Negative regulation of Hif1a expression and TH17 differentiation by the hypoxia-regulated microRNA miR-210.

The microRNA miR-210 is a signature of hypoxia. We found robust increase in the abundance of miR-210 (>100-fold) in activated T cells, especially in the TH17 lineage of helper T cells. Hypoxia acted in synergy with stimulation via the T cell antigen receptor (TCR) and coreceptor CD28 to accelerate and increase Mir210 expression. Mir210 was directly regulated by HIF-1α, a key transcriptional regulator of TH17 polarization. Unexpectedly, we identified Hif1a as a target of miR-210, which suggested negative feedback by miR-210 in inhibiting HIF-1α expression. Deletion of Mir210 promoted TH17 differentiation under conditions of limited oxygen. In experimental colitis, miR-210 reduced the abundance of Hif1a transcripts and the proportion of cells that produced inflammatory cytokines and controlled disease severity. Our study identifies miR-210 as an important regulator of T cell differentiation in hypoxia, which can limit immunopathology.

MZB1 is a GRP94 cochaperone that enables proper immunoglobulin heavy chain biosynthesis upon ER stress.

MZB1 (pERp1) is a B-cell-specific and endoplasmic reticulum (ER)-localized protein implicated in antibody secretion and integrin-mediated cell adhesion. Here, we examine the role of MZB1 in vivo by conditional gene inactivation in the mouse germline and at different stages of B lymphopoiesis. Deletion of MZB1 impairs humoral immune responses and antibody secretion in plasma cells that naturally undergo ER stress. In addition, we found that experimental induction of ER stress by tunicamycin injections in mice results in a block of pro-B-cell to pre-B-cell differentiation specifically in Mzb1(-/-) mice. A similar developmental block was observed in Mzb1(fl/fl)mb1(Cre) mice, whereby a Cre recombinase-induced genotoxic stress unmasks a role for MZB1 in the surface expression of immunoglobulin µ heavy chains (µHCs). MZB1 associates directly with the substrate-specific chaperone GRP94 (also called HSP90B1 or gp96) in an ATP-sensitive manner and is required for the interaction of GRP94 with µHCs upon ER stress. Thus, MZB1 seems to act as a substrate-specific cochaperone of GRP94 that enables proper biosynthesis of µHCs under conditions of ER stress.

Mzb1 protein regulates calcium homeostasis, antibody secretion, and integrin activation in innate-like B cells.

Marginal zone (MZ) B cells of the spleen and B1 cells, termed innate-like B cells, differ from follicular B cells by their attenuated Ca(2+) mobilization, fast antibody secretion, and increased cell adhesion. We identified and characterized Mzb1 as an endoplasmic reticulum-localized and B cell-specific protein that was most abundantly expressed in MZ B and B1 cells. Knockdown of Mzb1 in MZ B cells increased Ca(2+) mobilization and nuclear NFAT transcription factor localization, but reduced lipopolysaccharide-induced antibody secretion and integrin-mediated cell adhesion. Conversely, ectopic expression of an Lck-Mzb1 transgene in peripheral T cells resulted in attenuated Ca(2+) mobilization and augmented integrin-mediated cell adhesion. In addition to its interaction with the substrate-specific chaperone Grp94, Mzb1 augmented the function of the oxidoreductase ERp57 in favoring the expression of integrins in their activated conformation. Thus, Mzb1 helps to diversify peripheral B cell functions by regulating Ca(2+) stores, antibody secretion, and integrin activation.

Cell-type-specific function of BCL9 involves a transcriptional activation domain that synergizes with beta-catenin.

Transcriptional regulation by the canonical Wnt pathway involves the stabilization and nuclear accumulation of beta-catenin, which assembles with LEF1/TCF transcription factors and cofactors to activate Wnt target genes. Recently, the nuclear beta-catenin complex has been shown to contain BCL9, which interacts with beta-catenin and recruits Pygopus as a transcriptional coactivator. However, the presumed general functions of Pygopus and BCL9, which has been proposed to act as a scaffolding protein for Pygopus, have been challenged by the rather specific and modest developmental defects of targeted inactivations of both the Pygo1 and the Pygo2 genes. Here, we analyze the function of BCL9 in transcriptional activation by beta-catenin. We find that BCL9 acts in a cell-type-specific manner and, in part, independent of Pygopus. We show that BCL9 itself contains a transcriptional activation domain in the C terminus, which functionally synergizes in lymphoid cells with the C-terminal transactivation domain of beta-catenin. Finally, we identify amino acids in the transactivation domain of beta-catenin that are important for its function and association with the histone acetyltransferases CBP/p300 and TRRAP/GCN5. Thus, BCL9 may serve to modulate and diversify the transcriptional responses to Wnt signaling in a cell-type-specific manner.