Mutations in TPRN Cause a Progressive Form of Autosomal-Recessive Nonsyndromic Hearing Loss

We performed genome-wide homozygosity mapping in a large consanguineous family from Morocco and mapped the autosomal-recessive nonsyndromic hearing loss (ARNSHL) in this family to the DFNB79 locus on chromosome 9q34. By sequencing of 62 positional candidate genes of the critical region, we identified a causative homozygous 11 bp deletion, c.42_52del, in the TPRN gene in all seven affected individuals. The deletion is located in exon 1 and results in a frameshift and premature protein truncation (p.Gly15AlafsX150). Interestingly, the deleted sequence is part of a repetitive and CG-rich motive predicted to be prone to structural aberrations during crossover formation. We identified another family with progressive ARNSHL linked to this locus, whose affected members were shown to carry a causative 1 bp deletion (c.1347delG) in exon 1 of TPRN. The function of the encoded protein, taperin, is unknown; yet, partial homology to the actin-caping protein phostensin suggests a role in actin dynamics.     

Molecular cloning, expression and characterization of the zebrafish bram1 gene, a BMP receptor-associated molecule

Summary We have identified a cDNA clone encoding BMP receptor-associated molecule 1 (BRAM1) from the zebrafish expressed sequence tag (EST) database. The 2606 bp full-length bram1 cDNA was cloned, and further confirmed by nucleotide sequencing. The zebrafish sequence encodes a protein of 195 amino acids with an evolutionarily conserved MYND domain, which displays ∼ ∼98% homology with human and mouse BRAM1, and ∼ ∼64% homology with C. elegans BRA-1 and BRA-2. The bram1 gene, composed of five exons and four introns, spans ∼ ∼14 kb on linkage group 14 of the zebrafish genome. RT-PCR and whole mount in situ hybridization analyses disclosed that zebrafish BRAM1 is a maternal factor. The protein interacts directly with zebrafish BMP Receptor type IA, as observed from GST-pull down and co-immunoprecipitation assays. Furthermore, cotransfection of zebrafish BRAM1 with the corresponding BMP receptor resulted in down-regulation of BMP-mediated signaling. Our results collectively indicate that BRAM1 plays a biological role during zebrafish development.     

Isolation, cDNA, and Genomic Structure of a Conserved Gene (NOF) at Chromosome 11q13 Next to FAU and Oriented in the Opposite Transcriptional Orientation

In our effort to characterize a gene at chromosome 11q13 involved in a t(11;17)(q13;q21) translocation in B-non-Hodgkin lymphoma, we have identified a novel human gene, NOF (Neighbour of FAU). It maps right next to FAU in a head to head configuration separated by a maximum of 146 nucleotides. cDNA clones representing NOF hybridized to a 2.2-kb mRNA present in all tissues tested. The largest open reading frame appeared to contain 166 amino acids and is proline rich, and the sequence shows no homology with any known gene in the public databases. The NOF gene consists of 4 exons and 3 introns spanning approximately 5 kb, and the boundaries between exons and introns follow the GT/AG rule. The NOF locus is conserved during evolution, with the predicted protein having over 80% identity to three translated mouse and rat ESTs of unknown function. Moreover, the mouse ESTs map in the same organization, closely linked to the FAU gene, in the mouse genome. NOF, however, is not affected by the t(11;17)(q13;q21) chromosomal translocation.     

Identification of the hRDH-E2 gene,a novel member of the SDR family,and its increased expression in psoriatic lesion

To identify novel psoriasis-associated genes, we focused on several ESTs (expressed sequence tags) whose expression was predominantly increased in the affected skin in patients with psoriasis vulgaris, as assessed by microarray assay. In this paper, a full-length cDNA corresponding to one of those ESTs (AI440266) was isolated by screening of cultured human keratinocyte cDNA libraries. This cDNA has an open reading frame of a 309-amino-acid protein, sharing significant homology to one of the short-chain alcohol dehydrogenase/reductase (SDR) families that can catalyze the first and rate-limiting step that generates retinaldehyde from retinol. So, this gene was designated as hRDH-E2 (human epidermal retinal dehydrogenase 2). The hRDH-E2 gene has a single functional copy on chromosome 8q12.1, spanning approximately 20kb with seven exons. The deduced amino acid sequence contains three motifs that are conserved in the SDR family. Qualitative RT-PCR demonstrated that the mRNA levels of hRDH-E2 were significantly elevated in the affected skin in psoriasis patients as compared to the unaffected skin in patients and the normal skin in healthy individual. These results suggest that hRDH-E2 may be involved in the pathogenesis of psoriasis through its critical role in retinol metabolism in keratinocyte proliferation.     

