Homologous recombination but not nucleotide excision repair plays a pivotal role in tolerance of DNA-protein cross-links in mammalian cells.

DNA-protein cross-links (DPCs) are unique among DNA lesions in their unusually bulky nature. The steric hindrance imposed by cross-linked proteins (CLPs) will hamper DNA transactions, such as replication and transcription, posing an enormous threat to cells. In bacteria, DPCs with small CLPs are eliminated by nucleotide excision repair (NER), whereas oversized DPCs are processed exclusively by RecBCD-dependent homologous recombination (HR).

Phosphorylation of the cytokinesis regulator ECT2 at G2/M phase stimulates association of the mitotic kinase Plk1 and accumulation of GTP-bound RhoA.

The epithelial cell transforming gene 2 (ECT2) protooncogene encodes a Rho exchange factor, and regulates cytokinesis. ECT2 is phosphorylated in G2/M phases, but its role in the biological function is not known. Here we show that two mitotic kinases, Cdk1 and polo-like kinase 1 (Plk1), phosphorylate ECT2 in vitro.

TIM, a Dbl-related protein, regulates cell shape and cytoskeletal organization in a Rho-dependent manner.

The Dbl-like guanine nucleotide exchange factors (GEFs) have been implicated in direct activation of the Rho family of small GTPases. We previously isolated transforming immortalized mammary (TIM) as a Dbl-like protein. Here, we show that, when expressed in cells, TIM was a potent activator of RhoA.

Deregulation and mislocalization of the cytokinesis regulator ECT2 activate the Rho signaling pathways leading to malignant transformation.

The human ECT2 protooncogene encodes a guanine nucleotide exchange factor for the Rho GTPases and regulates cytokinesis. Although the oncogenic form of ECT2 contains an N-terminal truncation, it is not clear how the structural abnormality of ECT2 causes malignant transformation. Here we show that both the removal of the negative regulatory domain and alteration of subcellular localization are required to induce the oncogenic activity of ECT2.

Potential roles of the nucleotide exchange factor ECT2 and Cdc42 GTPase in spindle assembly in Xenopus egg cell-free extracts.

The ECT2 protooncogene encodes a guanine nucleotide exchange factor for the Rho family of small GTPases. ECT2 contains motifs of cell cycle regulators at its N-terminal domain. We previously showed that ECT2 plays a critical role in cytokinesis.

Rho exchange factor ECT2 is induced by growth factors and regulates cytokinesis through the N-terminal cell cycle regulator-related domains.

The ECT2 protooncogene plays a critical role in cytokinesis, and its C-terminal half encodes a Dbl homology-pleckstrin homology module, which catalyzes guanine nucleotide exchange on the Rho family of small GTPases. The N-terminal half of ECT2 (ECT2-N) contains domains related to the cell cycle regulator/checkpoint control proteins including human XRCC1, budding yeast CLB6, and fission yeast Cut5. The Cut5-related domain consists of two BRCT repeats, which are widespread to repair/checkpoint control proteins.

Suppression of anoikis by v-Src but not by activated c-H-ras in human gallbladder epithelial cells.

Detachment of anchorage-dependent normal epithelial cells from their substratum causes the type of apoptosis known as anoikis, whereas malignant cells can proliferate independently of anchorage. Because src and ras oncogenes are activated in many human cancers, we investigated their role and downstream signaling pathways in anoikis resistance, using HAG-1 human epithelial cells transfected with v-src or activated H-ras. Consequently, anchorage-dependent mock- or ras-transfected cells underwent anoikis.

Ostip2, a novel oncoprotein that associates with the Rho exchange factor Ost.

The ost protooncogene encodes a guanine nucleotide exchange factor for the Rho family of small GTPases, RhoA and Cdc42. The N-terminal domain of Ost (Ost-N) appears to negatively regulate the oncogenic activity of the protein, as deletion of this domain drastically increases its transforming activity in NIH 3T3 cells. Using a yeast two-hybrid system, we identified five genes encoding proteins that can interact with Ost-N.

Human ECT2 is an exchange factor for Rho GTPases, phosphorylated in G2/M phases, and involved in cytokinesis.

Animal cells divide into two daughter cells by the formation of an actomyosin-based contractile ring through a process called cytokinesis. Although many of the structural elements of cytokinesis have been identified, little is known about the signaling pathways and molecular mechanisms underlying this process. Here we show that the human ECT2 is involved in the regulation of cytokinesis.

Interferon-alpha and -gamma inhibit the growth and neoplastic potential of v-src-transformed human epithelial cells by reducing Src tyrosine kinase activity.

To investigate whether interferons (IFNs) selectively suppress the growth of solid tumor cells with elevated protein tyrosine kinase (PTK) activity, we evaluated the effect of recombinant IFN-alpha2a and IFN-gamma on the proliferative and neoplastic potentials triggered by p60v-src using v-src-transformed HAG-1 human epithelial cells. When compared with control cells harboring the pSV2neo gene, the monolayer growth of v-src-transformed cell lines was inhibited by both recombinant IFNs, in a dose-dependent manner, whereas growth of ras-transfected cell lines was not affected. Moreover, IFNs markedly reduced the clonogenic growth of v-src-transformed cells in soft-agar rather than monolayer growth, suggesting the preferential activity of IFNs on anchorage-independent growth.

Inhibition by 5-fluorouracil of ERCC1 and gamma-glutamylcysteine synthetase messenger RNA expression in a cisplatin-resistant HST-1 human squamous carcinoma cell line.

