Subcellular localization of Mitf in monocytic cells

Microphthalmia-associated transcription factor (Mitf) is a transcription factor that plays an important role in regulating the development of several cell lineages. The subcellular localization of Mitf is dynamic and is associated with its transcription activity. In this study, we examined factors that affect its subcellular localization in cells derived from the monocytic lineage since Mitf is present abundantly in these cells. We identified a domain encoded by Mitf exon 1B1b to be important for Mitf to commute between the cytoplasm and the nucleus. Deletion of this domain disrupts the shuttling of Mitf to the cytoplasm and results in its retention in the nucleus. M-CSF and RANKL both induce nuclear translocation of Mitf. We showed that Mitf nuclear transport is greatly influenced by ratio of M-CSF/Mitf protein expression. In addition, cell attachment to a solid surface also is needed for the nuclear transport of Mitf

Gamma frequency entrainment attenuates amyloid load and modifies microglia

Changes in gamma oscillations (20-50 Hz) have been observed in several neurological disorders. However, the relationship between gamma oscillations and cellular pathologies is unclear. Here we show reduced, behaviourally driven gamma oscillations before the onset of plaque formation or cognitive decline in a mouse model of Alzheimer's disease. Optogenetically driving fast-spiking parvalbumin-positive (FS-PV)-interneurons at gamma (40 Hz), but not other frequencies, reduces levels of amyloid-β (Aβ)[subscript 1-40] and Aβ [subscript 1-42] isoforms. Gene expression profiling revealed induction of genes associated with morphological transformation of microglia, and histological analysis confirmed increased microglia co-localization with Aβ. Subsequently, we designed a non-invasive 40 Hz light-flickering regime that reduced Aβ[subscript 1-40] and Aβ[subscript 1-42] levels in the visual cortex of pre-depositing mice and mitigated plaque load in aged, depositing mice. Our findings uncover a previously unappreciated function of gamma rhythms in recruiting both neuronal and glial responses to attenuate Alzheimer's-disease-associated pathology.National Institutes of Health (U.S.) (Grant 1R01EY023173)National Institutes of Health (U.S.) (Grant 1DP1NS087724)National Institutes of Health (U.S.) (Grant RF1AG047661)National Institutes of Health (U.S.) (Grant ROIGM104948

Towards estimating computer users' mood from interaction behaviour with keyboard and mouse

The purpose of this exploratory research was to study the relationship between the mood of computer users and their use of keyboard and mouse to examine the possibility of creating a generic or individualized mood measure. To examine this, a field study (n = 26) and a controlled study (n = 16) were conducted. In the field study, interaction data and self-reported mood measurements were collected during normal PC use over several days. In the controlled study, participants worked on a programming task while listening to high or low arousing background music. Besides subjective mood measurement, galvanic skin response (GSR) data was also collected. Results found no generic relationship between the interaction data and the mood data. However, the results of the studies found significant average correlations between mood measurement and personalized regression models based on keyboard and mouse interaction data. Together the results suggest that individualized mood prediction is possible from interaction behaviour with keyboard and mouse

The Role of Atypical Protein Kinase C in CSF-1-Dependent Erk Activation and Proliferation in Myeloid Progenitors and Macrophages

Colony stimulating factor-1 (CSF-1 or M-CSF) is the major physiological regulator of the proliferation, differentiation and survival of cells of the mononuclear phagocyte lineage. CSF-1 binds to a receptor tyrosine kinase, the CSF-1 receptor (CSF-1R). Multiple pathways are activated downstream of the CSF-1R; however, it is not clear which pathways regulate proliferation and survival. Here, we investigated the role of atypical protein kinase Cs (PKCζ) in a myeloid progenitor cell line that expressed CSF-1R (32D.R) and in primary murine bone marrow derived macrophages (BMMs). In 32D.R cells, CSF-1 induced the phosphorylation of PKCζ and increased its kinase activity. PKC inhibitors and transfections with mutant PKCs showed that optimal CSF-1-dependent Erk activation and proliferation depended on the activity of PKCζ. We previously reported that CSF-1 activated the Erk pathway through an A-Raf-dependent and an A-Raf independent pathway (Lee and States, Mol. Cell. Biol. 18, 6779). PKC inhibitors did not affect CSF-1 induced Ras and A-Raf activity but markedly reduced MEK and Erk activity, implying that PKCζ regulated the CSF-1-Erk pathway at the level of MEK. PKCζ has been implicated in activating the NF-κB pathway. However, CSF-1 promoted proliferation in an NF-κB independent manner. We established stable 32D.R cell lines that overexpressed PKCζ. Overexpression of PKCζ increased the intensity and duration of CSF-1 induced Erk activity and rendered cells more responsive to CSF-1 mediated proliferation. In contrast to 32D.R cells, PKCζ inhibition in BMMs had only a modest effect on proliferation. Moreover, PKCζ -specific and pan-PKC inhibitors induced a paradoxical increase in MEK-Erk phosphorylation suggesting that PKCs targeted a common negative regulatory step upstream of MEK. Our results demonstrated that CSF-1 dependent Erk activation and proliferation are regulated differentially in progenitors and differentiated cells

Absence of Colony Stimulation Factor-1 Receptor Results in Loss of Microglia, Disrupted Brain Development and Olfactory Deficits

