Biomedical subjects

T A Chan

Publications and source records attributed to T A Chan.

10 recordsLinked to original sources

PubMed2001

Ferredoxin reductase affects p53-dependent, 5-fluorouracil-induced apoptosis in colorectal cancer cells.

Loss of p53 gene function, which occurs in most colon cancer cells, has been shown to abolish the apoptotic response to 5-fluorouracil (5-FU). To identify genes downstream of p53 that might mediate these effects, we assessed global patterns of gene expression following 5-FU treatment of isogenic cells differing only in their p53 status. The gene encoding mitochondrial ferredoxin reductase (protein, FR; gene, FDXR) was one of the few genes significantly induced by p53 after 5-FU treatment. The FR protein was localized to mitochondria and suppressed the growth of colon cancer cells when over-expressed. Targeted disruption of the FDXR gene in human colon cancer cells showed that it was essential for viability, and partial disruption of the gene resulted in decreased sensitivity to 5-FU-induced apoptosis. These data, coupled with the effects of pharmacologic inhibitors of reactive oxygen species, indicate that FR contributes to p53-mediated apoptosis through the generation of oxidative stress in mitochondria.

Antimetabolites, Antineoplastic↗

PubMed2000

Cooperative effects of genes controlling the G(2)/M checkpoint.

It is believed that multiple effectors independently control the checkpoints permitting transitions between cell cycle phases. However, this has not been rigorously demonstrated in mammalian cells. The p53-induced genes p21 and 14-3-3sigma are each required for the G(2) arrest and allow a specific test of this fundamental tenet. We generated human cells deficient in both p21 and 14-3-3sigma and determined whether the double knockout was more sensitive to DNA damage than either single knockout. p21(-/-) 14-3-3sigma(-/-) cells were significantly more sensitive to DNA damage or to the exogenous expression of p53 than cells lacking only p21 or only 14-3-3sigma. Thus, p21 and 14-3-3sigma play distinct but complementary roles in the G(2)/M checkpoint, and help explain why genes at the nodal points of growth arrest pathways, like p53, are the targets of mutation in cancer cells.

14-3-3 Proteins↗

PubMed2000

High frequency of hypermethylation at the 14-3-3 sigma locus leads to gene silencing in breast cancer.

Expression of 14-3-3 final sigma (final sigma) is induced in response to DNA damage, and causes cells to arrest in G(2). By SAGE (serial analysis of gene expression) analysis, we identified final sigma as a gene whose expression is 7-fold lower in breast carcinoma cells than in normal breast epithelium. We verified this finding by Northern blot analysis. Remarkably, final sigma mRNA was undetectable in 45 of 48 primary breast carcinomas. Genetic alterations at final sigma such as loss of heterozygosity were rare (1/20 informative cases), and no mutations were detected (0/34). On the other hand, hypermethylation of CpG islands in the final sigma gene was detected in 91% (75/82) of breast tumors and was associated with lack of gene expression. Hypermethylation of final sigma is functionally important, because treatment of final sigma-non-expressing breast cancer cell lines with the drug 5-aza-2'-deoxycytidine resulted in demethylation of the gene and synthesis of final sigma mRNA. Breast cancer cells lacking final sigma expression showed increased number of chromosomal breaks and gaps when exposed to gamma-irradiation. Therefore, it is possible that loss of final sigma expression contributes to malignant transformation by impairing the G(2) cell cycle checkpoint function, thus allowing an accumulation of genetic defects. Hypermethylation and loss of final sigma expression are the most consistent molecular alterations in breast cancer identified so far.

14-3-3 Proteins↗

PubMed1999

The neural cell adhesion molecules L1 and NCAM-180 act in different steps of neurite outgrowth.

The formation of neurocircuitry depends on the control of neurite outgrowth that, in turn, can be divided into two processes: nerve growth cone protrusion and neurite extension. It has long been known that the neural cell adhesion molecules L1 and NCAM-180 promote neurite outgrowth, but how they function in growth cones is unclear. We addressed the roles of L1 and NCAM-180 in neurite outgrowth by using microscale chromophore-assisted laser inactivation (micro-CALI) of these proteins to perturb their functions at precise times in single growth cones of embryonic chick dorsal root ganglion neurons grown in culture. Micro-CALI of L1 causes neurite retraction after a 10 min lag period but does not affect growth cone protrusion. In contrast, micro-CALI of NCAM-180 causes rapid growth cone retraction but does not affect neurite extension. The simultaneous inactivation of both these molecules resulted in both distinct effects that were segregated in time. The behavior of growth cones after these micro-CALI treatments resemble the drug-induced perturbation of microtubules for L1 and F-actin for NCAM-180. These findings suggest distinct roles in the growth cone for L1 and NCAM-180 in different steps of neurite outgrowth: L1 functions in neurite extension,whereas NCAM-180 functions in growth cone protrusion.

Animals↗

PubMed1999

PPARdelta is an APC-regulated target of nonsteroidal anti-inflammatory drugs.

