Complete nucleotide sequence of the influenza B/Singapore/222/79 virus hemagglutinin gene and comparison with the B/Lee/40 Hemagglutinin

The complete nucleotide sequence of the hemagglutinin (HA) gene of the human type B influenza virus B/Singapore/222/79 is presented. Comparison with the only other known sequence of a B hemagglutinin (B/Lee/40) shows that antigenic drift in type B HA genes is essentially the same as already observed within the influenza A H3 subtype, i.e., an accumulation of point mutations. The main difference is that the apparent evolution is significantly slower, most likely due to the cumulative effect of a lower occurrence in the population (slower evolution) and/or less immunological pressure. There is a striking cluster of changes at positions 127 until 137 of the HA1 subunit which may represent one of the antigenic sites of the molecule

TDP2 promotes repair of topoisomerase I-mediated DNA damage in the absence of TDP1

The abortive activity of topoisomerases can result in clastogenic and/or lethal DNA damage in which the topoisomerase is covalently linked to the 3'- or 5'-terminus of a DNA strand break. This type of DNA damage is implicated in chromosome translocations and neurological disease and underlies the clinical efficacy of an important class of anticancer topoisomerase 'poisons'. Tyrosyl DNA phosphodiesterase-1 protects cells from abortive topoisomerase I (Top1) activity by hydrolyzing the 3'-phosphotyrosyl bond that links Top1 to a DNA strand break and is currently the only known human enzyme that displays this activity in cells. Recently, we identified a second tyrosyl DNA phosphodiesterase (TDP2; aka TTRAP/EAPII) that possesses weak 3'-tyrosyl DNA phosphodiesterase (3'-TDP) activity, in vitro. Herein, we have examined whether TDP2 contributes to the repair of Top1-mediated DNA breaks by deleting Tdp1 and Tdp2 separately and together in murine and avian cells. We show that while deletion of Tdp1 in wild-type DT40 cells and mouse embryonic fibroblasts decreases DNA strand break repair rates and cellular survival in response to Top1-induced DNA damage, deletion of Tdp2 does not. However, deletion of both Tdp1 and Tdp2 reduces rates of DNA strand break repair and cell survival below that observed in Tdp1(-)(/)(-) cells, suggesting that Tdp2 contributes to cellular 3'-TDP activity in the absence of Tdp1. Consistent with this idea, over-expression of human TDP2 in Tdp1(-)(/)(-)/Tdp2(-)(/)(-)(/)(-) DT40 cells increases DNA strand break repair rates and cell survival above that observed in Tdp1(-)(/)(-) DT40 cells, suggesting that Tdp2 over-expression can partially complement the defect imposed by loss of Tdp1. Finally, mice lacking both Tdp1 and Tdp2 exhibit greater sensitivity to Top1 poisons than do mice lacking Tdp1 alone, further suggesting that Tdp2 contributes to the repair of Top1-mediated DNA damage in the absence of Tdp1. In contrast, we failed to detect a contribution for Tdp1 to repair Top2-mediated damage. Together, our data suggest that Tdp1 and Tdp2 fulfil overlapping roles following Top1-induced DNA damage, but not following Top2-induced DNA damage, in vivo

TDP2-Dependent Non-Homologous End-Joining Protects against Topoisomerase II-Induced DNA Breaks and Genome Instability in Cells and In Vivo

Anticancer topoisomerase >poisons> exploit the break-and-rejoining mechanism of topoisomerase II (TOP2) to generate TOP2-linked DNA double-strand breaks (DSBs). This characteristic underlies the clinical efficacy of TOP2 poisons, but is also implicated in chromosomal translocations and genome instability associated with secondary, treatment-related, haematological malignancy. Despite this relevance for cancer therapy, the mechanistic aspects governing repair of TOP2-induced DSBs and the physiological consequences that absent or aberrant repair can have are still poorly understood. To address these deficits, we employed cells and mice lacking tyrosyl DNA phosphodiesterase 2 (TDP2), an enzyme that hydrolyses 5′-phosphotyrosyl bonds at TOP2-associated DSBs, and studied their response to TOP2 poisons. Our results demonstrate that TDP2 functions in non-homologous end-joining (NHEJ) and liberates DSB termini that are competent for ligation. Moreover, we show that the absence of TDP2 in cells impairs not only the capacity to repair TOP2-induced DSBs but also the accuracy of the process, thus compromising genome integrity. Most importantly, we find this TDP2-dependent NHEJ mechanism to be physiologically relevant, as Tdp2-deleted mice are sensitive to TOP2-induced damage, displaying marked lymphoid toxicity, severe intestinal damage, and increased genome instability in the bone marrow. Collectively, our data reveal TDP2-mediated error-free NHEJ as an efficient and accurate mechanism to repair TOP2-induced DSBs. Given the widespread use of TOP2 poisons in cancer chemotherapy, this raises the possibility of TDP2 being an important etiological factor in the response of tumours to this type of agent and in the development of treatment-related malignancy.Gobierno Español SAF2010-21017, BFU2010-11042-EEuropean Union PERG07- 2010-268466Queen Elisabeth Medical Foundation GSKE 111

The transcription factor Zeb2 regulates development of conventional and plasmacytoid DCs by repressing Id2

