The Epstein-Barr virus (EBV) is endemic in humans and can efficiently transform infected B cells under some circumstances. If an EBV carrier experiences immune suppression, EBV
B cells can turn into ...lymphoblasts and exhibit growth expansion that may cause lymphoproliferative diseases which often develop into lymphoma. Our immune system conducts surveillance for EBV
B cells in order to block spontaneous tumor formation. Here, we summarize the EBV products involved in tumorigenesis, EBV-associated lymphomas, and pathologically relevant mouse models. Preclinical mouse models for a range of EBV-associated diseases not only clear the path to new therapeutic approaches but also aid in our understanding of the nature of lymphomagenesis and immune surveillance.
Phosphatidylinositol 3′ OH kinase (PI3K) signaling and FOXO transcription factors play opposing roles at several B cell developmental stages. We show here abundant nuclear FOXO1 expression in the ...proliferative compartment of the germinal center (GC), its dark zone (DZ), and PI3K activity, downregulating FOXO1, in the light zone (LZ), where cells are selected for further differentiation. In the LZ, however, FOXO1 was expressed in a fraction of cells destined for DZ reentry. Upon FOXO1 ablation or induction of PI3K activity, GCs lost their DZ, owing at least partly to downregulation of the chemokine receptor CXCR4. Although this prevented proper cyclic selection of cells in GCs, somatic hypermutation and proliferation were maintained. Class switch recombination was partly lost due to a failure of switch region targeting by activation-induced deaminase (AID).
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•PI3K signaling and FOXO1 are antagonistic master regulators of the GC reaction•FOXO1 is essential for the proliferating GC compartment, the dark zone•Selection, but not generation, of somatic antibody mutants is controlled by FOXO1•FOXO1 controls isotype switching in GCs through switch region accessibility for AID
The germinal center (GC) reaction is of key importance for adaptive immunity as well as the origin of most human B cell lymphomas. Using conditional targeted mutagenesis, Sander, Rajewsky, and colleagues identify PI3K signaling and the transcription factor FOXO1 as essential antagonistic regulators of GC polarization, antibody diversification, and cellular selection.
Emerging evidence indicates that heterogeneity in ribosome composition can give rise to specialized functions. Until now, research mainly focused on differences in core ribosomal proteins and ...associated factors. The effect of posttranslational modifications has not been studied systematically. Analyzing ribosome heterogeneity is challenging because individual proteins can be part of different subcomplexes (40S, 60S, 80S, and polysomes). Here we develop polysome proteome profiling to obtain unbiased proteomic maps across ribosomal subcomplexes. Our method combines extensive fractionation by sucrose gradient centrifugation with quantitative mass spectrometry. The high resolution of the profiles allows us to assign proteins to specific subcomplexes. Phosphoproteomics on the fractions reveals that phosphorylation of serine 38 in RPL12/uL11, a known mitotic CDK1 substrate, is strongly depleted in polysomes. Follow-up experiments confirm that RPL12/uL11 phosphorylation regulates the translation of specific subsets of mRNAs during mitosis. Together, our results show that posttranslational modification of ribosomal proteins can regulate translation.
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•High-resolution polysome proteome profiling across ribosomal subcomplexes•Core ribosome of monosomes and polysomes does not differ significantly•Systematic identification of proteins associated with actively translating ribosomes•Phosphorylation of RPL12/uL11 regulates translation during mitosis
Multiple regulatory layers shape gene expression. Imami et al. show that protein production in mitosis can be regulated by phosphorylation of the ribosomal protein RPL12/uL11.
The Cre/loxP system is a powerful tool for generating conditional gene knockout (KO) mice and elucidate gene function in vivo. CD19-Cre and Mb1-iCre transgenic mice are commonly used for generating B ...cell-specific KO mice and investigate the development, as well as the physiological and pathophysiological roles of B cells. However, the CD19-Cre line low efficiency and the Mb1-iCre line occasional ectopic recombination represent challenges for their use. Thus, we developed a CD19-codon-improved Cre (CD19-iCre) knock-in mouse with the T2A-iCre sequence inserted into the Cd19 locus, just before the stop codon. The CD19-iCre mice were compared with existing models, crossed with the Rosa26-EYFP reporter mice, and their recombination activity in B cells carrying different Cre alleles was assessed. CD19-iCre mice showed more effective Cre recombination in the early B cell developmental stages compared with the CD19-Cre mice. The efficiencies of the CD19-iCre and Mb1-iCre lines were similar; however, the B lineage-specific recombination was more stringent in the CD19-iCre line. Furthermore, the utility value of the CD19-iCre model was superior than that of the CD19-Cre mice regarding deletion efficiency in IL10-floxed mice. Thus, the CD19-iCre line is a valuable tool for highly efficient gene targeting specific to the B cell compartment.
Although several long noncoding RNAs (lncRNAs) have recently been shown to encode small polypeptides, those in testis remain largely uncharacterized. Here we identify two sperm-specific polypeptides, ...Kastor and Polluks, encoded by a single mouse locus (Gm9999) previously annotated as encoding a lncRNA. Both Kastor and Polluks are inserted in the outer mitochondrial membrane and directly interact with voltage-dependent anion channel (VDAC), despite their different amino acid sequences. Male VDAC3-deficient mice are infertile as a result of reduced sperm motility due to an abnormal mitochondrial sheath in spermatozoa, and deficiency of both Kastor and Polluks also severely impaired male fertility in association with formation of a similarly abnormal mitochondrial sheath. Spermatozoa lacking either Kastor or Polluks partially recapitulate the phenotype of those lacking both. Cooperative function of Kastor and Polluks in regulation of VDAC3 may thus be essential for mitochondrial sheath formation in spermatozoa and for male fertility.
