spectroscopy to study the polymer conformation. These ordered nanostructures can be used in optoelectronic applications such as polymer light-emitting diodes, sensors and organic solar cells.
doi:10.1007/s11671-010-9788-6
PMCID: PMC3211439
Template wetting; Nanoporous alumina; PFO; Nanopillars; Luminescence; Raman spectroscopy
7.  Discrete amplifiable regions (amplicons) in the symbiotic plasmid of Rhizobium etli CFN42. 
Journal of Bacteriology  1995;177(4):973-980.
Frequent tandem amplification of defined regions of the genome, called amplicons, is a common characteristic in the genomes of some Rhizobium species, such as Rhizobium etli. In order to map these zones in a model Rhizobium replicon, we undertook an analysis of the plasticity patterns fostered by amplicons in the pSym (390 kb) of R. etli CFN42. Data presented in this article indicate the presence of four amplicons in pSym, used for the generation of tandem amplifications and deletions. The amplicons are large, ranging from 90 to 175 kb, and they are overlapping. Each amplicon is usually flanked by specific reiterated sequences. Formation of amplifications and deletions requires an active recA gene. All the amplicons detected are concentrated in a zone of roughly one-third of pSym, covering most of the symbiotic genes detected in this plasmid. No amplicons were detected in the remaining two-thirds of pSym. These data support the idea that most of the known symbiotic genes in this plasmid are located in a genomic region that is prone to the formation of frequent tandem amplification.
PMCID: PMC176691  PMID: 7860608
8.  Different plasmids of Rhizobium leguminosarum bv. phaseoli are required for optimal symbiotic performance. 
Journal of Bacteriology  1992;174(16):5183-5189.
Rhizobium leguminosarum bv. phaseoli CFN42 contains six plasmids (pa to pf), and pd has been shown to be the symbiotic plasmid. To determine the participation of the other plasmids in cellular functions, we used a positive selection scheme to isolate derivatives cured of each plasmid. These were obtained for all except one (pe), of which only deleted derivatives were recovered. In regard to symbiosis, we found that in addition to pd, pb is also indispensable for nodulation, partly owing to the presence of genes involved in lipopolysaccharide synthesis. The positive contribution of pb, pc, pe, and pf to the symbiotic capacity of the strain was revealed in competition experiments. The strains that were cured (or deleted for pe) were significantly less competitive than the wild type. Analysis of the growth capacity of the cured strains showed the participation of the plasmids in free-living conditions: the pf- strain was unable to grow on minimal medium, while strains cured of any other plasmid had significantly reduced growth capacity in this medium. Even on rich medium, strains lacking pb or pc or deleted for pe had a diminished growth rate compared with the wild type. Complementation of the cured strains with the corresponding wild-type plasmid restored their original phenotypes, thus confirming that the effects seen were due only to loss of plasmids. The results indicate global participation of the Rhizobium genome in symbiotic and free-living functions.
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PMCID: PMC206350  PMID: 1644746
9.  Fetal liver pro-B and pre-B lymphocyte clones: expression of lymphoid-specific genes, surface markers, growth requirements, colonization of the bone marrow, and generation of B lymphocytes in vivo and in vitro. 
Molecular and Cellular Biology  1992;12(2):518-530.
We describe here the development and characterization of the FLS4.1 stromal line derived from 15-day fetal liver of BALB/c embryos and defined culture conditions that efficiently support the cloning and long-term growth of nontransformed B-220+ 14-day fetal liver cells at two stages of B-cell development, namely, pro-B lymphocytes (immunoglobulin [Ig] genes in germ line configuration) and pre-B cells (JH-rearranged genes with both light-chain Ig genes in the germ line state). All B-cell precursor clones require recombinant interleukin-7 (rIL-7) and FLS4.1 stromal cells for continuous growth in culture, but pro-B lymphocyte clones can also proliferate in rIL-3. None proliferate in rIL-1, rIL-2, rIL-4, rIL-5, rIL-6, or leukemia inhibitory factor. FLS4.1 stromal cells synthesize mRNA for Steel factor but not for IL-1 to IL-7; all pro-B and pre-B clones express c-Kit, the receptor for Steel factor, and a c-Kit-specific antibody inhibits the enhanced proliferative response of fetal liver B-220+ B-cell precursors supported by FLS4.1 stromal cells and exogenous rIL-7 but does not affect that promoted by rIL-7 alone. Northern (RNA) blot analysis of the expression of the MB-1, lambda 5, Vpre-B, c mu, RAG-1, and RAG-2 genes in pro-B and pre-B clones show that transcription of the MB-1 gene precedes IgH gene rearrangement and RNA synthesis from c mu, RAG-1, RAG-2, lambda 5, and Vpre-B genes. All clones at the pre-B-cell stage synthesize mRNA for c mu, RAG-1, and RAG-2 genes; transcription of the lambda 5 and Vpre-B genes seems to start after D-to-JH rearrangement in B-cell precursors, indicating that the proteins encoded by either gene are not required for B-cell progenitors to undergo D-to-JH gene rearrangement. These findings mark transcription of the MB-1 gene as one of the earliest molecular events in commitment to develop along the B-lymphocyte pathway. Indeed, both pro-B and pre-B clones can generate in vitro and in vivo B lymphocytes but not T lymphocytes; moreover, these clones do not express the CD3-gamma T-cell-specific gene, nor do they have rearranged gamma, delta, or beta T-cell antigen receptor genes.