一个鼻咽癌相关EST的鉴定及其全长cDNA序列分析

鼻咽癌是我国南方及东南亚地区常见的恶性肿瘤之一.通过对鼻咽癌染色体高频率杂合性丢失区域3p21的表达序列标签(expressedsequencetag,EST)进行同源性比较分析,运用逆转录聚合酶链式反应的方法,筛选到一个在41.18%(14/34)的鼻咽癌活检组织及20.0%(1/5)的鼻咽癌细胞系中表达下调的ESTBG772301;并用Northern杂交方法,检测了该EST在多种正常成人组织中的表达状况及其所代表基因的转录本大小.在此基础上,对该EST来源的cDNA克隆(IMAGE:4839190)进行直接测序,获得了一个全长为2377bp的新cDNA序列;经生物信息学分析,发现它与已知基因序列无明显同源性,属于一个新基因,定位于染色体3p21.3,被命名为鼻咽癌表达下调基因(NPCEDRG,GenBank登录号:AF538150).其编码的蛋白质含169个氨基酸,与一个已报道的在进化上相对保守、功能未知的人类蛋白Nicolin1(简称NICN1)N端170个氨基酸残基的序列同源性为97%,但缺少NICN1蛋白C端43个氨基酸残基,可能是nicolin1基因不同剪接本的编码产物.     

cDNA, genomic sequence cloning and overexpression of ribosomal protein S25 gene (RPS25) from the Giant Panda

RPS25 is a component of the 40S small ribosomal subunit encoded by RPS25 gene, which is specific to eukaryotes. Studies in reference to RPS25 gene from animals were handful. The Giant Panda (Ailuropoda melanoleuca), known as a “living fossil”, are increasingly concerned by the world community. Studies on RPS25 of the Giant Panda could provide scientific data for inquiring into the hereditary traits of the gene and formulating the protective strategy for the Giant Panda. The cDNA of the RPS25 cloned from Giant Panda is 436 bp in size, containing an open reading frame of 378 bp encoding 125 amino acids. The length of the genomic sequence is 1,992 bp, which was found to possess four exons and three introns. Alignment analysis indicated that the nucleotide sequence of the coding sequence shows a high homology to those of Homo sapiens, Bos taurus, Mus musculus and Rattus norvegicus as determined by Blast analysis, 92.6, 94.4, 89.2 and 91.5%, respectively. Primary structure analysis revealed that the molecular weight of the putative RPS25 protein is 13.7421 kDa with a theoretical pI 10.12. Topology prediction showed there is one N-glycosylation site, one cAMP and cGMP-dependent protein kinase phosphorylation site, two Protein kinase C phosphorylation sites and one Tyrosine kinase phosphorylation site in the RPS25 protein of the Giant Panda. The RPS25 gene was overexpressed in E. coli BL21 and Western Blotting of the RPS25 protein was also done. The results indicated that the RPS25 gene can be really expressed in E. coli and the RPS25 protein fusioned with the N-terminally his-tagged form gave rise to the accumulation of an expected 17.4 kDa polypeptide. The cDNA and the genomic sequence of RPS25 were cloned successfully for the first time from the Giant Panda using RT-PCR technology and Touchdown-PCR, respectively, which were both sequenced and analyzed preliminarily; then the cDNA of the RPS25 gene was overexpressed in E. coli BL21 and immunoblotted, which is the first report on the RPS25 gene from the Giant Panda. The data will enrich and supplement the information about RPS25, which will contribute to the protection for gene resources and the discussion of the genetic polymorphism.     

Two Rep Genes in Small Cryptic Plasmid pKST21 of Escherichia coli

The complete nucleotide sequence of a small cryptic plasmid pKST21 from Escherichia coli was determined. This plasmid is 1,460 bp long with an overall GC content of 51 %. Based on sequence analysis, the presence of two segments with different average GC density was observed. The segment with higher GC content revealed 98–90 % similarity to several small plasmids of E. coli and to pCR1 from Gram-positive Corynebacterium renale. Plasmid pKST21 possesses two conversely oriented open reading frames encoding proteins with a high degree of amino acid identity to Rep proteins involved in replication. ORF1 encodes replication protein similar to RepA protein of Bartonella tribocorum or Bacillus cereus plasmids or to the putative plasmid Rep protein from ecologically close Selenomonas ruminantium. ORF2 similarly encodes a replication protein, which shares 97 % homology with Rep protein from C. renale. Genetic diversity observed in plasmid pKST21 indicates a mosaic structure of the plasmid with different segments acquired from different sources. Deletion analysis showed that both fragments carrying the repA and repB genes are necessary for the replication of pKST21 in E. coli. The presence of plasmid with the same gene composition was revealed in 14 % of tested E. coli isolates from the rumen of sheep. All these strains produced identical ERIC-PCR profiles indicating isogenic origin of the strain and lack of horizontal gene transfer of pKST21 plasmid.