Cryptochromes (CRY) are photolyase-like blue-light receptors that mediate light responses in plants and animals. How plant cryptochromes act in response to blue light is not well understood. We report here the identification and characterization of the Arabidopsis CIB1 (cryptochrome-interacting basic-helix-loop-helix) protein. CIB1 interacts with CRY2 (cryptochrome 2) in a blue light-specific manner in yeast and Arabidopsis cells, and it acts together with additional CIB1-related proteins to promote CRY2-dependent floral initiation. CIB1 binds to G box (CACGTG) in vitro with a higher affinity than its interaction with other E-box elements (CANNTG). However, CIB1 stimulates FT messenger RNA expression, and it interacts with chromatin DNA of the FT gene that possesses various E-box elements except G box. We propose that the blue light-dependent interaction of cryptochrome(s) with CIB1 and CIB1-related proteins represents an early photoreceptor signaling mechanism in plants.

SREBPs are key transcriptional regulators of lipid metabolism and cellular growth. It has been proposed that SREBP signaling regulates cellular growth through its ability to drive lipid biosynthesis. Unexpectedly, we find that loss of SREBP activity inhibits cancer cell growth and viability by uncoupling fatty acid synthesis from desaturation. Integrated lipid profiling and metabolic flux analysis revealed that cancer cells with attenuated SREBP activity maintain long-chain saturated fatty acid synthesis, while losing fatty acid desaturation capacity. We traced this defect to the uncoupling of Fatty Acid Synthase activity from SCD1-mediated desaturation. This deficiency in desaturation drives an imbalance between the saturated and monounsaturated fatty acid pools resulting in severe lipotoxicity. Importantly, replenishing the monounsaturated fatty acid pool restored growth to SREBP-inhibited cells. These studies highlight the importance of fatty acid desaturation in cancer growth and provide a novel mechanistic explanation for the role of SREBPs in cancer metabolism.

Summary Dendritic cell (DC) vaccination is emerging as a promising therapeutic option for malignant glioma patients. However, the optimal antigen formulation for loading these cells has yet to be established. The objective of this study was to compare the safety, feasibility, and immune responses of malignant glioma patients on two different DC vaccination protocols. 28 patients were treated with autologous tumor lysate (ATL)-pulsed DC vaccination, while 6 patients were treated with glioma-associated antigen (GAA) peptide-pulsed DCs. Safety, toxicity, feasibility and correlative immune monitoring assay results were compared between patients on each trial. Due to HLA subtype restrictions on the GAA-DC trial, 6/15 screened patients were eligible for treatment, while 28/32 patients passed eligibility screening for the ATL-DC trial. Elevated frequencies of activated natural killer (NK) cells were observed in the peripheral blood from GAA-DC patients compared with the ATL-DC patients. In addition, a significant correlation was observed between decreased regulatory T lymphocyte (Treg) ratios (post/pre vaccination) and overall survival (OS; p=0.004) in patients on both trials. In fact, Treg ratios were independently prognostic for OS in these patients, while tumor pathology was not in multivariate analyses. In conclusion, these results suggest that ATL-DC vaccination is associated with wider patient eligibility compared with GAA-DC vaccination. Decreased post/pre-vaccination Treg ratios and decreased frequencies of activated NK cells were associated with prolonged survival in patients from both trials, suggesting that these lymphocyte subsets may be relevant immune monitoring endpoints for immunotherapy protocols in malignant glioma patients.

PurposeDendritic cell (DC) vaccines have recently emerged as an innovative therapeutic option for glioblastoma patients. To identify novel surrogates of anti-tumor immune responsiveness, we studied the dynamic expression of activation and inhibitory markers on peripheral blood lymphocyte (PBL) subsets in glioblastoma patients treated with DC vaccination at UCLA.Experimental DesignPre-treatment and post-treatment PBL from 24 patients enrolled in two Phase I clinical trials of dendritic cell immunotherapy were stained and analyzed using flow cytometry. A univariate Cox proportional hazards model was utilized to investigate the association between continuous immune monitoring variables and survival. Finally, the immune monitoring variables were dichotomized and a recursive partitioning survival tree was built to obtain cut-off values predictive of survival.ResultsThe change in regulatory T cell (CD3+CD4+CD25+CD127low) frequency in PBL was significantly associated with survival (p = 0.0228; hazard ratio = 3.623) after DC vaccination. Furthermore, the dynamic expression of the negative co-stimulatory molecule, CTLA-4, was also significantly associated with survival on CD3+CD4+ T cells (p = 0.0191; hazard ratio = 2.840) and CD3+CD8+ T cells (p = 0.0273; hazard ratio = 2.690), while that of activation markers (CD25, CD69) was not. Finally, a recursive partitioning tree algorithm was utilized to dichotomize the post/pre fold change immune monitoring variables. The resultant cut-off values from these immune monitoring variables could effectively segregate these patients into groups with significantly different overall survival curves.ConclusionsOur results suggest that monitoring the change in regulatory T cell frequencies and dynamic expression of the negative co-stimulatory molecules on peripheral blood T cells, before and after DC vaccination, may predict survival. The cut-off point generated from these data can be utilized in future prospective immunotherapy trials to further evaluate its predictive validity.

The intestinal epithelium is a paradigm of adult tissue in constant regeneration that is supported by intestinal stem cells (ISCs). The mechanisms regulating ISC homeostasis after injury are poorly understood. We previously demonstrated that IκBα, the main regulator of NF‐κB, exerts alternative nuclear functions as cytokine sensor in a subset of PRC2‐regulated genes. Here, we show that nuclear IκBα is present in the ISC compartment. Mice deficient for IκBα show altered intestinal cell differentiation with persistence of a fetal‐like ISC phenotype, associated with aberrant PRC2 activity at specific loci. Moreover, IκBα‐deficient intestinal cells produce morphologically aberrant organoids carrying a PRC2‐dependent fetal‐like transcriptional signature. DSS treatment, which induces acute damage in the colonic epithelium of mice, results in a temporary loss of nuclear P‐IκBα and its subsequent accumulation in early CD44‐positive regenerating areas. Importantly, IκBα‐deficient mice show higher resistance to damage, likely due to the persistent fetal‐like ISC phenotype. These results highlight intestinal IκBα as a chromatin sensor of inflammation in the ISC compartment.