Pathogen-associated molecular patterns decisively influence antiviral immune responses, whereas the contribution of endogenous signals of tissue damage, also known as damage-associated molecular patterns or alarmins, remains ill defined. We show that interleukin-33 (IL-33), an alarmin released from necrotic cells, is necessary for potent CD8(+) T cell (CTL) responses to replicating, prototypic RNA and DNA viruses in mice. IL-33 signaled through its receptor on activated CTLs, enhanced clonal expansion in a CTL-intrinsic fashion, determined plurifunctional effector cell differentiation, and was necessary for virus control. Moreover, recombinant IL-33 augmented vaccine-induced CTL responses. Radio-resistant cells of the splenic T cell zone produced IL-33, and efficient CTL responses required IL-33 from radio-resistant cells but not from hematopoietic cells. Thus, alarmin release by radio-resistant cells orchestrates protective antiviral CTL responses.

Sequence similarity between aB-crystallin and small heat shock proteins (HSPs) has prompted us to investigate whether aB-crystallin expression is induced by heat shock. Indeed, accumulation of aB-crystallin was detected immunologically in NIH 3T3 cells after incubation at elevated temperatures and after addition of Cd2+ or sodium arsenite to these cells. Two-dimensional gel electrophoresis revealed identity between aB-crystallin from eye lenses and from heattreated fibroblasts. The promoter of the aB-crystallin gene was fused to the bacterial chloramphenicol acetyltransferase gene and was shown to confer heat inducibility on this reporter gene in transient transfection assays. A perfect heat shock element within the promoter region is likely to mediate this response. Small HSPs and aB-crystallin were shown to share the following two physical properties: (i) they form supramolecular structures with sedimentation values around 17 S and (ii) they are associated with the nucleus at high temperatures and are localized in the cytoplasm under normal conditions. We conclude that aB-crystallin has to be considered a member of the class of small HSPs.Heat shock and numerous other stress conditions lead to the rapid induction of several genes whose protein products are collectively called heat shock proteins (HSPs) (for recent reviews see refs. 1-3). The HSPs have been grouped into several classes on the basis of their size and sequence homology. Members of the class of small HSPs have molecular masses in the range 15-40 kDa. All analyzed organisms possess at least one small HSP gene. In mammals, birds, and yeast this class of HSPs is represented by a single member (4-9), whereas in Drosophila melanogaster and plants there appear to be multiple small HSPs (10, 11). Small HSPs aggregate to form characteristic ring-shaped structures called heat shock granules (4,(12)(13)(14)(15). These structures resemble prosomes or proteosomes but are distinct entities (16). Their biochemical function is unknown. Under heat shock conditions the small HSPs associate with the nucleus. Following a heat shock they slowly relocalize to the cytoplasm (17)(18)(19). It is still a matter of debate whether the small HSPs are actually transported into the nucleus at high temperatures or whether they are entrapped by the intermediate filaments, which, under heat shock conditions, collapse onto the nucleus (for a review see ref.3). The amino acid sequences of the small HSPs from different organisms are only poorly conserved. However, striking sequence similarities exist between vertebrate a-crystallins and small HSPs (5, 20-23). a-Crystallins were originally found in eye lenses, where they are among the most abundant proteins (24, 25). There exist two forms of a-crystallins, aA and aB, which are closely related (26). Considerable amounts of aB-crystallin, but not aAcrystallin, are present in many nonlenticular tissues (27-31). Moreover, aB-crystallin gene expression has been observed in various diseased cells, including astrocytes of patients s...

Genetic and molecular studies on the expression of Antennapedia (Antp) have suggested that this gene specifies mainly the second thoracic segment. On the basis of our molecular analysis of dominant gain-of-function mutants we have postulated that the transformation of antennae into second legs is due to the ectopic overexpression of the Antp+ protein. This hypothesis was tested by inserting the complementary DNA encoding the normal Antp protein into a heat-shock expression vector and subsequent germ-line transformation. As predicted, heat induction at defined larval stages leads to the transformation of antennae into second legs. The dorsal part of the head can also be transformed into second thoracic structures (scutum) indicating that Antp indeed specifies the second thoracic segment. By ectopic overexpression of the Antp protein the body plan of the fruit fly can be altered in a predictable way.

