Regulation of Immature Dendritic Cell Migration by RhoA Guanine Nucleotide Exchange Factor Arhgef5
Program for Vascular Biology and Therapeutics and Department of Pharmacology, Yale University School of Medicine, New Haven, Connecticut 06520,, USA. Journal of Biological Chemistry
(Impact Factor: 4.57).
09/2009; 284(42):28599-606. DOI: 10.1074/jbc.M109.047282
There are a large number of Rho guanine nucleotide exchange factors, most of which have no known functions. Here, we carried out a short hairpin RNA-based functional screen of Rho-GEFs for their roles in leukocyte chemotaxis and identified Arhgef5 as an important factor in chemotaxis of a macrophage phage-like RAW264.7 cell line. Arhgef5 can strongly activate RhoA and RhoB and weakly RhoC and RhoG, but not Rac1, RhoQ, RhoD, or RhoV, in transfected human embryonic kidney 293 cells. In addition, Gbetagamma interacts with Arhgef5 and can stimulate Arhgef5-mediated activation of RhoA in an in vitro assay. In vivo roles of Arhgef5 were investigated using an Arhgef-5-null mouse line. Arhgef5 deficiency did not affect chemotaxis of mouse macrophages, T and B lymphocytes, and bone marrow-derived mature dendritic cells (DC), but it abrogated MIP1alpha-induced chemotaxis of immature DCs and impaired migration of DCs from the skin to lymph node. In addition, Arhgef5 deficiency attenuated allergic airway inflammation. Therefore, this study provides new insights into signaling mechanisms for DC migration regulation.
Available from: Petr Draber
- "Although active RhoA transiently decreased after FcεRI triggering, more in NTAL KO cells than in WT cells, it is likely that differences in regulation of RhoA activity in NTAL-deficient cells and WT cells are responsible for the enhanced NTAL-regulated chemotaxis. It should be stressed that previous reports have shown that RhoA regulates chemotaxis in other cell types, such as neutrophils –, macrophages , dendritic cells  and lymphocytes . "
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ABSTRACT: Non-T cell activation linker (NTAL; also called LAB or LAT2) is a transmembrane adaptor protein that is expressed in a subset of hematopoietic cells, including mast cells. There are conflicting reports on the role of NTAL in the high affinity immunoglobulin E receptor (FcεRI) signaling. Studies carried out on mast cells derived from mice with NTAL knock out (KO) and wild type mice suggested that NTAL is a negative regulator of FcεRI signaling, while experiments with RNAi-mediated NTAL knockdown (KD) in human mast cells and rat basophilic leukemia cells suggested its positive regulatory role. To determine whether different methodologies of NTAL ablation (KO vs KD) have different physiological consequences, we compared under well defined conditions FcεRI-mediated signaling events in mouse bone marrow-derived mast cells (BMMCs) with NTAL KO or KD. BMMCs with both NTAL KO and KD exhibited enhanced degranulation, calcium mobilization, chemotaxis, tyrosine phosphorylation of LAT and ERK, and depolymerization of filamentous actin. These data provide clear evidence that NTAL is a negative regulator of FcεRI activation events in murine BMMCs, independently of possible compensatory developmental alterations. To gain further insight into the role of NTAL in mast cells, we examined the transcriptome profiles of resting and antigen-activated NTAL KO, NTAL KD, and corresponding control BMMCs. Through this analysis we identified several genes that were differentially regulated in nonactivated and antigen-activated NTAL-deficient cells, when compared to the corresponding control cells. Some of the genes seem to be involved in regulation of cholesterol-dependent events in antigen-mediated chemotaxis. The combined data indicate multiple regulatory roles of NTAL in gene expression and mast cell physiology.