The brain contains numerous mononuclear phagocytes called microglia. These cells express the transmembrane tyrosine kinase receptor for the macrophage growth factor colony stimulating factor-1 (CSF-1R). Using a CSF-1R-GFP reporter mouse strain combined with lineage defining antibody staining we show in the postnatal mouse brain that CSF-1R is expressed only in microglia and not neurons, astrocytes or glial cells. To study CSF-1R function we used mice homozygous for a null mutation in the Csflr gene. In these mice microglia are >99% depleted at embryonic day 16 and day 1 post-partum brain. At three weeks of age this microglial depletion continues in most regions of the brain although some contain clusters of rounded microglia. Despite the loss of microglia, embryonic brain development appears normal but during the post-natal period the brain architecture becomes perturbed with enlarged ventricles and regionally compressed parenchyma, phenotypes most prominent in the olfactory bulb and cortex. In the cortex there is increased neuronal density, elevated numbers of astrocytes but reduced numbers of oligodendrocytes. Csf1r nulls rarely survive to adulthood and therefore to study the role of CSF-1R in olfaction we used the viable null mutants in the Csf1 (Csf1op) gene that encodes one of the two known CSF-1R ligands. Food-finding experiments indicate that olfactory capacity is significantly impaired in the absence of CSF-1. CSF-1R is therefore required for the development of microglia, for a fully functional olfactory system and the maintenance of normal brain structure

Deletion of a Csf1r enhancer selectively impacts CSF1R expression and development of tissue macrophage populations.

The proliferation, differentiation and survival of mononuclear phagocytes depend on signals from the receptor for macrophage colony-stimulating factor, CSF1R. The mammalian Csf1r locus contains a highly conserved super-enhancer, the fms-intronic regulatory element (FIRE). Here we show that genomic deletion of FIRE in mice selectively impacts CSF1R expression and tissue macrophage development in specific tissues. Deletion of FIRE ablates macrophage development from murine embryonic stem cells. Csf1r mice lack macrophages in the embryo, brain microglia and resident macrophages in the skin, kidney, heart and peritoneum. The homeostasis of other macrophage populations and monocytes is unaffected, but monocytes and their progenitors in bone marrow lack surface CSF1R. Finally, Csf1r mice are healthy and fertile without the growth, neurological or developmental abnormalities reported in Csf1r rodents. Csf1r mice thus provide a model to explore the homeostatic, physiological and immunological functions of tissue-specific macrophage populations in adult animals

The evolution of the macrophage-specific enhancer (Fms intronic regulatory element) within the CSF1R locus of vertebrates

The Csf1r locus encodes the receptor for macrophage colony-stimulating factor, which controls the proliferation, differentiation and survival of macrophages. The 300 bp Fms intronic regulatory element (FIRE), within the second intron of Csf1r, is necessary and sufficient to direct macrophage-specific transcription. We have analysed the conservation and divergence of the FIRE DNA sequence in vertebrates. FIRE is present in the same location in the Csf1r locus in reptile, avian and mammalian genomes. Nearest neighbor analysis based upon this element alone largely recapitulates phylogenies inferred from much larger genomic sequence datasets. One core element, containing binding sites for AP1 family and the macrophage-specific transcription factor, PU.1, is conserved from lizards to humans. Around this element, the FIRE sequence is conserved within clades with the most conserved elements containing motifs for known myeloid-expressed transcription factors. Conversely, there is little alignment between clades outside the AP1/PU.1 element. The analysis favours a hybrid between "enhanceosome" and "smorgasbord" models of enhancer function, in which elements cooperate to bind components of the available transcription factor milieu

Analysis of the human monocyte-derived macrophage transcriptome and response to lipopolysaccharide provides new insights into genetic aetiology of inflammatory bowel disease

The FANTOM5 consortium utilised cap analysis of gene expression (CAGE) to provide an unprecedented insight into transcriptional regulation in human cells and tissues. In the current study, we have used CAGE-based transcriptional profiling on an extended dense time course of the response of human monocyte-derived macrophages grown in macrophage colony-stimulating factor (CSF1) to bacterial lipopolysaccharide (LPS). We propose that this system provides a model for the differentiation and adaptation of monocytes entering the intestinal lamina propria. The response to LPS is shown to be a cascade of successive waves of transient gene expression extending over at least 48 hours, with hundreds of positive and negative regulatory loops. Promoter analysis using motif activity response analysis (MARA) identified some of the transcription factors likely to be responsible for the temporal profile of transcriptional activation. Each LPS-inducible locus was associated with multiple inducible enhancers, and in each case, transient eRNA transcription at multiple sites detected by CAGE preceded the appearance of promoter-associated transcripts. LPS-inducible long non-coding RNAs were commonly associated with clusters of inducible enhancers. We used these data to re-examine the hundreds of loci associated with susceptibility to inflammatory bowel disease (IBD) in genome-wide association studies. Loci associated with IBD were strongly and specifically (relative to rheumatoid arthritis and unrelated traits) enriched for promoters that were regulated in monocyte differentiation or activation. Amongst previously-identified IBD susceptibility loci, the vast majority contained at least one promoter that was regulated in CSF1-dependent monocyte-macrophage transitions and/or in response to LPS. On this basis, we concluded that IBD loci are strongly-enriched for monocyte-specific genes, and identified at least 134 additional candidate genes associated with IBD susceptibility from reanalysis of published GWA studies. We propose that dysregulation of monocyte adaptation to the environment of the gastrointestinal mucosa is the key process leading to inflammatory bowel disease

Trophic macrophages in development and disease

Specialized phagocytes are found in the most primitive multicellular organisms. Their roles in homeostasis and in distinguishing self from non-self have evolved with the complexity of organisms and their immune systems. Equally important, but often overlooked, are the roles of macrophages in tissue development. As discussed in this Review, these include functions in branching morphogenesis, neuronal patterning, angiogenesis, bone morphogenesis and the generation of adipose tissue. In each case, macrophage depletion impairs the formation of the tissue and compromises its function. I argue that in several diseases, the unrestrained acquisition of these developmental macrophage functions exacerbates pathology. For example, macrophages enhance tumour progression and metastasis by affecting tumour-cell migration and invasion, as well as angiogenesis