Identification and Characterization of Genes That Interact with Lin-12 in Caenorhabditis Elegans

We identified and characterized 14 extragenic mutations that suppressed the dominant egg-laying defect of certain lin-12 gain-of-function mutations. These suppressors defined seven genes: sup-17, lag-2, sel-4, sel-5, sel-6, sel-7 and sel-8. Mutations in six of the genes are recessive suppressors, whereas the two mutations that define the seventh gene, lag-2, are semi-dominant suppressors. These suppressor mutations were able to suppress other lin-12 gain-of-function mutations. The suppressor mutations arose at a very low frequency per gene, 10-50 times below the typical loss-of-function mutation frequency. The suppressor mutations in sup-17 and lag-2 were shown to be rare non-null alleles, and we present evidence that null mutations in these two genes cause lethality. Temperature-shift studies for two suppressor genes, sup-17 and lag-2, suggest that both genes act at approximately the same time as lin-12 in specifying a cell fate. Suppressor alleles of six of these genes enhanced a temperature-sensitive loss-of-function allele of glp-1, a gene related to lin-12 in structure and function. Our analysis of these suppressors suggests that the majority of these genes are part of a shared lin-12/glp-1 signal transduction pathway, or act to regulate the expression or stability of lin-12 and glp-1.     

Identification and Characterization of Autosomal Genes That Interact with Glass in the Developing Drosophila Eye

The glass gene encodes a zinc finger, DNA-binding protein that is required for photoreceptor cell development in Drosophila melanogaster. In the developing compound eye, glass function is regulated at two points: (1) the protein is expressed in all cells' nuclei posterior to the morphogenetic furrow and (2) the ability of the Glass protein to regulate downstream genes is largely limited to the developing photoreceptor cells. We conducted a series of genetic screens for autosomal dominant second-site modifiers of the weak allele glass(3), to discover genes with products that may regulate glass function at either of these levels. Seventy-six dominant enhancer mutations were recovered (and no dominant suppressors). Most of these dominant mutations are in essential genes and are associated with recessive lethality. We have assigned these mutations to 23 complementation groups that include multiple alleles of Star and hedgehog as well as single alleles of Delta, roughened eye, glass and hairy. Mutations in 18 of the complementation groups are embryonic lethals, and of these, 13 show abnormal adult retinal phenotypes in homozygous clones, usually with altered numbers of photoreceptor cells in some of the ommatidia.     

杜氏盐藻两种碳酸酐酶基因启动子的克隆和功能研究

将克隆得到的杜氏盐藻DCA7和CA基因的启动子区与bar基因和NOS polyA终止子片段融合,分别构建成pMDDC-B和pMDC-B转基因杜氏盐藻表达载体。用基因枪法将两种表达载体转化人杜氏盐藻细胞,通过除草剂草丁膦筛选培养获得转化藻株,对转化藻株进行分析。对转化杜氏盐藻藻株的筛选培养结果表明：pMDDC-B和pMDC-B载体中的外源bar基因能在杜氏盐藻细胞中稳定或瞬时表达。同时在氦气压力为690kPa条件下,微弹轰击2次比微弹轰击1次或3次的效果更好。对pMDDC-B转化杜氏盐藻得到的稳定表达的转化藻株进行的PCR和Southern印迹分析的结果表明：外源的bar基因确已整合到杜氏盐藻基因组中。Northern印迹分析表明：DCA7基因启动子驱动bar基因在杜氏盐藻细胞中的表达效率受氯化钠浓度梯度调控。推测首次克隆得到的DCA7基因启动子可能是一种活性高、安全性好的高渗诱导性启动子;杜氏盐藻DCA7和CA基因启动子区的GT高度重复序列,可能与杜氏盐藻高度耐盐的分子机制有关。     

Evidence for a Set of Closely Linked Autosomal Genes That Interact with Sex-Chromosome Heterochromatin in DROSOPHILA MELANOGASTER

It is proposed that there exists a special region in the euchromatin of the left arm of chromosome 2 (contained within sections 31-32 of the standard salivary gland chromosome map) that is defined by a set of genes, each one of which interacts with a specific sex-chromosome heterochromatic segment. The evidence for the existence of this region is, first, the exhibition, mapping, and analysis of five different maternal-effect, embryonic semi-lethals located in region 31-32. Secondly, in each case the consequence of the maternal effect is markedly influenced by the amount of X- or Y-chromosome heterochromatin carried by the progeny of mutant mothers. The nature of this interaction and possible reasons for the existence of the cluster of autosomal genes are discussed.     

