Sparse distribution of γ/δ T lymphocytes around human epithelial tumors predominantly infiltrated by primed/memory T cells

Summary Evidence from the mouse system has suggested that T lymphocytes accumulating in non-lymphoid tissue, in particular epithelia, may preferentially express the T cell receptor (TCR) . In this study, we characterize the T cell receptor or phenotype of lymphocytes infiltrating human tumors of epithelial origin using monoclonal antibodies (mAb) for immunohistology and flow cytometry on cells extracted by enzyme digestion. This report shows that the majority of CD3⁺ tumor-infiltrating lymphocytes are TCR ⁺ but a small percentage of TCR can be clearly defined scattered throughout the tumor tissue with apparently no microanatomical selection. So far there has been little evidence for an accumulation of activated T cells in human tumor tissues as defined by mAb against molecules appearing transiently during the acute phase of activation. Now mAb are available that can identify primed or memory T cells such as mAb UCHL-1 recognizing the CD45RO antigen. Here we show that CD3⁺ tumor-infiltrating lymphocytes have a statistically significant accumulation of primed T cells, as compared to the autologous peripheral blood lymphocytes, suggesting their immune stimulation by tumor cells.     

No Orcadian Rhythms of Serotoninergic, Alpha-, Beta-Adrenergic and Imipramine Binding Sites in Rat Brain Regions

B_max values of the specific binding of [³H]-WB 4101, [³H]-dihydroalprenolol, [³H]-spiperone and [³H]-imipramine to various rat brain regions were determined at 4 hr intervals over 24 hr under circadian conditions. No significant circadian rhythm of binding sites number was found for any receptor investigated in cerebral cortex, hypothalamus or brain stem. Some methodological issues are discussed.     

The class I PITP giotto is required for Drosophila cytokinesis

Phosphatidylinositol transfer proteins (PITPs) are highly conserved polypeptides that bind phosphatidylinositol or phosphatidylcholine monomers, facilitating their transfer from one membrane compartment to another . Although PITPs have been implicated in a variety of cellular functions, including lipid-mediated signaling and membrane trafficking, the precise biological roles of most PITPs remain to be elucidated . Here we show for the first time that a class I PITP is involved in cytokinesis. We found that giotto (gio), a Drosophila gene that encodes a class I PITP, serves an essential function required for both mitotic and meiotic cytokinesis. Neuroblasts and spermatocytes from gio mutants both assemble regular actomyosin rings. However, these rings fail to constrict to completion, leading to cytokinesis failures. Moreover, gio mutations cause an abnormal accumulation of Golgi-derived vesicles at the equator of spermatocyte telophases, suggesting that Gio is implicated in membrane-vesicle fusion. Consistent with these results, we found that Gio is enriched at the cleavage furrow, the ER, and the spindle envelope. We propose that Gio mediates transfer of lipid monomers from the ER to the equatorial membrane, causing a specific local enrichment in phosphatidylinositol. This change in membrane composition would ultimately facilitate vesicle fusion, allowing membrane addition to the furrow and/or targeted delivery of proteins required for cytokinesis.     

Synthetic Lipid Vesicles Recruit Native-Like Aggregates and Affect the Aggregation Process of the Prion Ure2p: Insights on Vesicle Permeabilization and Charge Selectivity

The yeast prion Ure2p polymerizes into native-like fibrils, retaining the overall structure and binding properties of the soluble protein. Recently we have shown that, similar to amyloid oligomers, the native-like Ure2p fibrils and their precursor oligomers are highly toxic to cultured mammalian cells when added to the culture medium, whereas Ure2p amyloid fibrils generated by heating the native-like fibrils are substantially harmless. We show here that, contrary to the nontoxic amyloid fibrils, the toxic, native-like Ure2p assemblies induce a significant calcein release from negatively charged phosphatidylserine vesicles. A minor and less-specific effect was observed with zwitterionic phosphatidylcholine vesicles, suggesting that the toxic aggregates preferentially bind to negatively charged sites on lipid membranes. We also found that cholesterol-enriched phospholipid membranes are protected against permeabilization by native-like Ure2p assemblies. Moreover, vesicle permeabilization appears charge-selective, allowing calcium, but not chloride, influx to be monitored. Finally, we found that the interaction with phosphatidylserine membranes speeds up Ure2p polymerization into oligomers and fibrils structurally and morphologically similar to the native-like Ure2p assemblies arising in free solution, although less cytotoxic. These data suggest that soluble Ure2p oligomers and native-like fibrils, but not amyloid fibrils, interact intimately with negatively charged lipid membranes, where they allow selective cation influx.     

