Structural basis for inhibition of the replication licensing factor Cdt1 by geminin

To maintain chromosome stability in eukaryotic cells, replication origins must be licensed by loading mini-chromosome maintenance (MCM2-7) complexes once and only once per cell cycle. This licensing control is achieved through the activities of geminin and cyclin-dependent kinases. Geminin binds tightly to Cdt1, an essential component of the replication licensing system, and prevents the inappropriate reinitiation of replication on an already fired origin. The inhibitory effect of geminin is thought to prevent the interaction between Cdt1 and the MCM helicase. Here we describe the crystal structure of the mouse geminin-Cdt1 complex using tGeminin (residues 79-157, truncated geminin) and tCdt1 (residues 172-368, truncated Cdt1). The amino-terminal region of a coiled-coil dimer of tGeminin interacts with both N-terminal and carboxy-terminal parts of tCdt1. The primary interface relies on the steric complementarity between the tGeminin dimer and the hydrophobic face of the two short N-terminal helices of tCdt1 and, in particular, Pro 181, Ala 182, Tyr 183, Phe 186 and Leu 189. The crystal structure, in conjunction with our biochemical data, indicates that the N-terminal region of tGeminin might be required to anchor tCdt1, and the C-terminal region of tGeminin prevents access of the MCM complex to tCdt1 through steric hindrance.     

The Cdt1 protein is required to license DNA for replication in fission yeast

To maintain genome stability in eukaryotic cells, DNA is licensed for replication only after the cell has completed mitosis, ensuring that DNA synthesis (S phase) occurs once every cell cycle. This licensing control is thought to require the protein Cdc6 (Cdc18 in fission yeast) as a mediator for association of minichromosome maintenance (MCM) proteins with chromatin. The control is overridden in fission yeast by overexpressing Cdc18 (ref. 11) which leads to continued DNA synthesis in the absence of mitosis. Other factors acting in this control have been postulated and we have used a re-replication assay to identify Cdt1 (ref. 14) as one such factor. Cdt1 cooperates with Cdc18 to promote DNA replication, interacts with Cdc18, is located in the nucleus, and its concentration peaks as cells finish mitosis and proceed to S phase. Both Cdc18 and Cdt1 are required to load the MCM protein Cdc21 onto chromatin at the end of mitosis and this is necessary to initiate DNA replication. Genes related to Cdt1 have been found in Metazoa and plants (A. Whitaker, I. Roysman and T. Orr-Weaver, personal communication), suggesting that the cooperation of Cdc6/Cdc18 with Cdt1 to load MCM proteins onto chromatin may be a generally conserved feature of DNA licensing in eukaryotes.     

Two forms of transforming growth factor-beta distinguished by multipotential haematopoietic progenitor cells

Type-beta transforming growth factors (TGF-beta s) are polypeptides that act hormonally to control proliferation and differentiation of many cell types. Two distinct homodimeric TGF-beta polypeptides, TGF-beta 1 and TGF-beta 2 have been identified which show approximately 70% amino-acid sequence similarity. Despite their structural differences, TGF-beta 1 and TGF-beta 2 are equally potent at inhibiting epithelial cell proliferation and adipogenic differentiation. The recent immunohistochemical localization of high levels of TGF-beta in the bone marrow and haematopoietic progenitors of the fetal liver has raised the possibility that TGF-beta s might be involved in the regulation of haematopoiesis. Here we show that TGF-beta 1, but not TGF-beta 2, is a potent inhibitor of haematopoietic progenitor cell proliferation. TGF-beta 1 inhibited colony formation by murine factor-dependent haematopoietic progenitor cells in response to interleukin-3 (IL-3) or granulocyte-macrophage colony stimulating factor (GM-CSF), as well as colony formation by marrow progenitor cells responding to CSF-1 (M-CSF). The progenitor cell lines examined were approximately 100-fold more sensitive to TGF-beta 1 than TGF-beta 2, and displayed type-I TGF-beta receptors with affinity approximately 20-fold higher for TGF-beta 1 than TGF-beta 2. These results identify TGF-beta 1 as a novel regulator of haematopoiesis that acts through type-I TGF-beta receptors to modulate proliferation of progenitor cells in response to haematopoietic growth factors.     

