Biomedical subjects

Zilong Wen

Publications and source records attributed to Zilong Wen.

8 recordsLinked to original sources

PubMed2006

The 5' zebrafish scl promoter targets transcription to the brain, spinal cord, and hematopoietic and endothelial progenitors.

The stem cell leukemia (SCL) gene encodes a basic helix-loop-helix transcription factor and is essential for embryonic angiogenesis, hematopoietic stem cell specification, and erythrocyte maturation. Here, we report the isolation and characterization of the zebrafish scl promoter. We show that a 5-kilobase (kb) genomic fragment immediately upstream of the translation start site is capable of targeting the enhanced green fluorescence protein (EGFP) expression to the anterior and posterior lateral mesoderm where the endogenous scl normally expresses. Detailed analysis of the stable transgenic fish reveals that this 5-kb upstream sequence is sufficient to direct the EGFP transcription to the brain, spinal cord, and hematopoietic-endothelial progenitors, possibly the hemangioblast, but not primitive erythrocyte, suggesting that the zebrafish scl transcription in hematopoietic-endothelial progenitors and erythrocyte is regulated by distinct cis element(s). Our study has defined the cis regulatory element(s) for zebrafish scl expression in the brain, spinal cord, and hematopoietic-endothelial progenitors and established a valuable transgenic line Tg(5'5kbscl:EGFP) for studying hematopoietic lineage development.

Animals↗

PubMed2005

Loss of function of def selectively up-regulates Delta113p53 expression to arrest expansion growth of digestive organs in zebrafish.

Transcription factor p53 forms a network with associated factors to regulate the cell cycle and apoptosis in response to environmental stresses. However, there is currently no direct genetic evidence to show if or how the p53 pathway functions during organogenesis. Here we present evidence to show that the zebrafish def (digestive-organ expansion factor) gene encodes a novel pan-endoderm-specific factor. A loss-of-function mutation in def confers hypoplastic digestive organs and selectively up-regulates the expression of Delta113p53, counterpart to a newly identified isoform of p53 produced by an alternative internal promoter in intron 4 of the p53 gene in human. The increased Delta113p53 expression is limited to within the mutant digestive organs, and this increase selectively induces the expression of p53-responsive genes to trigger the arrest of the cell cycle but not apoptosis, resulting in compromised organ growth in the mutant. Our data demonstrate that, while induction of expression of p53 and/or its isoforms is crucial to suppress abnormal cell growth, Delta113p53 is tightly regulated by an organ/tissue-specific factor Def, especially during organogenesis, to prevent adverse inhibition of organ/tissue growth.

Animals↗

PubMed2005

Genome-wide identification of female-enriched genes in zebrafish.

Characteristic differences in morphology, physiology, and behavior between a male and female are correlated to the differential selection of sex-dependent transcriptomes. By using a cDNA array carrying approximately 9,000 zebrafish unique genes, we identified a group of genes whose expression are enriched in the female fish. A subset of these genes have been confirmed and further grouped as egg-enriched genes, as both somatic- and egg-enriched genes or as somatic-enriched genes by means of RNA gel blot hybridization. Most importantly, a significant proportion of these genes are either functionally unknown or are novel genes. Thus, future studies of this group of genes will help us greatly to understand more about sex-determination and sex-related physiology and behavior. In addition, comparison of zebrafish female-enriched genes with that in Drosophila, we found that only germline genes are shared between vertebrate and invertebrate, suggesting that the process of oogenesis is highly conserved during the evolution.

Animals↗

PubMed2005

Microarray analysis of zebrafish cloche mutant using amplified cDNA and identification of potential downstream target genes.

Zebrafish is an excellent model organism for studying vertebrate development and human disease. With the availability of increased numbers of zebrafish mutants and microarray chips, gene expression profiling has become a powerful tool for identification of downstream target genes perturbed by a specific mutation. One of the obstacles often encountered, however, is to isolate large numbers of zebrafish mutant embryos that are indistinguishable in morphology from the wild-type siblings for microarray analysis. Here, we report a method using amplified cDNA derived from five embryos for gene expression profiling of the 18-somite zebrafish cloche (clo) mutant, in which development of hematopoietic and endothelial lineages is severely impaired. In total, 31 differentially expressed target genes are identified, of which 13 have not been reported previously. We further determine that of these 13 new targets, 8 genes, including coproporphyrinogen oxidase (cpo), carbonic anhydrase (cahz), claudin g (cldn g), zinc-finger-like gene 2 (znfl2), neutrophil cytosol factor 1 (ncf1), matrix metalloproteinase 13 (mmp13), dual specificity phosphatase 5 (dusp5), and a novel gene referred as zebrafish vessel-specific gene 1 (zvsg1) are predominantly expressed in hematopoietic and endothelial cells. Comparative analysis demonstrates that this method is comparable and complementary to that of the conventional approach using unamplified sample. Our study provides valuable information for studying hematopoiesis and vessel formation. The method described here offers a powerful tool for gene expression profiling of zebrafish mutants in general.

Animals↗

PubMed2005

Identification of the conserved serine/threonine residues important for gibberellin-sensitivity of Arabidopsis RGL2 protein.

