Transcription factor access to regulatory elements is prevented by the nucleosome. Heat shock factor 1 (HSF1) is a winged helix transcription factor that plays roles in control and stressed conditions by gaining access to target elements, but mechanisms of HSF1 access are not well known in mammalian cells. Here, we show the physical interaction between the wing motif of human HSF1 and replication protein A (RPA), which is involved in DNA metabolism. Depletion of RPA1 abolishes HSF1 access to the promoter of HSP70 in unstressed condition and delays its rapid activation in response to heat shock. The HSF1-RPA complex leads to preloading of RNA polymerase II and opens the chromatin structure by recruiting a histone chaperone, FACT. Furthermore, this interaction is required for melanoma cell proliferation. These results provide a mechanism of constitutive HSF1 access to nucleosomal DNA, which is important for both basal and inducible gene expression.

The febrile response is a complex physiological reaction to disease, including a cytokine-mediated increase in body temperature and the activation of inflammatory systems. Fever has beneficial roles in terms of disease prognosis, partly by suppressing the expression of inflammatory cytokines. However, the molecular mechanisms underlining the fever-mediated suppression of inflammatory gene expression have not been clarified. In this study, we showed that heat shock suppresses LPS-induced expression of IL-6, a major pyrogenic cytokine, in mouse embryonic fibroblasts and macrophages. Heat shock transcription factor 1 (HSF1) activated by heat shock induced the expression of activating transcription factor (ATF) 3, a negative regulator of IL-6, and ATF3 was necessary for heat-mediated suppression of IL-6, indicating a fever-mediated feedback loop consisting of HSF1 and ATF3. A comprehensive analysis of inflammatory gene expression revealed that heat pretreatment suppresses LPS-induced expression of most genes (86%), in part (67%) via ATF3. When HSF1-null and ATF3-null mice were injected with LPS, they expressed much higher levels of IL-6 than wild-type mice, resulting in an exaggerated febrile response. These results demonstrate a novel inhibitory pathway for inflammatory cytokines.

Heat shock transcription factor 1 (HSF1) is an important regulator of protein homeostasis (proteostasis) by controlling the expression of major heat shock proteins (Hsps) that facilitate protein folding. However, it is unclear whether other proteostasis pathways are mediated by HSF1. Here, we identified novel targets of HSF1 in mammalian cells, which suppress the aggregation of polyglutamine (polyQ) protein. Among them, we show that one of the nuclear factor of activated T cells (NFAT) proteins, NFATc2, significantly inhibits polyQ aggregation in cells and is required for HSF1-mediated suppression of polyQ aggregation. NFAT deficiency accelerated disease progression including aggregation of a mutant polyQ-huntingtin protein and shortening of lifespan in R6/2 Huntington's disease mice. Furthermore, we found that HSF1 and NFAT cooperatively induce the expression of the scaffold protein PDZK3 and aB-crystallin, which facilitate the degradation of polyQ protein. These results show the first mechanistic basis for the observation that HSF1 has a much more profound effect on proteostasis than individual Hsp or combination of different Hsps, and suggest a new pathway for ameliorating protein-misfolding diseases.

We have tried to obtain new insight into the development of the medulla oblongata by using the quail-to-chick chimera system. Five types of isotopic and isochronic grafts were carried out, between quail and chick embryos, at the 10- to 12-somite stage: exchanges of (I) the entire myelencephalon, (II) the dorsal half of the myelencephalon, (III) the ventral half of the myelencephalon, (IV) the right half of the myelencephalon and (V) the dorsal quarter of the myelencephalon. Before analyzing the chimeric embryos, we studied the ontogeny of the various nuclei in the medulla oblongata of normal birds. The first appearance of nuclei in quail embryos preceded in many cases that of their chick counterpart by 12 to 24 h. The adult pattern of the nuclei was established by E8 in quail and E9 in chick. Similarly, during early development of chimeras, the migration of quail cells began earlier than that of chick cells. This shows that the species specific temporal sequence of proliferation and migration is not significantly altered by transplantation into the host. The possibility of grafting selectively the ventral or dorsal half of the neural tube allowed us to distinguish the fate of the cells belonging respectively to the alar and the basal plate. The nuclei with a total or partial motor function, such as the nucleus nervi abducentis, the nucleus nervi facialis, the nucleus nervi glossopharyngei and the nucleus motorius dorsalis nervi vagi, have either an exclusive or predominant origin from the basal plate. In contrast, the nuclei with essentially or exclusively sensory components (i.e., nucleus angularis, nucleus laminaris, nucleus magnocellularis) arise from the alar plate. The reticular formation such as the nucleus reticularis gigantocellularis and the nucleus reticularis subtrigeminalis was strikingly mixed, with both alar and basal plate origin of neurons. Active dorsoventral migrations of cells originating migrations from the dorsal neural tube, the "rhombic lip", contribute the ventral nuclei (i.e., nuclei pontis medialis, lateralis and olivaris inferior), whose functions are essentially associative. This study shows different types of cell migration. Dorsoventral and ventrodorsal movements are essentially active from E5 to E8. In the medulla oblongata, the dorsoventral stream is highly predominant. From E8 to E9, cells belonging to the marginal stream cross the midline laterally in both directions. Beyond E12, longitudinal migrations occur ventrally in both rostrocaudal and caudorostral directions. The immunohistochemical analyses carried out on chimeras generated in experiment V revealed the existence of fibers in marginal zones prior to the onset of the migration of cell bodies.(ABSTRACT TRUNCATED AT 400 WORDS)

