Antitumour activities of levans produced by Zymomonas mobilis strains

Levans produced by four Zymomonas mobilis strains showed antitumour activity against sarcoma 180 and Ehrlich carcinoma in Swiss albino mice. Levans from two strains (ZAP and CP4) had the highest effects. NMR analysis showed that the polymers were composed only of fructose units. The results suggested that the antineoplasic effect is associated to the polysaccharide molecular weight and that a particular molecular weight range may be responsible for this effect.     

Perforin is present only in normal activated Lyt2+ T lymphocytes and not in L3T4+ cells, but the serine protease granzyme A is made by both subsets.

We show that among the subsets of peripheral T lymphocytes (Lyt2+ L3T4- and L3T4+ Lyt2- cells) activated in short-term cultures by stimulation with H-2 incompatible leukocytes 97% of the cytolytic activity and all detectable perforin activity resides in the Lyt2+ cells. But both populations contain approximately equal amounts of a serine protease, granzyme A, the expression of which was previously thought to be restricted to cytolytic T lymphocytes.     

Cell specificity of granzyme gene expression

Granzymes are serine proteases present in secretory granules of cytolytic T lymphocyte lines. We have studied the expression of the granzyme family (granzyme A, B, C, D, E, F, and G) in different lymphoid cell populations and cell lines as well as in nonlymphoid cells and tissues. Our data show that with few exceptions expression of granzyme genes is restricted to T cells and their thymic precursors. In mature T cells granzymes are expressed only upon activation. The same is true for thymocytes, with the exception of grazyme A that is expressed also in non-stimulated cells. In T cells and thymocytes the distribution of mRNAs coding for different granzymes depends on the subpopulation tested and the activation protocol. Highly cytolytic PEL express granzymes A and B but none of the other granzymes.     

Activity and interleukin 1 responsiveness of SV40 enhancer motifs in a rodent immature T cell line.

We have analysed the enhancer activity and the interleukin 1 (IL1) responsiveness of individual motifs of the SV40 enhancer in an immature rodent T cell line, PC60. Transient transfection assays showed that tetramers of GT-I plus GT-IIC motifs, the TC-II or the P motif have significant enhancer activity in PC60, while neither Octamer nor SphI+II motifs have a detectable effect on promoter strength. Two motifs, TC-II and P, strongly respond to stimulation by IL1. DNase I and methylation protection experiments with nuclear extracts show specific footprints in the TC-II region of the SV40 enhancer. Exposure of PC60 cells to IL1 increases their intensity. The TC-II sequence forms several complexes detected in band shift assays. The molecules involved all have similar sequence specificity as NF-kappa B. Surprisingly, band shifts with extracts from control or IL1 treated cells differ only slightly. However, if GTP is added to the binding reactions the intensity of bands formed by extracts from control cells is strongly reduced, whereas extracts from IL1 treated cells form a single retarded complex that co-migrates with NF-kappa B from a pre-B cell line. The results suggest that in PC60 IL1 induces NF-kappa B activity by activating molecules that are already in the nucleus.     

Variation in the ribosomal internal transcribed spacers (ITS1 and ITS2) among eight taxa of the Mimulus guttatus species complex

The internal transcribed spacer region (ITS1 and ITS2) of the 18S-25S nuclear ribosomal DNA sequence and the intervening 5.8S region were sequenced from three individuals in each of eight taxa of the Mimulus guttatus species complex. Three discrete variants, or "types," of ITS sequences were found, among which 30%-40% of sites differed, compared with 1%-2% within types. Dot plots indicate that these types were not related by conspicuous rearrangements or inversions. More than one ITS type was often found in the same taxon, and two of three ITS types span species boundaries, indicating their presence prior to speciation. These ITS sequences showed essentially no positional homology with the nearest sequenced relative, tomato. In contrast, the 5.8S region was relatively unvaried, with 8 of 162 sites varied in the sample among all eight taxa. The phylogeny inferred by the most common ITS sequence type, rooted by the two other ITS types, agreed with isozymes in showing the distinctness of M. nudatus, M. laciniatus, and M. tilingii from the other five taxa.      

Variation in heat shock proteins within tropical and desert species of poeciliid fishes

The 70-kilodalton heat shock protein (hsp70) family of molecular chaperones, which contains both stress-inducible and normally abundant constitutive members, is highly conserved across distantly related taxa. Analysis of this protein family in individuals from an outbred population of tropical topminnows, Poeciliopsis gracilis, showed that while constitutive hsp70 family members showed no variation in protein isoforms, inducibly synthesized hsp70 was polymorphic. Several species of Poeciliopsis adapted to desert environments exhibited lower levels of inducible hsp70 polymorphism than the tropical species, but constitutive forms were identical to those in P. gracilis, as they were in the confamilial species Gambusia affinis. These differences suggest that inducible and constitutive members of this family are under different evolutionary constraints and may indicate differences in their function within the cell. Also, northern desert species of Poeciliopsis synthesize a subset of the inducible hsp70 isoforms seen in tropical species. This distribution supports the theory that ancestral tropical fish migrated northward and colonized desert streams; the subsequent decrease in variation of inducible hsp70 may have been due to genetic drift or a consequence of adaptation to the desert environment. Higher levels of variability were found when the 30- kilodalton heat shock protein (hsp30) family was analyzed within different strains of two desert species of Poeciliopsis and also in wild-caught individuals of Gambusia affinis. In both cases the distribution of hsp30 isoform diversity was similar to that seen previously with allozyme polymorphisms.      

In vitro secondary MLR. I. Kinetics of proliferation and specificity of in vitro primed responder cells.

We have examined the kinetics and specificity of secondary in vitro mixed lymphocyte reactions (MLR). With limited numbers of primed responder cells (PRC) in the presence of "excess antigen" it was possible to obtain proliferative responses that were proportional to the number of PRC initially placed in culture. The responding cells, after an initial lag period, seem to grow exponentially until day 3 of culture. The responses of PRC (with the strain combinations and culture conditions described in this report) seemed to be directed toward stimulator cell determinants whose expression was determined by genes in the I region of the MHC. In one case, the relevant incompatibilities could be further restricted to the I-A region. Although PRC responded best to stimulator cells sharing the I region with the priming stimulator cell, apparent cross-reactivity could be observed by restimulating PRC with stimulator cells that did not carry the MHC haplotype of the priming stimulator cell. The rate of proliferation (measured as 3H-thymidine incorporation) in these apparent cross-reactions was reproducible and comparable to the rate observed in response to the priming stimulator cell. It was possible, therefore, to estimate the proportion of PRC that reacted in the presence of third party stimulator cells compared to the response of these PRC to the priming stimulator cells. We have estimated that the response of A (B6) PRC against H-2d and H-2s haplotype stimulator cells is about half of the response of these PRC to H-2b, the priming stimulator cell.     

Characterization of cloned murine cytolytic T cell lines

Murine cytolytic T lymphocytes can be kept in continuous culture apparently indefinitely by repeated passage in a concanavalin A-induced growth promoting medium. Some of these long-term cell lines maintain their cytolytic activity. Starting from three such populations, several cloned cytolytic T cell lines were derived and subsequently subcloned one or more times. Considerable variation in the levels of cytolytic activity was observed between different subclones; some initially active subclones lost activity with prolonged culture. In addition, one of the clones appeared to progressively lose the relative specificity demonstrated during the earlier passages of the parent cell line.