In T cells, cyclosporin A (CsA) exerts its immunosuppressive effect by preventing transcriptional induction of the expression of interleukin(IL)-2. This is achieved by a mechanism that involves binding of a CsA-cyclophilin complex to calcineurin, which in turn inhibits the phosphatase-controlled translocation of transcription factor NFAT to the nucleus. We have previously identified IL-3 as an autocrine oncogenic regulator in tumour cell lines generated by introducing the v-H-ras oncogene into IL-3-dependent mast cells. Here we report that CsA specifically blocks autocrine tumour cell growth. The mechanism involves down-regulation of IL-3 expression by destabilization of the messenger RNA and requires ongoing transcription. Transcripts from exogenous IL-3 genes lacking the (A+U)-rich element (ARE) in the 3' untranslated terminal repeat could not be destabilized, suggesting that at least part of this sequence, which is known to mediate decay of short-lived mRNA, participates in a CsA-sensitive regulatory mechanism.

Autocrine interleukin 3 (IL-3)-secreting tumors were generated from an IL-3-dependent mouse mast cell line (PB-3c) after introduction of the v-H-ras oncogene. Tumor progression was characterized by four distinct phenotypes. The first corresponded to immortalized mast cells unresponsive to the oncogenic effect of v-H-ras. The second was expressed in a clonable subpopulation of PB-3c cells and was marked by the competence to form v-H-ras-dependent tumors (immortalized transformation competence). The third was a direct effect of v-H-ras expression on all PB-3c cells and was characterized in vitro by a reduced IL-3 requirement. Upon injection of v-H-ras-expressing, transformation-competent cells into mice, the final, fully malignant phenotype developed with a long latency period and was marked in vitro by independence of exogenous IL-3 and by autocrine IL-3 stimulation. Northern (RNA) blot analysis and an RNase A-T1 protection assay showed that IL-3 production was strictly associated with the tumor phenotype. Two of six tumors showed an alteration at the 5' region of the IL-3 gene. We conclude that v-H-ras required complementation by IL-3 gene rearrangement or an alternate event to generate autocrine mastocytomas.

Critical illness in COVID-19 is an extreme and clinically homogeneous disease phenotype that we have previously shown1 to be highly efficient for discovery of genetic associations2. Despite the advanced stage of illness at presentation, we have shown that host genetics in patients who are critically ill with COVID-19 can identify immunomodulatory therapies with strong beneficial effects in this group3. Here we analyse 24,202 cases of COVID-19 with critical illness comprising a combination of microarray genotype and whole-genome sequencing data from cases of critical illness in the international GenOMICC (11,440 cases) study, combined with other studies recruiting hospitalized patients with a strong focus on severe and critical disease: ISARIC4C (676 cases) and the SCOURGE consortium (5,934 cases). To put these results in the context of existing work, we conduct a meta-analysis of the new GenOMICC genome-wide association study (GWAS) results with previously published data. We find 49 genome-wide significant associations, of which 16 have not been reported previously. To investigate the therapeutic implications of these findings, we infer the structural consequences of protein-coding variants, and combine our GWAS results with gene expression data using a monocyte transcriptome-wide association study (TWAS) model, as well as gene and protein expression using Mendelian randomization. We identify potentially druggable targets in multiple systems, including inflammatory signalling (JAK1), monocyte–macrophage activation and endothelial permeability (PDE4A), immunometabolism (SLC2A5 and AK5), and host factors required for viral entry and replication (TMPRSS2 and RAB2A).

We analyzed the effect of rapamycin on autocrine mast cell tumor lines with abnormally stable interleukin-3 (IL-3) transcripts due to a defect in mRNA degradation. Rapamycin inhibited IL-3 mRNA expression specifically, while transcripts of IL-4 and IL-6 were not affected. As indicated by the use of the transcriptional inhibitor actinomycin D or by reporter constructs, inhibition was posttranscriptional and resulted from destabilization of the mRNA. Transcripts from transgenes lacking the AU-rich 3 untranslated region were refractory to drug-induced degradation, suggesting that these 3 sequences contain the target of the rapamycin effect. Rapamycin did not promote IL-3 mRNA degradation in cells of a tumor variant lacking expression of FKBP12, the binding protein of rapamycin. Experiments with wortmannin indicated that rapamycin does not act via p70S6 kinase. FK-506, another ligand of FKBP12 affecting the phosphatase calcineurin, did not antagonize but shared the effect of rapamycin. Our data fit a model whereby both FKBP12 and calcineurin target an unknown regulator of IL-3 mRNA turnover.Lymphokines and their related signal transduction pathways play an important role in the control of cell activation, proliferation, and oncogenesis. These processes can be conveniently studied in murine PB-3c mast cells, which are interleukin-3

Tumors obtained from v-Ha-ras-transformed PB-3c cells are characterized by autocrine interleukin-3 (IL3) expression, which occurs either without (class I tumors) or with enhanced transcription (class II tumors). To address possible post-transcriptional mechanisms of IL3 expression, IL3 mRNA stability was examined in both tumor classes. Increased stability of IL3 mRNA was detected in class I tumor lines (t1/2 > 3 h), whereas rapid decay of IL3 transcripts (t1/2 < 0.5 h) was found in class II tumor lines. In both tumor classes, the c-myc and interleukin-6 transcripts were short-lived. Transcripts of a constitutively expressed IL3 reporter gene were stable in class I tumor cells but unstable in class II tumor cells, suggesting that IL3 mRNA stabilization involved a trans-acting mechanism. Rapid decay of IL3 reporter transcripts was observed in untransformed PB-3c as well as in v-Ha-ras expressing precursor cells linking transcript stabilization to the tumor stage. Reporter transcript stabilization in class I tumor cells correlated with increased IL3 production. Deletion of the AU-rich element from the IL3 reporter gene further augmented IL3 mRNA levels as well as IL3 production, suggesting that the stabilizing mechanism in class I tumor cells is not equivalent to AU-rich element deletion.