The expression pattern of dormancy-associated genes in multiple life-history stages in the rotifer Brachionus plicatilis

Rotifer resting eggs retain their viability for several decades in a non-desiccated form and are of interest in discerning the processes associated with dormancy, since in most organisms this phenomenon is linked with desiccation. The expression pattern of candidate genes with biological functions associated with dormancy in several other organisms was examined in rotifers. High-throughput transcriptome profiling revealed three patterns of gene expression in resting eggs: (1) relatively highly expressed genes coding for LEA proteins and putative paralogs of the small heat shock protein family (shsp); (2) genes coding for ferritin (ferr), glutathione-6-transferase (gts) and HSP70, where some of the putative gene paralogs of these families show relatively high expression levels and other putative paralogs show relatively low expression levels in resting eggs; and (3) genes with relatively low expression levels in resting eggs, for trehalose-6-phosphate synthase (tps), fatty-acid binding proteins (fab) and of lipoprotein lipase (lpl) and the aquaporins gene family (aqp). Changes in the expression pattern of some members of putative gene families occurred during the obligatory dormant period of resting eggs. A transition was observed from an expression pattern of diapausing embryos to an expression pattern of amictic females, during hatching. Differences were also found in the expression pattern in the different types of females, especially in those carrying resting eggs, and in males compared with females. These results suggest putative functional significance to genes associated with dormancy in non-desiccated resting eggs. It could also be proposed that their occurrence in resting eggs is developmentally programmed to facilitate survival in case of desiccation

BMC Genomics

BACKGROUND Microscopic monogonont rotifers, including the euryhaline species Brachionus plicatilis, are typically found in water bodies where environmental factors restrict population growth to short periods lasting days or months. The survival of the population is ensured via the production of resting eggs that show a remarkable tolerance to unfavorable conditions and remain viable for decades. The aim of this study was to generate Expressed Sequence Tags (ESTs) for molecular characterisation of processes associated with the formation of resting eggs, their survival during dormancy and hatching. RESULTS Four normalized and four subtractive libraries were constructed to provide a resource for rotifer transcriptomics associated with resting-egg formation, storage and hatching. A total of 47,926 sequences were assembled into 18,000 putative transcripts and analyzed using both Blast and GO annotation. About 28-55% (depending on the library) of the clones produced significant matches against the Swissprot and Trembl databases. Genes known to be associated with desiccation tolerance during dormancy in other organisms were identified in the EST libraries. These included genes associated with antioxidant activity, low molecular weight heat shock proteins and Late Embryonic Abundant (LEA) proteins. Real-time PCR confirmed that LEA transcripts, small heat-shock proteins and some antioxidant genes were upregulated in resting eggs, therefore suggesting that desiccation tolerance is a characteristic feature of resting eggs even though they do not necessarily fully desiccate during dormancy. The role of trehalose in resting-egg formation and survival remains unclear since there was no significant difference between resting-egg producing females and amictic females in the expression of the tps-1 gene. In view of the absence of vitellogenin transcripts, matches to lipoprotein lipase proteins suggest that, similar to the situation in dipterans, these proteins may serve as the yolk proteins in rotifers. CONCLUSION: The 47,926 ESTs expand significantly the current sequence resource of B. plicatilis. It describes, for the first time, genes putatively associated with resting eggs and will serve as a database for future global expression experiments, particularly for the further identification of dormancy related genes

Induction of plasminogen activator inhibitor type-1 (PAI-1) by hypoxia and irradiation in human head and neck carcinoma cell lines

Contains fulltext : 53187.pdf ( ) (Open Access)BACKGROUND: Squamous cell carcinoma of the head and neck (SCCHN) often contain highly radioresistant hypoxic regions, nonetheless, radiotherapy is a common treatment modality for these tumours. Reoxygenation during fractionated radiotherapy is desired to render these hypoxic tumour regions more radiosensitive. Hypoxia additionally leads to up-regulation of PAI-1, a protein involved in tumour progression and an established prognostic marker for poor outcome. However, the impact of reoxygenation and radiation on PAI-1 levels is not yet clear. Therefore, we investigated the kinetics of PAI-1 expression and secretion after hypoxia and reoxygenation, and determined the influence of ionizing radiation on PAI-1 levels in the two human SCCHN cell lines, BHY and FaDu. METHODS: HIF-1alpha immunoblot was used to visualize the degree of hypoxia in the two cell lines. Cellular PAI-1 expression was investigated by immunofluorescence microscopy. ELISA was used to quantify relative changes in PAI-1 expression (cell lysates) and secretion (cell culture supernatants) in response to various lengths (2-4 h) of hypoxic exposure (< 0.66% O2), reoxygenation (24 h, 20% O2), and radiation (0, 2, 5 and 10 Gy). RESULTS: HIF-1alpha expression was induced between 2 and 24 h of hypoxic exposure. Intracellular PAI-1 expression was significantly increased in BHY and FaDu cells as early as 4 h after hypoxic exposure. A significant induction in secreted PAI-1 was seen after 12 to 24 h (BHY) and 8 to 24 h (FaDu) hypoxia, as compared to the normoxic control. A 24 h reoxygenation period caused significantly less PAI-1 secretion than a 24 h hypoxia period in FaDu cells. Irradiation led to an up-regulation of PAI-1 expression and secretion in both, BHY and FaDu cells. CONCLUSION: Our data suggest that both, short-term (approximately 4-8 h) and long-term (approximately 20-24 h) hypoxic exposure could increase PAI-1 levels in SCCHN in vivo. Importantly, radiation itself could lead to PAI-1 up-regulation in head and neck tumours, whereas reoxygenation of hypoxic tumour cells during fractionated radiotherapy could counteract the increased PAI-1 levels

Specific binding of TES-23 antibody to tumour vascular endothelium in mice, rats and human cancer tissue: a novel drug carrier for cancer targeting therapy

The tissue distribution of anti-tumour vascular endothelium monoclonal antibody (TES-23) produced by immunizing with plasma membrane vesicles from isolated rat tumour-derived endothelial cells (TECs) was assessed in various tumour-bearing animals. Radiolabelled TES-23 dramatically accumulated in KMT-17 fibrosarcoma, the source of isolated TECs after intravenous injection. In Meth-A fibrosarcoma, Colon-26 adenocarcinoma in BALB/c mice and HT-1080 human tumour tissue in nude mice, radioactivities of 125I-labelled TES-23 were also up to 50 times higher than those of control antibody with little distribution to normal tissues. The selective recognition of TES-23 to TECs was competitively blocked by preadministration of unlabelled TES-23 in vivo. Furthermore, immunostaining of human tissue sections showed specific binding of TES-23 on endothelium in oesophagus cancers. These results indicate that tumour vascular endothelial cells express common antigen in different tumour types of various animal species. In order to clarify the efficacy of TES-23 as a drug carrier, an immunoconjugate, composed of TES-23 and neocarzinostatin, was tested for its anti-tumour effect in rats bearing KMT-17 fibrosarcomas. The immunoconjugate (TES-23-NCS) caused marked regression of the tumour, accompanied by haemorrhagic necrosis. Thus, from a clinical view, TES-23 would be a novel drug carrier because of its high specificity to tumour vascular endothelium and its application to many types of cancer. © 1999 Cancer Research Campaig