PPARB was identified as a target of APC through the analysis of global gene expression profiles in human colorectal cancer (CRC) cells. PPARdelta expression was elevated in CRCs and repressed by APC in CRC cells. This repression was mediated by beta-catenin/Tcf-4-responsive elements in the PPARdelta promotor. The ability of PPARs to bind eicosanoids suggested that PPARdelta might be a target of chemopreventive non-steroidal anti-inflammatory drugs (NSAIDs). Reporters containing PPARdelta-responsive elements were repressed by the NSAID sulindac. Furthermore, sulindac was able to disrupt the ability of PPARdelta to bind its recognition sequences. These findings suggest that NSAIDs inhibit tumorigenesis through inhibition of PPARdelta, the gene for which is normally regulated by APC.

Adenomatous Polyposis Coli Protein↗

PubMed1999

14-3-3Sigma is required to prevent mitotic catastrophe after DNA damage.

14-3-3Sigma is a member of a family of proteins that regulate cellular activity by binding and sequestering phosphorylated proteins. It has been suggested that 14-3-3sigma promotes pre-mitotic cell-cycle arrest following DNA damage, and that its expression can be controlled by the p53 tumour suppressor gene. Here we describe an improved approach to the generation of human somatic-cell knockouts, which we have used to generate human colorectal cancer cells in which both 14-3-3sigma alleles are inactivated. After DNA damage, these cells initially arrested in the G2 phase of the cell cycle, but, unlike cells containing 14-3-3sigma, the 14-3-3sigma-/- cells were unable to maintain cell-cycle arrest. The 14-3-3sigma-/- cells died ('mitotic catastrophe') as they entered mitosis. This process was associated with a failure of the 14-3-3sigma-deficient cells to sequester the proteins (cyclin B1 and cdc2) that initiate mitosis and prevent them from entering the nucleus. These results may indicate a mechanism for maintaining the G2 checkpoint and preventing mitotic death.

14-3-3 Proteins↗

PubMed1998

Mechanisms underlying nonsteroidal antiinflammatory drug-mediated apoptosis.

Nonsteroidal antiinflammatory drugs (NSAIDs) can inhibit colorectal tumorigenesis and are among the few agents known to be useful for the chemoprevention of neoplasia. Here, we show that the tumor suppressive effects of NSAIDs are not likely to be related to a reduction in prostaglandins but rather are due to the elevation of the prostaglandin precursor arachidonic acid (AA). NSAID treatment of colon tumor cells results in a dramatic increase in AA that in turn stimulates the conversion of sphingomyelin to ceramide, a known mediator of apoptosis. These results have significant implications for understanding and improving colon cancer chemoprevention.

Anti-Inflammatory Agents, Non-Steroidal↗

PubMed1996

Converting cancer genes into killer genes.

Over the past decade, it has become clear that tumorigenesis is driven by alterations in genes that control cell growth or cell death. Theoretically, the proteins encoded by these genes provide excellent targets for new therapeutic agents. Here, we describe a gene therapy approach to specifically kill tumor cells expressing such oncoproteins. In outline, the target oncoprotein binds to exogenously introduced gene products, resulting in transcriptional activation of a toxic gene. As an example, we show that this approach can be used to specifically kill cells overexpressing a mutant p53 gene in cell culture. The strategy may be generally applicable to neoplastic diseases in which the underlying patterns of genetic alterations or abnormal gene expression are known.

Cell Line↗

PubMed1994

Identification of a gene encoding a novel protein-tyrosine kinase containing SH2 domains and ankyrin-like repeats.

Using the polymerase chain reaction with primers corresponding to conserved regions in the kinase domain of protein-tyrosine kinases, we amplified segments of several protein-tyrosine kinase genes from Hydra vulgaris, a member of the ancient metazoan phylum Cnidaria. Characterization of cDNA clones for one of these genes, HTK16, revealed that it encodes a non-receptor protein-tyrosine kinase with two SH2 domains but no SH3 domain. In this regard the predicted HTK16 protein resembles two mammalian non-receptor protein-tyrosine kinases, the products of the ZAP-70 and syk genes. However, the HTK16 protein contains five ankyrin-like repeats, a structural motif which has not previously been found in protein-tyrosine kinases. The HTK16 protein also contains a potential tyrosine phosphorylation site in its carboxyl-terminal tail which resembles the phosphorylation site in members of the src family. RNA hybridization analysis indicates that the HTK16 gene is expressed in epithelial cells, cells which also express the Hydra homologue of the src protein. Our finding of the HTK16 gene in Hydra indicates that diversification of genes encoding non-receptor protein-tyrosine kinases was a very early event in metazoan evolution.

Amino Acid Sequence↗

PubMedDate not supplied

IVE (Image Visualization Environment): a software platform for all three-dimensional microscopy applications.

IVE (Image Visualization Environment) is a software platform designed from the outset to handle all aspects of modern computerized multidimensional microscopy. This platform provides users with an execution environment in which 5D data (XYZ, wavelength, and time) can be easily manipulated for the purpose of data collection, processing, display, and analysis. During the entire process, powerful data display functions are readily available for extracting complicated three-dimensional information through data visualization. By employing both the shared memory and multitasking features of the UNIX operation system, individual functions can be implemented as separate programs, and multiple programs can access the same data pool simultaneously. This enables users to combine the functionalities of different programs to facilitate each unique data analysis task. Furthermore, by defining an appropriate program execution model, commonly shared functional components such as data display, data I/O and user interface, etc. can be implemented using simple IVE library calls. This dramatically reduces the program development time and ensures consistency throughout the entire software system. As a result, users can quickly master the microscopy software system and new functions can be easily integrated, as different functional requirements arise for different research projects.

Chromosomes↗