We describe new vectors suitable for P-element mediated germ line transformation of Drosophila melanogaster using passenger genes whose expression does not result in a readily detectable phenotypic change of the transformed flies. The P-element vectors contain the white gene fused to the heat shock protein 70 (hsp70) gene promoter. Expression of the white gene rescues the white phenotype of recipient flies partly or completely even without heat treatment. Transformed descendents of most founder animals (GO) fall into two classes which are distinguishable by their orange and red eye colours. The different levels of white expression are presumably due to position effects associated with different chromosomal sites of insertion. Doubling of the gene dose in orange eyed fly stocks results in an easily visible darkening of the eye colour. Consequently, the generation of homozygous transformants is easily possible by simple inbreeding due to the phenotypic distinction of homo- and heterozygous transformants. Cloning into these P-element vectors is facilitated by the presence of polylinkers with 8 and 12 unique restriction sites.

We describe the molecular identification of a gene, designated T1, whose expression in mouse NIH 3T3 cells is strongly induced by the Ha-ras(EJ) and v-mos oncogenes and by serum. The T1 gene encodes a 38.5-kDa protein, as predicted from its primary sequence and shown by in vitro translation. The protein was processed at its amino terminus and extensively modified by N-linked glycosylation in vitro in the presence of microsomal vesicles. Sequence comparison of T1 with the MIPSX data base (Max-Planck-Institut fir Biochemie, Martinsried/Munich) revealed similarity to the human carcinoembryonic antigen, a tumor marker which is overexpressed in colon adenocarcinomas and in fetal tissues.Considerable sequence similarity has also been observed to the short conserved region of other proteins which, like carcinoembryonic antigen, are encoded by members of the immunoglobulin gene superfamily.The binding of growth factors to their cell surface receptors induces a cascade ofevents which culminates in the induction of DNA synthesis and mitosis. The initiation of a mitogenic stimulus requires the expression of genes which presumably control these events (1). Oncogenically transformed cells exhibit a reduced requirement for growth factors. One might therefore expect oncogene expression to lead to the activation of those genes whose expression is also induced through the action of growth factors. Other genes are selectively induced by oncoproteins (ref. 2 and references therein), and their products are thought to bring about the transformed phenotype. We have tried to identify specific alterations in gene expression mediated by the conditional expression of the Ha-ras(EJ) and the v-mos oncogenes. We have previously isolated a gene, designated T1, whose expression is transiently stimulated by the two oncogene products as well as by serum growth factors in quiescent cells (32). Here we describe the molecular identification of the T1 gene. § It encodes a glycoprotein with sequence similarity to the carcinoembryonic antigen (CEA) and it is probably a member of the immunoglobulin gene superfamily. METHODSCell Lines and Culture Conditions. The NIH 3T3 cell line, stably transfected with the viral mos oncogene under the control of the mouse mammary tumor virus (MMTV) promoter (NIH[LTR v-mos]) has been described (3). Cells were grown in Dulbecco's modified Eagle's medium supplemented with 10% fetal calf serum. Growth arrest was induced by the addition of 1.5% fetal calf serum.RNA Preparation and Northern Blots. For the isolation of total RNA, cells were directly lysed in a suspension of 4 ml of 150 mM Tris-HCI, pH 9.0/5 mM EDTA/1% sodium dodecyl sulfate (SDS) and 8 ml of water-saturated, neutralized phenol on a 15-cm culture dish. This phenol extraction was followed by two phenol/chloroform extractions at 40C. The RNA was precipitated once in 2.5 M LiCl and once in 70% (vol/vol) ethanol. Total RNA (3 Ag) was glyoxylated, electrophoresed (33), blotted onto nitrocellulose membranes, prehybridized, and hybridized to a probe labeled...