HSF1 is a master regulator of the heat-shock response in mammalian cells, whereas in avian cells, HSF3, which was considered as an avian-specific factor, is required for the expression of classical heat-shock genes. Here, the authors identify mouse HSF3, and demonstrate that it has the potential to activate only nonclassical heat-shock genes.

bThe heat shock response is an evolutionally conserved adaptive response to high temperatures that controls proteostasis capacity and is regulated mainly by an ancient heat shock factor (HSF). However, the regulation of target genes by the stress-inducible HSF1 transcription complex has not yet been examined in detail in mammalian cells. In the present study, we demonstrated that HSF1 interacted with members of the ATF1/CREB family involved in metabolic homeostasis and recruited them on the HSP70 promoter in response to heat shock. The HSF1 transcription complex, including the chromatin-remodeling factor BRG1 and lysine acetyltransferases p300 and CREB-binding protein (CBP), was formed in a manner that was dependent on the phosphorylation of ATF1. ATF1-BRG1 promoted the establishment of an active chromatin state and HSP70 expression during heat shock, whereas ATF1-p300/CBP accelerated the shutdown of HSF1 DNA-binding activity during recovery from acute stress, possibly through the acetylation of HSF1. Furthermore, ATF1 markedly affected the resistance to heat shock. These results revealed the unanticipated complexity of the primitive heat shock response mechanism, which is connected to metabolic adaptation. All living cells maintain a balance among the synthesis, folding, and clearance of individual proteins in order to maintain the proper conformations and physiological concentrations of proteins, and this is referred to as protein homeostasis or proteostasis (1). To survive temperature elevations, which cause protein unfolding and misfolding, cells induce the expression of a small number of highly conserved heat shock proteins (HSPs or chaperones) and hundreds of non-HSP proteins involved in diverse functions, including protein degradation (2, 3). Thus, this universal adaptive response, which is known as the heat shock response, controls the proteostasis capacity or buffering capacity against protein misfolding in a cell (4) and is regulated mainly at the level of transcription by the ancient transcription factor 32 in Escherichia coli (5) or heat shock factor (HSF) in eukaryotes (6, 7).In contrast to the E. coli genome, which is compressed into a small space through supercoiling (8), eukaryotic genomes are packaged into nucleosomes, which are composed of DNA wrapped around the histone octamer and occlude DNA from interacting with most DNA-binding proteins (9). To induce transcription during heat shock, HSF binds to regulatory elements and recruits coactivators, including chromatin-modifying enzymes and nucleosome-remodeling complexes that move or displace histones at the promoter and gene body (10). Metazoan HSF remains mostly as an inactive monomer in unstressed cells and is converted to an active trimer that binds to the heat shock response element (HSE) during heat shock (11). In Drosophila, the GAGA factor restricts the nucleosome occupancy of the HSP70 promoter, thereby allowing the establishment of paused RNA polymerase II (Pol II) in unstressed cells (12). In response to heat shock, the increased level...

Heat-shock response is an adaptive response to proteotoxic stresses including heat shock, and is regulated by heat-shock factor 1 (HSF1) in mammals. Proteotoxic stresses challenge all subcellular compartments including the mitochondria. Therefore, there must be close connections between mitochondrial signals and the activity of HSF1. Here, we show that heat shock triggers nuclear translocation of mitochondrial SSBP1, which is involved in replication of mitochondrial DNA, in a manner dependent on the mitochondrial permeability transition pore ANT–VDAC1 complex and direct interaction with HSF1. HSF1 recruits SSBP1 to the promoters of genes encoding cytoplasmic/nuclear and mitochondrial chaperones. HSF1–SSBP1 complex then enhances their induction by facilitating the recruitment of a chromatin-remodelling factor BRG1, and supports cell survival and the maintenance of mitochondrial membrane potential against proteotoxic stresses. These results suggest that the nuclear translocation of mitochondrial SSBP1 is required for the regulation of cytoplasmic/nuclear and mitochondrial proteostasis against proteotoxic stresses.

Most cancer immunotherapies under present investigation are based on the belief that cytotoxic T cells are the most important anti-tumoral immune cells, whereas intra-tumoral macrophages would rather play a pro-tumoral role. We have challenged this antagonistic point of view and searched for collaborative contributions by tumor-infiltrating T cells and macrophages, reminiscent of those observed in anti-infectious responses. We demonstrate that, in a model of therapeutic vaccination, cooperation between myeloid cells and T cells is indeed required for tumor rejection. Vaccination elicited an early rise of CD11b+ myeloid cells that preceded and conditioned the intra-tumoral accumulation of CD8+ T cells. Conversely, CD8+ T cells and IFNγ production activated myeloid cells were required for tumor regression. A 4-fold reduction of CD8+ T cell infiltrate in CXCR3KO mice did not prevent tumor regression, whereas a reduction of tumor-infiltrating myeloid cells significantly interfered with vaccine efficiency. We show that macrophages from regressing tumors can kill tumor cells in two ways: phagocytosis and TNFα release. Altogether, our data suggest new strategies to improve the efficiency of cancer immunotherapies, by promoting intra-tumoral cooperation between macrophages and T cells.