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PMCID: PMC364211  PMID: 1346335
10.  Amplification and deletion of a nod-nif region in the symbiotic plasmid of Rhizobium phaseoli. 
Journal of Bacteriology  1991;173(8):2435-2441.
One remarkable characteristic of the genomes of some Rhizobium species is the frequent occurrence of rearrangements. In some instances these rearrangements alter the symbiotic properties of the strains. However, no detailed molecular mechanisms have been proposed for the generation of these rearrangements. To understand the mechanisms involved in the formation of rearrangements in the genome of Rhizobium phaseoli, we have designed a system which allows the positive selection for amplification and deletion events. We have applied this system to investigate the stability of the symbiotic plasmid of R. phaseoli. High-frequency amplification events were detected which increase the copy number of a 120-kb region carrying nodulation and nitrogen fixation genes two to eight times. Deletion events that affect the same region were also found, albeit at a lower frequency. Both kinds of rearrangements are generated by recombination between reiterated nitrogenase (nifHDK) operons flanking the 120-kb region.
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PMCID: PMC207805  PMID: 2013567
11.  Structural complexity of the symbiotic plasmid of Rhizobium leguminosarum bv. phaseoli. 
Journal of Bacteriology  1991;173(8):2411-2419.
The complete physical map of the symbiotic plasmid of Rhizobium leguminosarum bv. phaseoli strain CFN42 was established. The data support the concept that Rhizobium symbiotic genes are part of a complex genomic structure which contains a large amount of reiterated DNA sequences. This plasmid is a circular structure of 390 kb with approximately 10 families of internally reiterated DNA sequences of two to three elements each. One family includes two directly oriented nitrogenase operons situated 120 kb apart. We also found several stretches of pSym that are reiterated in other replicons of the cell. Localization of symbiotic gene sequences by heterologous hybridization revealed that nodABC sequences are separated in two regions, each of which contains a nod boxlike element, and it also suggested the presence of two copies of the nifA and nodD gene sequences. We propose that the complex structure of the symbiotic plasmid allows interactions between repeated DNA sequences which, in turn, might result in frequent rearrangements.
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PMCID: PMC207802  PMID: 2013564
12.  Genetic structure of a soil population of nonsymbiotic Rhizobium leguminosarum. 
The genetic structure of a population of nonsymbiotic Rhizobium leguminosarum strains was determined by the electrophoretic mobilities of eight metabolic enzymes. Nonsymbiotic strains were isolated from the rhizosphere of bean plants and characterized by growth on differential media and at different temperatures, intrinsic antibiotic resistance, the lack of homology to a nifH probe, and their inability to form nodules on bean roots. All the isolates clustered with R. leguminosarum bv. phaseoli reference strains and did not encompass any other Rhizobium taxa. Their rRNA operon restriction fragment length polymorphisms and the nucleotide sequence of a fragment of the 16S rRNA gene were also found to be identical to those of R. leguminosarum bv. phaseoli reference strains. When complemented with an R. leguminosarum bv. phaseoli symbiotic plasmid (p42d), the nonsymbiotic isolates were able to fix nitrogen in symbiosis with bean roots at levels similar to those of the parental strain. The symbiotic isolates were found at a relative frequency of 1 in 40 nonsymbiotic R. leguminosarum strains.
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PMCID: PMC182727  PMID: 1707606
13.  High-frequency rearrangements in Rhizobium leguminosarum bv. phaseoli plasmids. 
Journal of Bacteriology  1991;173(3):1344-1346.
