Somatic hypermutation in B cells is initiated by activation-induced cytidine deaminase-catalyzed C→U deamination at immunoglobulin variable regions. Here we investigate the role of the germinal centre-associated nuclear protein (GANP) in enhancing the access of activation-induced cytidine deaminase (AID) to immunoglobulin variable regions. We show that the nuclear export factor GANP is involved in chromatin modification at rearranged immunoglobulin variable loci, and its activity requires a histone acetyltransferase domain. GANP interacts with the transcription stalling protein Spt5 and facilitates RNA Pol-II recruitment to immunoglobulin variable regions. Germinal centre B cells from ganp-transgenic mice showed a higher AID occupancy at the immunoglobulin variable region, whereas B cells from conditional ganp-knockout mice exhibit a lower AID accessibility. These findings suggest that GANP-mediated chromatin modification promotes transcription complex recruitment and positioning at immunoglobulin variable loci to favour AID targeting.

RNA export factor germinal center–associated nuclear protein (GANP) interacts with activation-induced cytidine deaminase (AID) and shepherds it from the cytoplasm to the nucleus and toward the IgV region loci in B cells. In this study, we demonstrate a role for GANP in the repair of AID-initiated DNA damage in chicken DT40 B cells to generate IgV region diversity by gene conversion and somatic hypermutation. GANP plays a positive role in IgV region diversification of DT40 B cells in a nonhomologous end joining–proficient state. DNA-PKcs physically interacts with GANP, and this interaction is dissociated by dsDNA breaks induced by a topoisomerase II inhibitor, etoposide, or AID overexpression. GANP affects the choice of DNA repair mechanism in B cells toward homologous recombination rather than nonhomologous end joining repair. Thus, GANP presumably plays a critical role in protection of the rearranged IgV loci by favoring homologous recombination of the DNA breaks under accelerated AID recruitment.

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Zinc finger protein St18 was initially reported as candidate tumor suppressor gene, and also suggested that fibroblast St18 positively regulates NF-kB activation. Despite the pleiotropic functions of St18, little is known about its roles in macrophages. Here, we report that myeloid St18 is a potent inhibitor of VEGF-A. Mice lacking St18 in myeloid lineages exhibit increased retinal vasculature with enhanced serum VEGF-A concentrations. Despite the normal activation of NF-kB target genes, these mice are highly susceptible to LPS-induced shock, polymicrobial sepsis, and experimental colitis, accompanied by enhanced vascular and intestinal leakage. Pharmacological inhibition of VEGF signaling rescued the high mortality rate of myeloid-specific St18-deficient mice in response to inflammation. Mechanistically, St18 directly binds to Sp1 and attenuates its activity, leading to the suppression of Sp1 target gene VEGF-A. Using mouse genetic and pharmacological models, we reveal myeloid St18 as a critical septic death protector.

GANP maintains antibody structural integrity during SHM

The single-stranded DNA-dependent deoxycytidine deaminase APOBEC3G (A3G) is a potent restrictive factor against HIV-1 virus lacking viral-encoded infectivity factor (Vif) in CD4+ T cells. A3G antiretroviral activity requires its encapsulation into HIV-1 virions. Here we show that germinal center-associated nuclear protein (GANP) is induced in activated CD4+ T cells and physically interacts with A3G. Overexpression of GANP augments the A3G encapsidation into the virion-like particles and ΔVif HIV-1 virions. GANP is encapsidated in HIV-1 virion and modulates A3G packaging into the cores together with cellular RNAs including 7SL RNA, and with unspliced HIV-1 genomic RNA. GANP upregulation leads to a significant increase in A3G-catalyzed G→A hypermutation in the viral genome and suppression of HIV-1 infectivity in a single-round viral infection assay. Conversely, GANP knockdown caused a marked increase in HIV-1 infectivity in a multiple rounds infection assay. The data suggest that GANP is a cellular factor that facilitates A3G encapsidation into HIV-1 virions to inhibit the viral infectivity.

Chronic myeloid leukemia (CML) respond dramatically to molecular target therapy; imatinib (IM), a first generation tyrosine kinase inhibitor (TKIs). Quantitation of cytokines like Interleukin-6, Interleukin-7 and Transforming growth factor-α plasma levels before IM therapy, could assess early molecular response (EMR) to IM and predict imatinib failure. A case-control study of 30 CML patients and 30 controls. Levels of IL-6, IL-7 and TGF-α were assayed by ELISA (R&D systems, USA) for both controls and patients. The patients' BCR-ABL1 transcript was assayed by real time-quantitative polymerase chain reaction, using ipsogen® BCR-ABL1 Mbcr Kit on the Rotor-Gene Q MDx (Qiagen, USA). Cytokines and BCR-ABL1 levels were done both before therapy and at 3 months follow up.Three months following IM therapy, the patients were divided into improved (n= 27) and non-improved (n= 3) groups; based on the establishment of EMR. Plasma levels of IL-7, IL-6 and TGF-α were significantly higher in CML patients (p< 0.05). Cytokines plasma levels dropped significantly after IM therapy (p< 0.05). Correlation studies revealed a strong positive correlation between pretreatment levels of both IL-6 and TGF-α and posttreatment levels of BCR-ABL transcript (r= 0.89 and 0.84, respectively). IL-7 showed a poor correlation with posttreatment levels of BCR-ABL transcript (r= 0.32). Our study revealed a possible role of IL-6, IL-7 and TGF-α as mediators of CML. The initial high levels of IL-6 and TGF-α was associated with the failure of achieving EMR. The initial high levels of IL-7 in CML patients appears to facilitate the disease process.