Senescent cells were recently shown to play a role in aging-related malfunctions and pathologies. This consensus has been facilitated by evidence from senolytic model mice capable of eliminating ...senescent cells in tissues using well-characterized senescent markers, such as p16INK4a (hereafter p16). However, since the incomplete or artificial gene expression regulatory regions of manipulated marker genes affect their cognate expression, it currently remains unclear whether these models accurately reflect physiological senescence. We herein describe a novel approach to eliminate p16-expressing cells from mice at any given point in time, generating a new type of knock-in model, p16hCD2 mice and a toxin-conjugated anti-human CD2 antibody (hCD2-SAP) as an inducer. p16hCD2 mice possess an intact Cdkn2a locus that includes a p16 coding region and human CD2 (hCD2) expression unit. We confirmed cognate p16-associated hCD2 expression in mouse embryonic fibroblasts (MEFs) and in several tissues, such as the spleen, liver, and skin. We detected chronological increases in the hCD2-positive population in T lymphocytes that occurred in a p16-dependent manner, which reflected physiological aging. We then confirmed the high sensitivity of hCD2-SAP to hCD2 and validated its efficacy to remove p16-positive cells, particularly in T lymphocytes. The multiple administration of hCD2-SAP for a prolonged p16-positive cell deficiency partially restored aging-related phenotypes in T lymphocytes, such as the contraction of the CD4+ naïve population and expansion of senescence-associated T cells. Our novel approach of targeting p16-positive senescent cells will provide novel insights into the mechanisms underlying physiological aging in vivo.
•A novel knock-in model p16hCD2 mouse expressed the surface marker hCD2, which correlated with the induction of p16INK4a.•A hCD2-targeting immunotoxin eliminated hCD2-positive MEFs derived from p16hCD2 mice.•Age-associated p16INK4a-positive T cells in p16hCD2 mice were removed by hCD2-SAP in a hCD2 expression-dependent manner.•The administration of hCD2-SAP to p16hCD2 mice partially recovered the age-related phenotypes of CD4+ T cells.
The CARMA1–Bcl10–MALT1 (CBM) signalosome is a crucial module of NF‐κB activation in B cell receptor (BCR) signaling. Biophysical studies have shown that the E3 ubiquitin ligase TRAF6 cooperatively ...modifies the CBM signalosome; however, the specific details regarding how TRAF6 is involved in BCR signal‐induced CBM formation remain unclear. In this study, we aimed to reveal the influences of TRAF6 on CBM formation and TAK1 and IKK activities using DT40 B cells which lack all the exons of TRAF6. In TRAF6‐null cells we found: (i) attenuation of TAK1 activity and abolishment of IKK activity and (ii) sustained binding of CARMA1 to Bcl10. To account for the molecular mechanism causing these dynamics, we performed a mathematical model analysis. The mathematical model analysis showed that the regulation of IKK activation by TRAF6 can reproduce TAK1 and IKK activities in TRAF6 null cells, and that the TRAF6 related signal‐dependent inhibitor suppresses CARMA1 binding to Bcl10 in wild‐type cells. These results suggest that TRAF6 contributes to the positive regulation of IKK activation via TAK1, alongside the negative signal‐dependent regulation of CARMA1 binding to Bcl10.
G protein-coupled receptor 3 (GPR3) constitutively activates Gαs proteins without any ligands and is predominantly expressed in neurons. Since the expression and physiological role of GPR3 in immune ...cells is still unknown, we examined the possible role of GPR3 in T lymphocytes. The expression of GPR3 was upregulated 2 h after phorbol 12-myristate 13-acetate (PMA)/ionomycin stimulation and was sustained in Jurkat cells, a human T lymphocyte cell line. In addition, the expression of nuclear receptor 4 group A member 2 (NR4A2) was highly modulated by GPR3 expression. Additionally, GPR3 expression was linked with the transcriptional promoter activity of NR4A in Jurkat cells. In mouse CD4+ T cells, transient GPR3 expression was induced immediately after the antigen receptor stimulation. However, the expression of NR4A2 was not modulated in CD4+ T cells from GPR3-knockout mice after stimulation, and the population of Treg cells in thymocytes and splenocytes was not affected by GPR3 knockout. By contrast, spontaneous effector activation in both CD4+ T cells and CD8+ T cells was observed in GPR3-knockout mice. In summary, GPR3 is immediately induced by T cell stimulation and play an important role in the suppression of effector T cell activation.
Although the cellular concentration of miRNAs is critical to their function, how miRNA expression and abundance are regulated during ontogeny is unclear. We applied miRNA-, mRNA-, and ChIP-Seq to ...characterize the microRNome during lymphopoiesis within the context of the transcriptome and epigenome. We show that lymphocyte-specific miRNAs are either tightly controlled by polycomb group-mediated H3K27me3 or maintained in a semi-activated epigenetic state prior to full expression. Because of miRNA biogenesis, the cellular concentration of mature miRNAs does not typically reflect transcriptional changes. However, we uncover a subset of miRNAs for which abundance is dictated by miRNA gene expression. We confirm that concentration of 5p and 3p miRNA strands depends largely on free energy properties of miRNA duplexes. Unexpectedly, we also find that miRNA strand accumulation can be developmentally regulated. Our data provide a comprehensive map of immunity's microRNome and reveal the underlying epigenetic and transcriptional forces that shape miRNA homeostasis.
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► H3K27me3 inhibits expression of induced miRNAs during lymphopoiesis ► Lymphocyte-specific, poised miRNAs are not downregulated by H3K27me3 ► Fluctuations in 25% of all miRNAs are dictated by transcription in B cells ► miRNA strand accumulation can be developmentally regulated
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