Pretreatment of 5-fluorouracil (5-FU), but not posttreatment, has been shown to augment the cytotoxicity of cisplatin (CDDP) or even circumvent CDDP resistance by inhibiting repair of platinum-DNA interstrand crosslinks as well as by reducing the cellular glutathione (GSH) contents in CDDP-resistant HST-1/CP0.2 human squamous carcinoma cells. Because exogenous thymidine, which compensates for 5-FU-mediated inhibition of de novo DNA synthesis via salvage pathway, did not affect this schedule-dependent synergism, the modulatory effect of 5-FU on CDDP resistance would be attributed to the 5-FU-induced RNA damage.

Inhibition of cis-diamminedichloroplatinum (II)-induced DNA interstrand cross-link removal by 7-ethyl-10-hydroxy-camptothecin in HST-1 human squamous-carcinoma cells.

The combination of cis-diamminedichloroplatinum(II) (CDDP) and 7-ethyl-10-[4-(1-piperidino)-1-piperidino]carbonyloxycamptothecin (CPT-11), a topoisomerase-I inhibitor, has been shown to be synergistic in vitro and clinically active against several human cancers refractory to chemotherapy. To elucidate the mechanism of the synergistic cytotoxicity of CDDP and 7-ethyl-10-hydroxycamptothecin (SN-38), an active metabolite of CPT-11, we studied the interaction of these agents using an HST-1 human squamous-carcinoma cell line. Cells were exposed to the IC50 concentration of SN-38 (5.

Direct tumorigenic conversion of human gallbladder carcinoma cells by v-src but not by activated c-H-ras oncogene.

The roles of activated ras and src oncogene products in the acquisition of fully neoplastic phenotype by human gallbladder adenocarcinoma cells were investigated by co-transfecting non-tumorigenic HAG-I human gallbladder carcinoma cells with the pSV2neo plasmid and a plasmid carrying either activated c-H-ras or v-src oncogene. G418-resistant clones were isolated and assessed for the acquisition of anchorage-independent growth potential. Neither the 10 established clones transfected with pSV2neo alone nor the 17 clones transfected with activated c-H-ras, including 4 clones expressing the mutated p21H-ras protein, could form colonies in soft agar.

Sequence-dependent modulation of anticancer drug activities by 7-ethyl-10-hydroxycamptothecin in an HST-1 human squamous carcinoma cell line.

Background: We studied the modulatory effects of 7-ethyl-10-hydroxy-camptothecin (SN-38), an active metabolite of 7-ethyl-10-[4-(1-piperidino)-1-piperidino] carbonyloxycamptothecin, on the antitumor activities of clinically important anticancer drugs including cisplatin (CDDP), 5-fluorouracil (5-FU), mitomycin C (MMC), etoposide (VP-16), and adriamycin (ADR).

Materials And Methods: The HST-1 human carcinoma cells were treated with graded concentrations of these anticancer drugs either alone or in combination with IC50 concentration of SN-38, administered in several different treatment schedules, and antitumor activity was evaluated by the growth inhibition assay.

Results: SN-38 potentiated the antitumor activity of CDDP in all schedules with a maximal effect observed with a simultaneous administration, while SN-38 enhanced the cytotoxicity of MMC, 5-FU, or VP-16 only in certain schedules.

Inhibitory effects of cucurbitane triterpenoids on Epstein-Barr virus activation and two-stage carcinogenesis of skin tumors.

To search for possible anti-tumor-promoters, we carried out a primary screening of 21 cucurbitane triterpenoids using an in vitro assay system. Of these triterpenoids, scandenoside R6 (6), 23,24-dihydrocucurbitacin F (14), 25-acetyl-23,24-dihydrocucurbitacin F (15), 2-O-beta-D-glucopyranosyl-23,24-dihydrocucurbitacin F (17) and cucurbitacin F (18) exhibited significant inhibitory effects on Epstein-Barr virus (EBV) activation induced by the tumor promoter, 12-O-tetradecanoyl-phorbol-13-acetate (TPA). Further, compounds 14 and 17 exhibited remarkable anti-tumor-promotion effects on mouse skin tumor promotion in an in vivo two-stage carcinogenesis test.

Inhibition by 5-fluorouracil of cis-diamminedichloroplatinum(II)-induced DNA interstrand cross-link removal in a HST-1 human squamous carcinoma cell line.

To elucidate the mechanism of the synergistic cytotoxicity of 5-fluorouracil (5-FU) and cis-diamminedichloroplatinum(II) (CDDP), we studied the interaction of these agents using a human squamous carcinoma cell line (HST-1). Exposure to 5-FU for 24 h and to CDDP for 1 h produced a 50% inhibitory concentration of 1.0 micrograms/ml (7.

Identification of new epitopes recognized by human monoclonal antibodies with neutralizing and antibody-dependent cellular cytotoxicity activities specific for human T cell leukemia virus type 1.

We have generated a number of EBV-transformed B cell lines producing human mAb against human T cell leukemia virus type 1 (HTLV-1) from the peripheral blood B lymphocytes obtained from patients with HTLV-1-associated myelopathy/tropical spastic paraparesis. Various synthetic peptides corresponding to antigenic regions of HTLV-1 gag and env proteins were used for the screening of antibodies in ELISA. In our study, four IgG mAb to the gag p19 amino acids 100 to 130, and 5 IgG mAb to the env p46 amino acids 175 to 199 were characterized.

[An autopsy case of citrullinemia type II complicated with chronic pancreatitis].

A 34-year-old woman had developed frequent episodes of disorientation at night since a year ago. Her blood ammonia level was found to be markedly increased. Serum amino acid pattern and biochemical analysis of urea cycle enzymes in the liver indicated type II citrullinemia.