Colonization of heterochromatic genes by transposable elements in Drosophila

As a further step toward understanding transposable element-host genome interactions, we investigated the molecular anatomy of introns from five heterochromatic and 22 euchromatic protein-coding genes of Drosophila melanogaster. A total of 79 kb of intronic sequences from heterochromatic genes and 355 kb of intronic sequences from euchromatic genes have been used in Blast searches against Drosophila transposable elements (TEs). The results show that TE-homologous sequences belonging to 19 different families represent about 50% of intronic DNA from heterochromatic genes. In contrast, only 0.1% of the euchromatic intron DNA exhibits homology to known TEs. Intraspecific and interspecific size polymorphisms of introns were found, which are likely to be associated with changes in TE-related sequences. Together, the enrichment in TEs and the apparent dynamic state of heterochromatic introns suggest that TEs contribute significantly to the evolution of genes located in heterochromatin.     

The multidrug transporters belonging to major facilitator superfamily in Mycobacterium tuberculosis

BACKGROUND: Both intrinsic and acquired multidrug resistance play an important role in the insurgence of tuberculosis. Detailed knowledge of the molecular basis of drug recognition and transport by multidrug transport systems is required for the development of new antibiotics that are not extruded or of inhibitors that block the multidrug transporter and allow traditional antibiotics to be effective. MATERIALS AND METHODS: We have undertaken the inventory of the drug transporters subfamily, included in the major facilitator superfamily (MFS), encoded by the complete genome of Mycobacterium tuberculosis (MTB). These proteins were identified on the basis of their characteristic stretches of amino acids and transmembrane segments (TMS) number. CONCLUSIONS: Genome analysis and searches of homology between the identified transporters and proteins characterized in other organisms revealed 16 open reading frames encoding putative drug efflux pumps belonging to MFS. In the case of two of them, we also have demonstrated that they function as drug efflux proteins.     

Cytogenetic analysis of the third chromosome heterochromatin of Drosophila melanogaster

Previous cytological analysis of heterochromatic rearrangements has yielded significant insight into the location and genetic organization of genes mapping to the heterochromatin of chromosomes X, Y, and 2 of Drosophila melanogaster. These studies have greatly facilitated our understanding of the genetic organization of heterochromatic genes. In contrast, the 12 essential genes known to exist within the mitotic heterochromatin of chromosome 3 have remained only imprecisely mapped. As a further step toward establishing a complete map of the heterochomatic genetic functions in Drosophila, we have characterized several rearrangements of chromosome 3 by using banding techniques at the level of mitotic chromosome. Most of the rearrangement breakpoints were located in the dull fluorescent regions h49, h51, and h58, suggesting that these regions correspond to heterochromatic hotspots for rearrangements. We were able to construct a detailed cytogenetic map of chromosome 3 heterochromatin that includes all of the known vital genes. At least 7 genes of the left arm (from l(3)80Fd to l(3)80Fj) map to segment h49-h51, while the most distal genes (from l(3)80Fa to l(3)80Fc) lie within the h47-h49 portion. The two right arm essential genes, l(3)81Fa and l(3)81Fb, are both located within the distal h58 segment. Intriguingly, a major part of chromosome 3 heterochromatin was found to be "empty," in that it did not contain either known genes or known satellite DNAs.     

In silico determination of potential antisense targets for human beta-globin variants

The functional characterization of available genomic sequences is the major task of the research in the post-genome era. This complex task requires an integrative approach of high-throughput systems with in vitro and in vivo models in order to have a reliable evaluation of the biological function. The oligonucleotide antisense technology is one of the most promising approaches for the investigation of gene function; the crucial point of antisense experiments is the identification of optimal target sites for hybridisation. In this paper we have applied a bioinformatic tool for the recognition of optimal antisense targets. In order to evaluate the effect of mutational events on target selection we have tested the program on a sample of human beta-hemoglobin variants. The proposed algorithm software will be integrated in a web based tool at the site: http://www.nettab.org/agewa.