转录因子GATA—1基因在白血病中的表达

转录因子ＧＡＴＡ家族在造血细胞的正常发育中起着重要的作用。利用聚合酶链反应方法分析了１１６例各种白血病中红系统特异转录因子ＧＡＴＡ－１的表达情况。ＡＮＬＬ、ＣＭＬ、Ｃ－ＡＬＬ和ＣＬＬ中的表达率分别为４３．７５％、８８．２４％、１４．２９％和３３．３３％；３例Ｔ－ＡＬＬ均不表达该基因。     

Leukaemia inhibitory factor is identical to the myeloid growth factor human interleukin for DA cells

Leukaemia inhibitory factor (LIF) is a cytokine that induces macrophage differentiation of the murine M1 myeloid leukaemia cell line. We have isolated a cDNA clone encoding a novel human haemopoietic growth factor, human interleukin for DA cells (HILDA) that supports the proliferation of the murine interleukin-3-dependent leukaemic cell line, DA-la (refs 3-5). HILDA proved to be identical to LIF. The demonstration that the differentiation factor LIF will also serve as a growth factor for at least one myeloid leukaemic cell line provides further evidence that the distinction between growth-promoting and differentiation-inducing activities are largely determined by the target cell type.     

As2O3对脐血红系祖细胞HOXB6基因表达的调控作用

目的：探讨脐血造血干祖细胞向红系祖细胞增殖过程中HOXB6mRNA表达及As2O3对HOXB6mRNA表达的影响．方法：①采用外培养技术,以As2O3干扰造血干祖细胞,观察HSPC经促红素诱导后,CFU-E集落生成情况．②采用实时荧光定量HR技术检测造血祖细胞增殖分化过程中HOXB6基因的表达．结果：①造血于祖细胞红系祖细胞增殖过程中,各组细胞HOXB6基因均表达,②与正常对照组比较,As2O3可下调HOXB6基因的表达．结论：①HOXB6可能是造血干祖细胞向红系祖细胞增殖分化中的调控基因之一．②As2O3能下调HOXB6基因的表达．     

Changes in geniculate cell size following brief monocular blockade of retinal activity in kittens

When a kitten is subjected to monocular lid suture early in life, cells in laminae of the lateral geniculate nucleus (LGN) connected to the sutured eye grow less than normal and cells in those laminae connected to the non-sutured eye grow more than normal. These changes are seen primarily in the binocular segment of the LGN, which corresponds to the central visual field, and are due to competition either between intracortical afferents originating from the different LGN laminae, or directly among cells within the LGN. The afferent deprivation induced by lid suture, however, is not complete, as retinal ganglion cells fire tonically both in darkness and in light. It is generally thought that this tonic retinal activity is necessary to maintain neuronal excitability at normal threshold in the central visual pathway. In the visual cortex of developing kittens, we previously showed a long-lasting change in ocular dominance of binocular cells by a brief blockade of retinal activity in one optic nerve. We report here that a complete blockade of retinal activity in one eye causes major changes in LGN cell size within 1 week. These changes occur throughout the LGN, including the monocular segment where binocular competition does not occur. The results indicate that tonic retinal activity may have an important role in the control of geniculate cell size.     

rhsBLyS活性检测方法的比较

以FluroBeads B荧光磁珠法体外分离并培养人B淋巴细胞作为研究材料，通过MTT法、^3H-TdR掺入法测定重组入B淋巴细胞刺激因子(rhsBLyS)对人B淋巴细胞的促增殖活性并进行相应比较．结果表明：rhsBLyS非融合蛋白在引发剂Anti-IgM的协同作用下表现出显著刺激B淋巴细胞增殖的活性，且两种方法所得结果一致，充分验证了rhsBLyS样品的活性；MTT法检测可替代放射性同位素法．