High-frequency genomic rearrangements affecting the plasmids of Rhizobium leguminosarum bv. phaseoli CFN42 were analyzed. This strain contains six large plasmids ranging in size from 200 to 600 kb. In the absence of any selective pressure, we found 11 strains from 320 analyzed colonies that presented different kinds of plasmid-borne rearrangements, including sequence amplification, deletion, cointegration, and loss of plasmids. These data support the concept that the R. leguminosarum bv. phaseoli genome is a dynamic structure and imply that strains are mixtures of similar but not identical cells.
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PMCID: PMC207262  PMID: 1991727
14.  Genomic instability in Rhizobium phaseoli. 
Journal of Bacteriology  1988;170(3):1191-1196.
Experience from different laboratories indicates that Rhizobium strains can generate variability in regard to some phenotypic characteristics such as colony morphology or symbiotic properties. On the other hand, several reports suggest that under certain stress conditions or genetic manipulations Rhizobium cells can present genomic rearrangements. In search of frequent genomic rearrangements, we analyzed three Rhizobium strains under laboratory conditions that are not considered to cause stress in bacterial populations. DNAs from direct descendants of a single cell were analyzed in regard to the hybridization patterns obtained, using as probes different recombinant plasmids or cosmids; while most of the probes utilized did not show differences in the hybridization patterns, some of them revealed the occurrence of frequent genomic rearrangements. The implications and possible biological significance of these observations are discussed.
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PMCID: PMC210891  PMID: 3343217
15.  Reiterated DNA sequences in Rhizobium and Agrobacterium spp. 
Journal of Bacteriology  1987;169(12):5782-5788.
Repeated DNA sequences are a general characteristic of eucaryotic genomes. Although several examples of DNA reiteration have been found in procaryotic organisms, only in the case of the archaebacteria Halobacterium halobium and Halobacterium volcanii [C. Sapienza and W. F. Doolittle, Nature (London) 295:384-389, 1982], has DNA reiteration been reported as a common genomic feature. The genomes of two Rhizobium phaseoli strains, one Rhizobium meliloti strain, and one Agrobacterium tumefaciens strain were analyzed for the presence of repetitive DNA. Rhizobium and Agrobacterium spp. are closely related soil bacteria that interact with plants and that belong to the taxonomical family Rhizobiaceae. Rhizobium species establish a nitrogen-fixing symbiosis in the roots of legumes, whereas Agrobacterium species is a pathogen in different plants. The four strains revealed a large number of repeated DNA sequences. The family size was usually small, from 2 to 5 elements, but some presented more than 10 elements. Rhizobium and Agrobacterium spp. contain large plasmids in addition to the chromosomes. Analysis of the two Rhizobium strains indicated that DNA reiteration is not confined to the chromosome or to some plasmids but is a property of the whole genome.
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PMCID: PMC214138  PMID: 3450286
16.  Nitrogen-fixing nodules induced by Agrobacterium tumefaciens harboring Rhizobium phaseoli plasmids. 
Journal of Bacteriology  1987;169(6):2828-2834.
Rhizobium phaseoli CFN299 forms nitrogen-fixing nodules in Phaseolus vulgaris (bean) and in Leucaena esculenta. It has three plasmids of 185, 225, and 410 kilobases. The 410-kilobase plasmid contains the nitrogenase structural genes. We have transferred these plasmids to the plasmid-free strain Agrobacterium tumefaciens GMI9023. Transconjugants containing different combinations of the R. phaseoli plasmids were obtained, and they were exhaustively purified before nodulation was assayed. Only transconjugants harboring the 410-kilobase plasmid nodulate P. vulgaris and L. esculenta. Nodules formed by all such transconjugants are able to reduce acetylene. Transconjugants containing the whole set of plasmids from CFN299 nodulate better and fix more nitrogen than the transconjugants carrying only the Sym plasmid. Microscopic analysis of nodules induced by A. tumefaciens transconjugants reveals infected cells and vascular bundles. None of the A. tumefaciens transconjugants, not even the one with the whole set of plasmids from CFN299, behaves in symbiosis like the original R. phaseoli strain; the transconjugants produce fewer nodules and have lower acetylene reduction (25% as compared to the original R. phaseoli strain) and more amyloplasts per nodule. More than 2,000 bacterial isolates from nodules of P. vulgaris and L. esculenta formed by the transconjugants were analyzed by different criteria. Not a single rhizobium could be detected. Our results show that R. phaseoli plasmids may be expressed in the A. tumefaciens background and direct the formation of effective, differentiated nodules.
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PMCID: PMC212195  PMID: 3584072
17.  Abelson virus abrogation of interleukin-3 dependence in a lymphoid cell line. 
Molecular and Cellular Biology  1986;6(11):4133-4135.
