Cloning, structure, expression analysis, and assignment to mouse Chromosome 7 of the gene Zfp296 encoding a zinc finger protein

Received: 4 May 2000 / Accepted: 16 June 2000     

Structure, expression and in vitro functional characterization of a novel RNA binding zinc finger protein from Xenopus.

Large multigene families of zinc finger proteins are expressed in vertebrates. One way of approaching their function is to characterize their structure, expression and biochemical properties. XFG 5-1 is a Xenopus zinc finger protein which is widely transcribed in oocytes, embryos and adult tissues. It carries a novel, non-finger repeat structure, which is common to a subfamily of Xenopus zinc finger proteins. The bacterially expressed protein exhibits specific RNA homopolymer binding activities with the zinc finger domain being sufficient for this ability. These findings suggest that XFG 5-1 serves a general biological function involving its RNA binding capacity.     

甘蔗锌指蛋白基因ShSAP1的克隆与表达模式

植物中具有A20/AN1锌指结构域的蛋白与逆境应答密切相关,在甘蔗热带种Saccharum officinarum拔地拉Badila中克隆到一个具有A20/AN1锌指结构域的锌指蛋白基因ShSAP1。为研究ShSAP1的基因结构和表达特性,采用PCR和Southern blotting分析了ShSAP1的基因组结构,通过半定量RT-PCR对ShSAP1在甘蔗不同部位、不同胁迫和不同激素处理下的表达模式进行了分析。结果表明,ShSAP1的5'UTR区有两段内含子,大小分别为202 bp和1 052 bp,在     

一个与小鼠锌指蛋白基因ZF—12相关的假基因的克隆和鉴定

小鼠类Krueppel锌指蛋白基因ZF-12为人的类Krueppel锌指蛋白基因ZNF191的同源基因,它们都编码368个氨基酸残基,N端存在SCAN结构域,C端有4个连续的锌指模体,最近的研究表明ZNF191是肝癌发生的相关基因,我们以人锌指蛋白基因ZNF191的vDNA为探针,筛选小鼠λ噬菌体基因组文库,意外地获得了1个与小鼠锌指蛋白基因ZF-12相类似的基因,多种组织的RT-PCR和启动子序列分析,暗示该基因不表达,且该基因无内含子,与ZF-12高度相似,存在突变,暗示其为与ZF-12相关的假基因序列,经查新证实它为新的序列后,以ZF12p（ZF-12pseudogene)命名在GenBank登录（AY040222）。查录GenBank的人类基因组库以及Southern结果显示人类基因组中无ZNF191假基因序列,ZF12p与ZF-12高度相似,暗示ZF12p在进化过程中产生的时间较晚,这对研究锌指蛋白基因ZF-12的突变与进化具有重要的意义。     

Cloning and characterization of a novel human gene encoding a zinc finger protein with 25 fingers

This study reports cloning and characterization of a human cDNA encoding a novel human zinc finger protein, ZFD25. ZFD25 cDNA is 6118 bp long and has an open reading frame of 2352 bp that encodes a 783 amino acid protein with 25 C2H2-type zinc fingers. The ZFD25 cDNA also contains a region with high sequence similarity to the Krüppel-associated box A and B domain in the 5'-untranslated region, suggesting that ZFD25 belongs to the Krüppel-associated box zinc finger protein family. The ZFD25 gene was localized to chromosome 7q11.2. Northern blot analysis showed that ZFD25 was expressed in a wide range of human organs. In cultured endothelial cells, the mRNA level was decreased upon serum starvation.     