The DELLA proteins GAI, RGA, RGL1 and RGL2 in Arabidopsis are plant growth repressors, repressing diverse developmental processes. Studies have shown that gibberellin (GA) attenuates the repressive function of DELLA proteins by triggering their degradation via the proteasome pathway. However, it is not known if GA-induced protein degradation is the only pathway for regulating the bioactivity of DELLA proteins. We show here that tobacco BY2 cells represent a suitable system for studying GA signaling. RGL2 exists in a phosphorylated form in BY2 cells. RGL2 undergoes GA-induced degradation, and this process is blocked by proteasome inhibitors and serine/threonine phosphatase inhibitors; however, serine/threonine kinase inhibitors had no detectable effect, suggesting that dephosphorylation of serine/threonine is probably a prerequisite for degradation of RGL2 via the proteasome pathway. Site-directed substitution of all 17 conserved serine and threonine residues showed that six mutants (RGL2(S441D, RGL2(S542D), RGL2(T271E), RGL2(T319E), RGL2(T411E) and RGL2(T535E)) mimicking the status of constitutive phosphorylation are resistant to GA-induced degradation. This suggests that these sites are potential phosphorylation sites. A functional assay based on the expression of GA 20-oxidase revealed that RGL2(T271E) is probably a null mutant, RGL2(S441D), RGL2(S542D), RGL2(T319E) and RGL2(T411E) only retained about 4-17% of the activity of the wild type RGL2, whereas RGL2(T535E) retained about 66% of the activity of the wild type RGL2. However, expression of GA 20-oxidase in BY2 cells expressing these mutant proteins is still responsive to GA, suggesting that the stabilization of RGL2 protein is not the only pathway for regulating its bioactivity.

Arabidopsis↗

PubMed2003

15000 unique zebrafish EST clusters and their future use in microarray for profiling gene expression patterns during embryogenesis.

A total of 15590 unique zebrafish EST clusters from two cDNA libraries have been identified. Most significantly, only 22% (3437) of the 15590 unique clusters matched 2805 (of 15200) clusters in the Danio rerio UniGene database, indicating that our EST set is complementary to the existing ESTs in the public database and will be invaluable in assisting the annotation of genes based on the upcoming zebrafish genome sequence. Blast search showed that 7824 of our unique clusters matched 6710 known or predicted proteins in the nonredundant database. A cDNA microarray representing approximately 3100 unique zebrafish cDNA clusters has been generated and used to profile the gene expression patterns across six different embryonic stages (cleavage, blastula, gastrula, segmentation, pharyngula, and hatching). Analysis of expression data using K-means clustering revealed that genes coding for muscle-specific proteins displayed similar expression patterns, confirming that the coordinate gene expression is important for myogenesis. Our results demonstrate that the combination of microarray technology with the zebrafish model system can provide useful information on how genes are coordinated in a genetic network to control zebrafish embryogenesis and can help to identify novel genes that are important for organogenesis.

Animals↗

PubMed2002

Cloning and expression pattern of the lysozyme C gene in zebrafish.

Here, we report isolation and developmental expression pattern of the zebrafish lysozyme C gene. Amino acid sequence analysis showed that the zebrafish lysozyme C protein shared approximately 37-80% identities with the mouse, human, chicken, and carp counterparts. Whole-mount in situ hybridization showed that the lysozyme C gene was expressed in macrophages, as its expression was co-localized with the known myeloid lineage markers L-plastin and PU.1. At 20 hours postfertilization (hpf), most of the lysozyme C positive cells were localized in the yolksac and head mesenchyme but not in the intermediate cell mass, supporting the notion that the primitive macrophage originated from the yolksac (Development 126 (1999) 3735). At 36hpf, the lysozyme C positive cells scattered within the head and yolksac, and began to appear in the caudal part of axial vein. By 6 days postfertilization (dpf), the lysozyme C positive cells accumulated in the kidney where hematopoiesis had been indicated to take place after 4dpf (Dev. Dyn. 214 (1999) 323). Taken together, our results demonstrate that the lysozyme C gene is specifically expressed in myeloid lineage, suggesting that it could serve as an excellent marker for genetic screening of both primitive and definitive myeloid lineage development in zebrafish.

Amino Acid Sequence↗

PubMed2002

Chromatin-remodelling factor BRG1 selectively activates a subset of interferon-alpha-inducible genes.

Brahma-related gene 1 (BRG1 ) is a key component of the ATP-dependent chromatin-remodelling SWI2-SNF2 complex and has been implicated in regulating gene expression, cell-cycle control and tumorigenesis. Here we report that BRG1 interacts with signal transducer and activator of transcription 2 (STAT2) - a transcription factor that regulates gene expression mediated by interferon-alpha (IFN-alpha). BRG1 enhances the IFN-alpha-induced expression of 9-27 and IFI27 but not that of four other target genes tested, showing that the activation of different target genes by STAT2 may involve alternative chromatin modifiers. Our results also suggest that the recruitment and activation of BRG1 may require other cis-acting and trans-acting elements in addition to STAT2. Our study links the SWI2-SNF2 complex to the regulation of cytokine-induced gene expression and may identify a molecular mechanism of BRG1-mediated gene activation and tumorigenesis.

Amino Acid Sequence↗