Two Closely Linked Mutations in DROSOPHILA MELANOGASTER That Are Lethal to opposite Sexes and Interact with Daughterless

A new spontaneous mutation named Sex-lethal, Male-specific No. 1 (SxlM1) is described that is lethal to males, even in the presence of an Sxl+ duplication. Females homozygous for SxlM1 are fully viable. This dominant, male-specific lethal mutation is on the X chromosome approximately 0.007 map units to the right of a previously isolated female-specific mutation, Female-lethal, here renamed Sex-lethal, Female-specific No. 1 (SxlF1). SxlM1 and SxlF1 are opposite in nearly every repect, particularly with regard to their interaction with maternal effect of the autosomal mutation, daughterless (da). Females that are homozygous for da produce defective eggs that cannot support female (XX) development. A single dose of SxlM1 enables daughters to survive this da female-specific lethal maternal effect. A duplication of the Sxl locus weakly mimics this action of SxlM1. In contrast, SxlF1 and a deficiency for Sxl, strongly enhance the female-lethal effects of da. The actions of SxlM1 and SxlF1 are explained by a model in which expression of the Sxl locus is essential for females, lethal for males, and under the control of a product of the da locus. It is suggested that SxlM1 is a constitutive mutation at the Sxl locus.     

Identification of Mutations in Three Genes That Interact with Zeste in the Control of White Gene Expression in Drosophila Melanogaster

Three previously described genes, enhancer of yellow, 1, 2 and 3, are shown to cooperate with the zeste gene in the control of white gene expression. The mutations e(y)1(u1), e(y)3(u1), and to a lesser extent e(y)2(u1), enhance the effect of the zeste null allele z(v77h). Different combinations of e(y)1(u1), e(y)2(u1) and e(y)3(u1) mutations with several other z alleles also enhance the white mutant phenotype, but only to levels characteristic of white alleles containing a deletion of the upstream eye enhancer. Loss of zeste protein binding sites from the white locus does not eliminate the effect of e(y)1(u1) and e(y)3(u1) mutations, suggesting that the products of these genes interact with some other nucleotide sequences. Combinations of either e(y)1(u1) or e(y)2(u1) mutations with e(y)3(u1) are lethal. The products of these three genes may represent, together with zeste, a group of proteins involved in the organization of long-distance interactions between DNA sequences.     

Suppressors of a Lin-12 Hypomorph Define Genes That Interact with Both Lin-12 and Glp-1 in Caenorhabditis Elegans

The lin-12 gene of Caenorhabditis elegans is thought to encode a receptor which mediates cell-cell interactions required to specify certain cell fates. Reversion of the egg-laying defective phenotype caused by a hypomorphic lin-12 allele identified rare extragenic suppressor mutations in five genes, sel-1, sel-9, sel-10, sel-11 and sel(ar40) (sel = suppressor and/or enhancer of lin-12). Mutations in each of these sel genes suppress defects associated with reduced lin-12 activity, and enhance at least one defect associated with elevated lin-12 activity. None of the sel mutations cause any obvious phenotype in a wild-type background. Gene dosage experiments suggest that sel-1 and sel(ar40) mutations are reduction-of-function mutations, while sel-9 and sel-11 mutations are gain-of-function mutations. sel-1, sel-9, sel-11 and sel(ar40) mutations do not suppress amorphic lin-12 alleles, while sel-10 mutations are able to bypass partially the requirement for lin-12 activity in at least one cell fate decision. sel-1, sel-9, sel-10, sel-11 and sel(ar40) mutations are also able to suppress the maternal-effect lethality caused by a partial loss-of-function allele of glp-1, a gene that is both structurally and functionally related to lin-12. These sel genes may therefore function in both lin-12 and glp-1 mediated cell fate decisions.     

两个鼻咽癌负相关新基因的分离与特性

8个通过 c DNA代表差异分析法 ( c DNA representational difference analysis,c DNA RDA)分离的新 c DNA序列中 ,经 RT- PCR验证 ,发现其中一 c DNA序列 (登录号 :AF0 91 51 7)在 40 %的鼻咽癌活检组织中存在表达缺失和下调 .Northern杂交显示 ,AF0 91 51 7代表转录本为 1 .1 kb和 1 .4kb大小的两个基因 ,进而采用文库筛选 ,成功分离出 3′端完全不同的两个基因 ,命名为 NAG1 1和 NAG 1 2 (登录号分别为 AF 1 70 30 7和 AF 1 94971 ) .经过计算机预测 ,NAG 1 1编码 87个氨基酸组成的跨膜蛋白 ,NAG1 2编码 1 36个氨基酸组成的可溶性的核蛋白 ,两者无任何同源性 .NAG 1 1蛋白含有 3个 ATP结合区、两个蛋白激酶 C磷酸化位点和两个 N-肉豆寇酸化位点 ,NAG 1 2含有POU结构域和多个功能位点 .结果说明 NAG1 1和 NAG1 2的表达的缺失与下调可能参与了鼻咽癌的进程 ;NAG1 1基因产物可能与 ATP的跨膜转运有关 ;NAG1 2基因产物可能与转录翻译有关 .     