Identification of four CCCH zinc finger proteins in Xenopus, including a novel vertebrate protein with four zinc fingers and severely restricted expression

Tristetraprolin (TTP), the prototype of a class of CCCH zinc finger proteins, is a phosphoprotein that is rapidly and transiently induced by growth factors and serum in fibroblasts. Recent evidence suggests that a physiological function of TTP is to inhibit tumor necrosis factor alpha secretion from macrophages by binding to and destabilizing its mRNA (Carballo, E., Lai, W.S., Blackshear, P.J., 1998. Science, 281, 1001-1005). To investigate possible functions of CCCH proteins in early development of Xenopus, we isolated four Xenopus cDNAs encoding members of this class. Based on 49% overall amino acid identity and 84% amino acid identity within the double zinc finger domain, one of the Xenopus proteins (XC3H-1) appears to be the homologue of TTP. By similar analyses, XC3H-2 and XC3H-3 are homologues of ERF-1 (cMG1, TIS11B) and ERF-2 (TIS11D). A fourth protein, XC3H-4, is a previously unidentified member of the CCCH class of vertebrate zinc finger proteins; it contains four Cx8Cx5Cx3H repeats, two of which are YKTEL Cx8Cx5Cx3H repeats that are closely related to sequences found in the other CCCH proteins. Whereas XC3H-1, XC3H-2, and XC3H-3 were widely expressed in adult tissues, XC3H-4 mRNA was not detected in any of the adult tissues studied except for the ovary. Its expression appeared to be limited to the ovary, oocyte, egg and the early embryonic stages leading up to the mid-blastula transition. Its mRNA was highly expressed in oocytes of all ages, and was enriched in the animal pole cytosol of mature oocytes. Maternal expression was also seen with the other three messages, suggesting the possibility that these proteins are involved in regulating mRNA stability in oocyte maturation and/or early embryogenesis.     

Inducible gene expression in transgenic Xenopus embryos

The amphibian Xenopus laevis has been successfully used for many years as a model system for studying vertebrate development. Because of technical limitations, however, molecular investigations have mainly concentrated on early stages. We have developed a straightforward method for stage-specific induction of gene expression in transgenic Xenopus embryos [1] [2]. This method is based on the Xenopus heat shock protein 70 (Xhsp70 [3]) promoter driving the expression of desired gene products. We found that ubiquitous expression of the transgene is induced upon relatively mild heat treatment. Green fluorescent protein (GFP) was used as a marker to monitor successful induction of gene expression in transgenic embryos. We used this method to study the stage specificity of Wnt signalling function. Transient ectopic Wnt-8 expression during early neurulation was sufficient to repress anterior head development and this capacity was restricted to early stages of neurulation. By transient over-expression at different stages of development, we show that frizzled-7 disrupted morphogenesis sequentially from anterior to posterior along the dorsal axis as development proceeds. These results demonstrate that this method for inducible gene expression in transgenic Xenopus embryos will be a very powerful tool for temporal analysis of gene function and for studying molecular mechanisms of vertebrate organogenesis.     

Cloning and expression pattern of a Xenopus pronephros-specific gene, XSMP-30

The first step in kidney development is the formation of the pronephros which is derived from mesoderm. Xenopus is an appropriate model to study this process since the pronephros can be efficiently induced in animal cap explants by treatment with activin and retinoic acid (RA). Using this in vitro system, we isolated a Xenopus homologue of SMP-30 (Senescence marker protein-30), which is a Ca(2+)-binding protein that is highly conserved in vertebrates. This gene, termed XSMP-30, was found to be selectively expressed in pronephric tubules from the late tadpole stage, by whole mount in situ hybridization. Furthermore XSMP-30 was expressed in animal caps treated with both activin and RA, a condition in which the pronephros is formed in vitro. These data indicate that XSMP-30 is a specific marker for the pronephros.     

Differential antero-posterior expression of two proteins encoded by a homeobox gene in Xenopus and mouse embryos.

The X.laevis XlHbox 1 gene uses two functional promoters to produce a short and a long protein, both containing the same homeodomain. In this report we use specific antibodies to localize both proteins in frog embryos. The antibodies also recognize the homologous proteins in mouse embryos. In both mammalian and amphibian embryos, expression of the long protein starts more posteriorly than that of the short protein. This difference in spatial expression applies to the nervous system, the segmented mesoderm and the internal organs. This suggests that each promoter from this gene has precisely restricted regions of expression along the anterior-posterior axis of the embryo. Because the long and short proteins share a common DNA-binding specificity but differ by an 82 amino acid domain, their differential distribution may have distinct developmental consequences.