Among several tyrosine-protein kinases, only v-abl could abrogate interleukin 3 dependence of a lymphoblastoid cell line; v-src and v-fps proteins gave partial or no interleukin 3 independence, respectively. Lymphokine independence was achieved via a nonautocrine mechanism. Direct involvement of c-myc in this process was not evident.
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PMCID: PMC367185  PMID: 3025637
18.  Genetic and biochemical characterization of glutamine synthetase from Neurospora crassa glutamine auxotrophs and their revertants. 
Journal of Bacteriology  1983;156(3):993-1000.
In this paper we present the isolation and characterization of glutamine auxotrophs of Neurospora crassa and their revertants. The results show that although various enrichment procedures were used, we found only two types of auxotrophs. Genetic crosses performed between the different mutants showed that the mutations responsible for their phenotypes were highly linked and probably affected the same gene. The biochemical characterization of the glutamine synthetase polypeptides of the different mutants showed that both types contained the alpha monomer. However, in place of the normal beta monomer, each type had a new polypeptide differing from normal beta either in its molecular weight or in its isoelectric point. On the other hand, the revertants had only the alpha monomer and were capable of growing without glutamine. On the basis of these data, we propose that the lack of glutamine synthetase activity in the auxotrophs is due to the interaction of the altered beta with the alpha monomer, and as a consequence the alpha monomer of the revertants regains its activity because of the absence of the altered beta.
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PMCID: PMC217941  PMID: 6139363
19.  Physiology of ammonium assimilation in Neurospora crassa. 
Journal of Bacteriology  1982;150(1):105-112.
In Neurospora crassa the assimilation of high and low concentrations of ammonium occurs by two different pathways. When the fungi are growing exponentially on ammonium excess, this compound is fixed by a glutamic dehydrogenase and an octameric glutamine synthetase (GS). The synthesis of this GS polypeptide (beta) is regulated by the nitrogen source present in excess; being higher on glutamate, intermediate on ammonium, and lower on glutamine. When N. crassa is growing in fed-batch ammonium-limited cultures a different polypeptide of GS (alpha), arranged as a tetramer, is synthesized. In both conditions synthesis in vivo correlates with the data obtained with an in vitro translation system primed with N. crassa RNA. This different expression of alpha and beta GS polypeptides was also observed when the cultures were shifted from excess to low nitrogen, and vice versa. By agarose gel electrophoresis in the presence of methylmercury hydroxide, some separation of different mRNAs that direct the in vitro synthesis of alpha and beta GS polypeptides has been accomplished. Data are presented that establish the operation of the tetrameric alpha GS and of glutamate synthase in the assimilation of ammonium in low concentration.
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PMCID: PMC220087  PMID: 6120927
20.  T cell growth factor abrogates concanavalin A-induced suppressor cell function 
The Journal of Experimental Medicine  1981;153(5):1360-1365.
Concanavalin A (Con-A)-induced suppressor T cells were found to respond to T cell growth factor (TCGF) by proliferation. TCGF abrogated the suppressor activity exerted by these cells on phytohemagglutinin (PHA)- and alloantigen- induced lymphocyte proliferation and on pokeweed mitogen (PWM)-driven immunoglobulin secretion. The Con-A-activated suppressor T cells absorbed the TCGF activity, preincubation of these active suppressor cells with TCGF abolished their suppressor activity and addition of increasing numbers of Con-A-activated T cells reverted the abrogator,/ effect of TCGF. Altogether, these findings suggest that Con-A-induced suppressor T cells exert their function by decreasing the available levels of TCGF. Cyclosporin-A (CYA), which is known to inhibit the expression of receptors for TCGF on T cells, also inhibited the suppressor activity as determined in both indicator systems, namely PHA- or alloantigen-induced DNA synthesis and PWM-induced immunoglobulin synthesis. CYA made Con-A-treated T cells unresponsive to TCGF and unable to absorb the growth factor, supporting the notion that CYA inhibits the expression of TCGF receptors on T cells, a mechanism by which this drug seems to abrogate Con-A-induced suppressor T cell function.
PMCID: PMC2186149  PMID: 6454746
21.  Autologous rosette-forming T cells as the responding cells in human autologous mixed-lymphocyte reaction. 
Journal of Clinical Investigation  1980;65(6):1527-1530.