Plasmacytoid dendritic cells (DCs [pDCs]) develop from pre-pDCs, whereas two lineages of conventional DCs (cDCs; cDC1s and cDC2s) develop from lineage-committed pre-cDCs. Several transcription factors (TFs) have been implicated in regulating the development of pDCs (E2-2 and Id2) and cDC1s (Irf8, Id2, and Batf3); however, those required for the early commitment of pre-cDCs toward the cDC2 lineage are unknown. Here, we identify the TF zinc finger E box-binding homeobox 2 (Zeb2) to play a crucial role in regulating DC development. Zeb2 was expressed from the pre-pDC and pre-cDC stage onward and highly expressed in mature pDCs and cDC2s. Mice conditionally lacking Zeb2 in CD11c(+) cells had a cell-intrinsic reduction in pDCs and cDC2s, coupled with an increase in cDC1s. Conversely, mice in which CD11c(+) cells overexpressed Zeb2 displayed a reduction in cDC1s. This was accompanied by altered expression of Id2, which was up-regulated in cDC2s and pDCs from conditional knock-out mice. Zeb2 chromatin immunoprecipitation analysis revealed Id2 to be a direct target of Zeb2. Thus, we conclude that Zeb2 regulates commitment to both the cDC2 and pDC lineages through repression of Id2

Transcriptional repressor ZEB2 promotes terminal differentiation of CD8⁺ effector and memory T cell populations during infection

ZEB2 is a multi-zinc-finger transcription factor known to play a significant role in early neurogenesis and in epithelial-mesenchymal transition-dependent tumor metastasis. Although the function of ZEB2 in T lymphocytes is unknown, activity of the closely related family member ZEB1 has been implicated in lymphocyte development. Here, we find that ZEB2 expression is up-regulated by activated T cells, specifically in the KLRG1(hi) effector CD8(+) T cell subset. Loss of ZEB2 expression results in a significant loss of antigen-specific CD8(+) T cells after primary and secondary infection with a severe impairment in the generation of the KLRG1(hi) effector memory cell population. We show that ZEB2, which can bind DNA at tandem, consensus E-box sites, regulates gene expression of several E-protein targets and may directly repress Il7r and Il2 in CD8(+) T cells responding to infection. Furthermore, we find that T-bet binds to highly conserved T-box sites in the Zeb2 gene and that T-bet and ZEB2 regulate similar gene expression programs in effector T cells, suggesting that T-bet acts upstream and through regulation of ZEB2. Collectively, we place ZEB2 in a larger transcriptional network that is responsible for the balance between terminal differentiation and formation of memory CD8(+) T cells

Zeb2 is a negative regulator of midbrain dopaminergic axon growth and target innervation

Neural connectivity requires neuronal differentiation, axon growth, and precise target innervation. Midbrain dopaminergic neurons project via the nigrostriatal pathway to the striatum to regulate voluntary movement. While the specification and differentiation of these neurons have been extensively studied, the molecular mechanisms that regulate midbrain dopaminergic axon growth and target innervation are less clear. Here we show that the transcription factor Zeb2 cell-autonomously represses Smad signalling to limit midbrain dopaminergic axon growth and target innervation. Zeb2 levels are downregulated in the embryonic rodent midbrain during the period of dopaminergic axon growth, when BMP pathway components are upregulated. Experimental knockdown of Zeb2 leads to an increase in BMP-Smad-dependent axon growth. Consequently there is dopaminergic hyperinnervation of the striatum, without an increase in the numbers of midbrain dopaminergic neurons, in conditional Zeb2 (Nestin-Cre based) knockout mice. Therefore, these findings reveal a new mechanism for the regulation of midbrain dopaminergic axon growth during central nervous system development

MicroRNAs promote skeletal muscle differentiation of mesodermal iPSC-derived progenitors

Muscular dystrophies (MDs) are often characterized by impairment of both skeletal and cardiac muscle. Regenerative strategies for both compartments therefore constitute a therapeutic avenue. Mesodermal iPSC-derived progenitors (MiPs) can regenerate both striated muscle types simultaneously in mice. Importantly, MiP myogenic propensity is influenced by somatic lineage retention. However, it is still unknown whether human MiPs have in vivo potential. Furthermore, methods to enhance the intrinsic myogenic properties of MiPs are likely needed, given the scope and need to correct large amounts of muscle in the MDs. Here, we document that human MiPs can successfully engraft into the skeletal muscle and hearts of dystrophic mice. Utilizing non-invasive live imaging and selectively induced apoptosis, we report evidence of striated muscle regeneration in vivo in mice by human MiPs. Finally, combining RNA-seq and miRNA-seq data, we define miRNA cocktails that promote the myogenic potential of human MiPs

MiR-200 family controls late steps of postnatal forebrain neurogenesis via Zeb2 inhibition

During neurogenesis, generation, migration and integration of the correct numbers of each neuron sub-Type depends on complex molecular interactions in space and time. MicroRNAs represent a key control level allowing the flexibility and stability needed f

3D genome organization during lymphocyte development and activation

Chromosomes have a complex three-dimensional (3D) architecture comprising A/B compartments, topologically associating domains and promoter-enhancer interactions. At all these levels, the 3D genome has functional consequences for gene transcription and therefore for cellular identity. The development and activation of lymphocytes involves strict control of gene expression by transcription factors (TFs) operating in a three-dimensionally organized chromatin landscape. As lymphocytes are indispensable for tissue homeostasis and pathogen defense, and aberrant lymphocyte activity is involved in a wide range of human morbidities, acquiring an in-depth understanding of the molecular mechanisms that control lymphocyte identity is highly relevant. Here we review current knowledge of the interplay between 3D genome organization and transcriptional control during B and T lymphocyte development and antigen-dependent activation, placing special emphasis on the role of TFs