PLoS ONE 08/2014; 9(8):e105539. DOI:10.1371/journal.pone.0105539 · 3.23 Impact Factor
Available from: Lester Kobzik
- "Among these 57 genes which evidently interact with each other very well, there are 29 that we found have a known role in asthma or allergic inflammation. They are: Klf6 (Gene ID: 23849) , Lamin A/C (Gene ID: 16905) , E4BP4 (Gene ID: 18030) , Eomesodemin (Gene ID: 13813) , GATA-3 (Gene ID: 14462) , T-bet (Gene ID: 57765) , CXCR5 (Gene ID: 12145) , SOCS3(Gene ID: 12702) , Lck (Gene ID: 16818) and FGR(Gene ID: 14191) , MKP-1 (Gene ID: 19252) , ALOX12(Gene ID: 11684) , RhoB (Gene ID 11852) , G0/G1 switch 2(Gene ID: 14373) relevant to cell proliferation, NOXA (Gene ID: 58801) , Dematin (Gene ID: 13829) , PKC-alpha (Gene ID: 18750) , TRPC6 (Gene ID: 22068) , VEGFR-1(Gene ID: 14254) , CXCR4 (Gene ID: 12767) , Beta-2 adrenergic receptor(Gene ID: 11555) , Il12a (Gene ID: 16159) , NPY (Gene ID: 109648) , Slit2 (Gene ID: 20563) , Col1a2 (Gene ID: 12843) , Cxcl12 (Gene ID: 20315) , MMP-9 (Gene ID: 17395) , IBP3 (Gene ID: 16009) , Kallikrein 4 (Gene ID: 56640) . The rest of the factors (genes) are less well characterized and comprise opportunities for future research. "
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ABSTRACT: We investigated the link between epigenome-wide methylation aberrations at birth and genomic transcriptional changes upon allergen sensitization that occur in the neonatal dendritic cells (DC) due to maternal asthma. We previously demonstrated that neonates of asthmatic mothers are born with a functional skew in splenic DCs that can be seen even in allergen-naïve pups and can convey allergy responses to normal recipients. However, minimal-to-no transcriptional or phenotypic changes were found to explain this alteration. Here we provide in-depth analysis of genome-wide DNA methylation profiles and RNA transcriptional (microarray) profiles before and after allergen sensitization. We identified differentially methylated and differentially expressed loci and performed manually-curated matching of methylation status of the key regulatory sequences (promoters and CpG islands) to expression of their respective transcripts before and after sensitization. We found that while allergen-naive DCs from asthma-at-risk neonates have minimal transcriptional change compared to controls, the methylation changes are extensive. The substantial transcriptional change only becomes evident upon allergen sensitization, when it occurs in multiple genes with the pre-existing epigenetic alterations. We demonstrate that maternal asthma leads to both hyper- and hypomethylation in neonatal DCs, and that both types of events at various loci significantly overlap with transcriptional responses to allergen. Pathway analysis indicates that approximately 1/2 of differentially expressed and differentially methylated genes directly interact in known networks involved in allergy and asthma processes. We conclude that congenital epigenetic changes in DCs are strongly linked to altered transcriptional responses to allergen and to early-life asthma origin. The findings are consistent with the emerging paradigm that asthma is a disease with underlying epigenetic changes.
PLoS ONE 08/2013; 8(8):e70387. DOI:10.1371/journal.pone.0070387 · 3.23 Impact Factor
Available from: Mohamed Aittaleb
- "Pulldown assays using purified proteins demonstrated that the interaction between Tim and G␤␥ was direct. G␤␥ also activated Tim in a serum response element reporter assay as well as in vitro using purified proteins (Wang et al., 2009). It remains to be seen how regulation by G␤␥ is integrated into the layers of autoinhibition already mediated by the SH3 domain and N-terminal helix motif, or if it simply provides a mechanism to bypass them. "
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ABSTRACT: Activation of certain classes of G protein-coupled receptors (GPCRs) can lead to alterations in the actin cytoskeleton, gene transcription, cell transformation, and other processes that are known to be regulated by Rho family small-molecular-weight GTPases. Although these responses can occur indirectly via cross-talk from canonical heterotrimeric G protein cascades, it has recently been demonstrated that Dbl family Rho guanine nucleotide exchange factors (RhoGEFs) can serve as the direct downstream effectors of heterotrimeric G proteins. Heterotrimeric Galpha(12/13), Galpha(q), and Gbetagamma subunits are each now known to directly bind and regulate RhoGEFs. Atomic structures have recently been determined for several of these RhoGEFs and their G protein complexes, providing fresh insight into the molecular mechanisms of signal transduction between GPCRs and small molecular weight G proteins. This review covers what is currently known about the structure, function, and regulation of these recently recognized effectors of heterotrimeric G proteins.
Molecular pharmacology 10/2009; 77(2):111-25. DOI:10.1124/mol.109.061234 · 4.13 Impact Factor
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