Autologous rosette-forming cells (Tar cells) have surface and functional characteristics of post-thymic precursors and among these characteristics there are some that have been identified in the responsive cell of the autologous mixed-lymphocyte reaction (AMLR). We therefore did AMLR with circulating mononuclear cells from normal subjects using as responding cells either total T cells, T cells depleted of Tar cells, or purified Tar cells. The response of Tar cells in AMLR was significantly greater than that of total T cells and these responded significantly more than Tar-depleted T cells. Conversely, Tar cells responded less than total T cells or T cells depleted of Tar cells in allogeneic mixed-lymphocyte reactions. Increasing numbers of Tar cells gave significantly greater AMLR responses both alone and when added to diminishing proportions of Tar-depleted T cells to keep the number of T cells constant in the system. Tar cells are the responding cells in AMLR but not in allogeneic mixed-lymphocyte reactions.
PMCID: PMC371493  PMID: 6447710
22.  Immunochemical characterization of glutamine synthetase from Neurospora crassa glutamine auxotrophs. 
Journal of Bacteriology  1979;139(2):537-543.
Glutamine synthetase derived from two Neurospora crassa glutamine auxotrophs was characterized. Previous genetic studies indicated that the mutations responsible for the glutamine auxotrophy are allelic and map in chromosome V. When measured in crude extracts, both mutant strains had lower glutamine synthetase specific activity than that found in the wild-type strain. The enzyme from both auxotrophs and the wild-type strain was partially purified from cultures grown on glutamine as the sole nitrogen source, and immunochemical studies were performed in crude extracts and purified fractions. Quantitative rocket immunoelectrophoresis indicated that the activity per enzyme molecule is lower in the mutants than in the wild-type strain; immunoelectrophoresis and immunochemical titration of enzyme activity demonstrated structural differences between the enzymes from both auxotrophs. On the other hand, the monomer of glutamine synthetase of both mutants was found to be of a molecular weight similar to that of the wild-type strain. These data indicate that the mutations are located in the structural gene of N. crassa glutamine synthetase.
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PMCID: PMC216901  PMID: 37239
23.  Nitrogen source regulates glutamine synthetase mRNA levels in Neurospora crassa. 
Journal of Bacteriology  1978;136(3):880-885.
Neurospora crassa glutamine synthetase mRNA was measured by its capacity to direct the synthesis of the specific protein in a cell-free system derived from rabbit reticulocytes. N. crassa cultures grown on glutamate as the sole nitrogen source had higher mRNA activities than did those grown on glutamine. The differences were about 10-fold when polysomal RNA was used for translation and about 5-fold when either total cellular RNA or polyadenylic acid-enriched cellular RNA was used. These data indicate that in exponentially growing N. crassa, the nitrogen source regulates glutamine synthetase by adjusting specific mRNA levels.
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PMCID: PMC218521  PMID: 31352
24.  Genetics and physiology of Neurospora crassa glutamine auxotrophs. 
Journal of Bacteriology  1978;134(3):693-698.
This work reports on the isolation and characterization of two glutamine auxotrophs in Neurospora crassa. The mutations responsible for the glutamine-requiring phenotype were very closely linked, and one of them proved to be recessive to wild type. The mutations impaired the conversion of glutamic acid to glutamine and resulted in changes of both the activity and oligomeric structure of the enzyme glutamine synthetase.
PMCID: PMC222312  PMID: 26664
25.  THE COMPLEMENT FIXATION TEST IN THE DIAGNOSIS OF VIRUS INFECTIONS OF THE CENTRAL NERVOUS SYSTEM 
A specific complement fixation test can be obtained in various central nervous system virus infections by using as antigens emulsions of infected brain tissue, freezing and thawing the brain emulsion, and then centrifuging it in an angle head centrifuge at 3500 R.P.M. for 1 hour. The method has proved reliable in the case of rabies, St. Louis encephalitis, Japanese B encephalitis, lymphocytic choriomeningitis, Eastern equine encephalomyelitis, Western equine encephalomyelitis, louping ill, and spontaneous encephalomyelitis of mice (Theiler's disease). The specificity of the reaction, regardless of the virus involved, requires different temperatures of inactivation of the sera according to animal species: 56°C. for guinea pig, 60°C. for mouse, and 65°C. for rabbit and dog sera, all heated for 20 minutes. For human sera a temperature of inactivation of 60°C. also for 20 minutes has been adopted; at this temperature the reaction is in general specific. Complement-fixing antibodies in high titre were found in the sera of rabbits, guinea pigs, mice, and dogs immunized with rabies virus. Complement-fixing antibodies were present in high titre in sera drawn from two persons 8 years after an attack of louping ill, from five persons 2½ years after an attack of Eastern equine encephalomyelitis, and from two persons 2½ years after Western equine encephalomyelitis. In cases of St. Louis encephalitis and lymphocytic choriomeningitis, complement-fixing antibodies have been found shortly following infection but not after long periods.
PMCID: PMC2135201  PMID: 19871144

Results 1-25 (25)