Novel Mutations Found in Two Genes of Thai Patients with Isolated Methylmalonic Acidemia

Molecular genetic analysis of three patients diagnosed with isolated methylmalonic acidemia (MMA) revealed that one was mut ⁰ MMA, with a mutation in the MUT gene encoding the l-methylmalonyl-CoA mutase (MCM), and two were cblB MMA, with mutations in the MMAB gene required for synthesizing the deoxyadenosylcobalamin cofactor of MCM. The mut ⁰ patient was homozygous for a novel nonsense mutation in MUT, p.R31X (c.167C → T), and heterozygous for three previously described polymorphisms, p.K212K (c.712A → G), p.H532R (c.1671A → G), and p.V671I (c.2087G → A). The new MMAB mutation, p.E152X (c.454G → T), was found to be homozygous in one cblB patient and heterozygous in the other patient, who also had four intron polymorphisms in this gene.     

Mosaic Analysis of Two Genes That Affect Nervous System Structure in Caenorhabditis elegans

The mutation mec-4(e 1611), identified by M. Chalfie, leads to the degeneration and death of the six neurons, called the microtubule cells, that mediate the response of wild-type animals to light touch. The fates of two of these cells, PLML and PLMR, which are responsible for response to light touch in the tail of the animal, have been monitored in animals mosaic for the mec-4(e 1611) mutation. The results are consistent with the view that the mutation behaves cell autonomously in its killing effect; in particular, none of the neurons that make either chemical synapses or gap junctions to PLML or PLMR is responsible for the deaths of PLML or PLMR. The results of gene dosage and dominance tests suggest that the mec-4(+) gene product, which is required for wild-type microtubule cell function, is altered by the e 1611 mutation into a novel product that kills the microtubule cells. Mutation in the gene unc-3 leads to the derangement of the processes of the motor neurons of the ventral cord. Mosaic analysis strongly suggests that unc-3(+) expression is required only in the motor neurons themselves for normal neuronal development. In particular, the hypodermis surrounding the ventral cord is not the primary focus of unc-3 action (body muscle was excluded in earlier work). Finally, the mosaic analysis supports an earlier suggestion that a sensory defect caused by a daf-6 mutation is localized to a non-neuronal cell called the sheath cell.     

Expression of Ethylene Biosynthetic Genes Regulated by Pollination associated Factors in Doritaenopsis hybrida Flowers

Pollination of flowers initiates postpollination development in orchid ( Doritaenopsis hybrida Hort. ) flowers, including perianth senescence, stigma closure, and ovary development. Because ethylene is thought to play a key role in coordinating these developmental changes, the authors studied the temporal and spatial patterns of expression of genes encoding 1-aminocyclopropane-l-carboxylic acid (ACC) synthase and ACC oxidase following pollination-associated factor treatments in orchid flowers. Both ACC synthase and ACC oxidase mRNA accumulation in the various parts of the flowers is induced by auxin, and ethylene, but not by emasculation. The patterns of both ACC synthase and ACC oxidase mRNA accumulation are similar in all floral organs following auxin and ethylene treatments. Further, in situ hybridization analysis indicates that the ACC oxidase mRNA is localized in epidermal and parenchyma cells of the stigma after auxin and ethylene treatments. The putative roles of auxin, ethylene and emasculation are discussed in terms of the regulation of ACC synthase and ACC oxidase gene expression in flowers.     

Isolation of the Promoters of Atlantic Salmon MHCII Genes

The major histocompatibility complex class II (MHCII) has a central role in the immune response of vertebrates with its function of presenting antigenic peptides to the T-cell receptors. We have isolated the promoters and intron 1 of MHCII and MHCII genes of Atlantic salmon. To isolate these promoters, we constructed an Atlantic salmon (Salmo salar) promoter finder kit (analogous to the commercially available human promoter finder kit). By nucleotide sequence alignment of known MHCII promoter regions, we identified the 3 conserved regulatory X, X2, and Y boxes in the salmon promoters. The W box was not found. In contrast, a salmon-specific putative W box was identified. Both of the isolated Atlantic salmon MHCII and promoters (included in patent applications by Genomar A/S, Oslo, Norway) were found to be functional since they both gave positive yellow fluorescence protein signal when inserted as promoters in the pEYFP-1 reporter plasmid and transfected into the salmon head kidney cell line (SHK-1).     

与授粉有关的因子调节的乙烯生物合成基因在兰花花器官中的表达

分析了与授粉有关的因子调节的ＡＣＣ合酶和ＡＣＣ氧化酶基因在朵丽蝶兰（ＤｏｒｉｔａｅｎｏｐｓｉｓｈｙｂｒｉｄａＨｏｒｔ．）花中的表达。生长素和乙烯均可诱导ＡＣＣ合酶和ＡＣＣ氧化酶的ｍＲＮＡ在花器官中积累。然而,去雄却不能诱导这两个基因在花器官中表达。生长素和乙烯所诱导的ＡＣＣ合酶和ＡＣＣ氧化酶的ｍＲＮＡ在花器官中的积累模式相似。原位杂交结果表明,生长素和乙烯处理后ＡＣＣ氧化酶的ｍＲＮＡ在柱头的表皮和薄壁细胞中积累。根据ＡＣＣ合酶和ＡＣＣ氧化酶基因表达的结果,对生长素、乙烯和去雄在兰花授粉后乙烯生物合成过程中的作用进行了分析。