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Schistosoma infection is a major cause of morbidity and mortality worldwide. Schistosomiasis japonica is endemic in mainland China along the Yangtze River, typically distributed in two geographical categories of lake and mountainous regions. Study on schistosome genetic diversity is of interest in respect of understanding parasite biology and transmission, and formulating control strategy. Certain genetic variations may be associated with adaptations to different ecological habitats. The aim of this study is to gain insight into Schistosoma japonicum genetic variation, evolutionary origin and associated causes of different geographic lineages through examining homozygous Single Nucleotide Polymorphisms (SNPs) based on resequenced genome data. We collected S. japonicum samples from four sites, three in the lake regions (LR) of mid-east (Guichi and Tonglin in Anhui province, Laogang in Hunan province) and one in mountainous region (MR) (Xichang in Sichuan province) of south-west of China, resequenced their genomes using Next Generation Sequencing (NGS) technology, and made use of the available database of S. japonicum draft genomic sequence as a reference in genome mapping. A total of 14,575 SNPs from 2059 genes were identified in the four lineages. Phylogenetic analysis confirmed significant genetic variation exhibited between the different geographical lineages, and further revealed that the MR Xichang lineage is phylogenetically closer to LR Guich lineage than to other two LR lineages, and the MR lineage might be evolved from LR lineages. More than two thirds of detected SNPs were nonsynonymous; functional annotation of the SNP-containing genes showed that they are involved mainly in biological processes such as signaling and response to stimuli. Notably, unique nonsynonymous SNP variations were detected in 66 genes of MR lineage, inferring possible genetic adaption to mountainous ecological condition.

SummaryAttenuated DNA repair leads to genomic instability and tumorigenesis. BRCA1/BARD1 are the best known tumor suppressors that promote homology recombination (HR) and arrest cell cycle at G2/M checkpoint. As E3 ubiquitin ligases, their ubiquitinase activity has been known to involve in the HR and tumor suppression, but the mechanism remains ambiguous. Here, we demonstrated upon genotoxic stress, BRCA1 together with BARD1 catalyzed the K48 ployubiquitination on LARP7, a 7SK RNA binding protein known to control RNAPII pausing, and thereby degraded it through 26S ubiquitin-proteasome pathway. Depleting LARP7 suppressed the expression of CDK1 complex, arrested cell at G2/M DNA damage checkpoint and reduced BRCA2 phosphorylation which thereby facilitated RAD51 recruitment to damaged DNA to enhance HR. Importantly, LARP7 depletion observed in breast patients lead to the chemoradiotherapy resistance both in vitro and in vivo. Together, this study unveils a mechanism by which BRCA1/BARD1 utilizes their E3 ligase activity to control HR and cell cycle, and highlights LARP7 as a potential target for cancer prevention and therapy.HighlightsDNA damage response downregulates LARP7 through BRCA1/BARD1BRCA1/BARD1 catalyzes the K48 polyubiquitination on LARP7LARP7 promotes G2/M cell cycle transition and tumorigenesis via CDK1 complexLARP7 disputes homology-directed repair that leads to tumor therapy resistance

POSH (plenty of SH3) is a scaffold protein that has been shown to act as an E3 ubiquitin ligase. Here we report that POSH stimulates the ubiquitination of Kir1.1 (ROMK) and enhances the internalization of this potassium channel. Immunostaining reveals the expression of POSH in the renal cortical collecting duct. Immunoprecipitation of renal tissue lysate with ROMK antibody and glutathione S-transferase pulldown experiments demonstrated the association between ROMK and POSH. Moreover, immunoprecipitation of lysates of HEK293T cells transfected with ROMK1 or with constructs encoding the ROMK-N terminus or ROMK1-C-Terminus demonstrated that POSH binds to ROMK1 on its N terminus. To study the effect of POSH on ROMK1 channels, we measured potassium currents with electrophysiological methods in HEK293T cells and in oocytes transfected or injected with ROMK1 and POSH. POSH decreased potassium currents, and the inhibitory effect of POSH on ROMK channels was dose-dependent. Biotinylation assay further showed that POSH decreased surface expression of ROMK channels in HEK293T cells transfected with ROMK1 and POSH. The effect of POSH on ROMK1 channels was specific because POSH did not inhibit sodium current in oocytes injected with ENaC-alpha, beta, and gamma subunits. Moreover, POSH still decreased the potassium current in oocytes injected with a ROMK1 mutant (R1Delta373-378), in which a clathrin-dependent tyrosine-based internalization signal residing between amino acid residues 373 and 378 is deleted. However, the inhibitory effect of POSH on ROMK channels was absent in cells expressing with dominant negative dynamin and POSHDeltaRING, in which the RING domain was deleted. Expression of POSH also increased the ubiquitination of ROMK1, whereas expression of POSHDeltaRING diminished its ubiquitination in HEK293T cells. The notion that POSH may serve as an E3 ubiquitin ligase is also supported by in vitro ubiquitination assays in which adding POSH increased the ROMK ubiquitination. We conclude that POSH inhibits ROMK channels by enhancing dynamin-dependent and clathrin-independent endocytosis and by stimulating ubiquitination of ROMK channels.

The liver performs a number of essential functions for life. The development of such a complex organ relies on finely regulated gene expression profiles which change over time in the development and determine the phenotype and function of the liver. We used high-density oligonucleotide microarrays to study the gene expression and transcription regulation at 14 time points across the C57/B6 mouse liver development, which include E11.5 (embryonic day 11.5), E12.5, E13.5, E14.5, E15.5, E16.5, E17.5, E18.5, Day0 (the day of birth), Day3, Day7, Day14, Day21, and normal adult liver. With these data, we made a comprehensive analysis on gene expression patterns, functional preferences and transcriptional regulations during the liver development. A group of uncharacterized genes which might be involved in the fetal hematopoiesis were detected.

The cDNA of a Schistosoma japonicum myophilin-like protein was cloned, sequenced, and expressed in Escherichia coli as a recombined protein (rSj myophilin-like protein), and the protein was purified by affinity chromatography. The deduced amino acid sequences of the Sj myophilin-like protein showed significant homology to myophilin, calponin, Np22 and Mp20. Northern blot and RT-PCR analyzes revealed expression of the Sj myophilin-like protein mRNA in eggs, sporocysts, cercariae, hepatic schistosomula and adult worms. Confocal fluorescence microscopy localized the native protein to the muscle of the adult worm. In schistosome-infected rabbits, the rSj myophilin-like protein antibody level, assessed by ELISA, was elevated after infection but was reduced after praziquantel treatment. In humans, the myophilin-like protein antibody level was evaluated by ELISA in sera from 33 non-infected humans and 61 schistosomiasis patients; the results showed a highly significant difference between the two groups with a sensitivity of 57.4%. Taken together, the myophilin-like protein may prove useful for monitoring the therapeutic effect of praziquantel rather than in serodiagnosis of schistosomiasis.

A novel human gene, named as human CAP10-like protein 46 kDa (hCLP46), was isolated and identified from human acute myeloid leukemia transformed from myelodysplastic syndrome (MDS-AML) CD34(+) cells. hCLP46 (3q13.33) contains 11 exons encoding a putative protein of 392 amino acids, with a highly conserved CAP10 domain, a hydrophobic signal peptide at its N-terminus, and an endoplasmic reticulum (ER) retention signal motif KTEL at the C-terminus. The homologs of hCLP46 exist in different organisms from plants to animal kingdoms. Subcellular localization analysis showed that hCLP46 is an ER-resident protein. hCLP46 expressed in most human adult tissues at different intensities, with lengths of 3.5 kb and 1.9 kb. Transcript of hCLP46 was not detectable in colon, thymus, and small intestine, but was abundant in liver, indicating that hCLP46 may be involved in important physiological functions in the liver. hCLP46 over-expressed U937 cells had higher growth rate than the cells without exogenic hCLP46 protein expression, suggesting that hCLP46 protein possess the ability of promoting cell proliferation.

Pituitary, a master gland of neuroendocrine system, secretes hormones that orchestrate many physiological processes, under the regulation of multiple signaling pathways. To investigate the genes involved in hormones expression of human pituitary, homemade cDNA microarray containing 14,800 human genes/ESTs were used to profile the gene expression in both fetal and adult pituitaries. Seven hundred and twelve known genes changed over 2-fold between the both tissues. Of which, 23 genes were changed with hormones expression in aging were confirmed by RT-PCR, not only the known regulators such as Pit1, GATA4, ESRRA, GABA-A, and EMK, but also LOC55884, DUSP3, PNN, and RCL, which had not been reported to be involved in the hormones expression. Correspondingly, the mRNAs of GH, PRL, POMC, TSH-beta, FSH-beta, and LH-beta, was increased as much as 6- to 20-fold in adult pituitary than those in fetal pituitary, by real-time quantitative RT-PCR assay. In addition, the mRNAs of signaling pathways, such as cAMP-PKA-CREB, PI3K-Akt, and PKA-ERK were further investigated. Of them, it was only cAMP-PKA-CREB pathway, but not PI3K-Akt and PKA-ERK have the same expressing pattern as hormones. It suggested that cDNA microarray is highly advantages to profile the differential expressed genes that were involved in hormones expression of human pituitary, but it might ignore some responding proteins regulated posttranscriptionally.

The Chinese genome project was initiated in 1993 with the goal of contributing 1% to the Human Genome Program. The study of gene expression profiles with cDNA microarrays, and large-scale sequencing and analysis of 130 928 expressed sequence tags (ESTs), allowed isolation and characterization of over 1000 novel full-length human cDNAs derived from human hematopoietic stem/progenitor cells, neuroendocrine tissues, liver, and cardiovascular cells. In addition, EST sequencing for model organisms, including rat, zebrafish, Schistosoma japonicum and rice was performed, aiming at identifying genes associated with physiological and/or pathological characteristics. To cite this article: Z.-G. Han et al., C. R. Biologies 326 (2003).

In order to fully understand the physiological functions of adipose tissue, especially its secretory functions, and to provide a basis for the identification of novel obesity related genes, the gene expression profiling of human visceral adipose tissue was established by using cDNA array. 33 P -labelled cDNA, derived from visceral adipose tissue total RNA, was hybridized to a cDNA array containing over 16,000 expressed sequence-tagged clones which represent human singleton genes. The expressed sequence tag (EST) was considered to be expressed in visceral adipose tissue when the ratio of signal to noise was greater than or equal to 2. The results were analyzed with bioinformatics. Totally, 8230 genes were found to be expressed in visceral adipose tissue with 5200 known genes and 3030 known ESTs. Most of 84 secretory proteins, 120 receptors, and 74 transcription factors expressed in adipose tissue were newly identified. Many appetite-regulating related peptides or receptors and some reproduction-related genes were first found to be expressed in adipose tissue. Eight autocrine/paracrine systems were described for the first time in the visceral adipose tissue. These results clearly demonstrate that the visceral adipose tissue has important secretory functions and there is a complex local autocrine/paracrine regulatory network. The present work suggests that the visceral adipose tissue is an important component of the neuroendocrine-immune network and plays an important role in regulating appetite not only via endocrine but also via autocrine/paracrine systems. The visceral adipose tissue might also play a role in regulating reproduction and sexual function.