| Glucocorticoid Receptor (GR) in the biological evolution, is the most conservative transcription factor of initiation nucleus and vital to maintain the activities of life. It plays an important role in the process of trauma and diseases development and progress. It has been testified that mice knocking-out the GR gene are non-viable, indicating that GR is the vital protein mediating many essential physiological functions.The previous work in our lab have confirmed that for a paitient being severe scalded and burnt GR has the following features at the earlier stage: GR reduces in the many organs and tissues. The expression of GR cuts down. Especially the transfer of GR from cytoplasm to nucleuses reduces so as to decrease the transcriptional activation of GR. Meanwhile the cell factors joining in the systemic inflammatory response, such as TNF- # JL-1 * ,IL-6, rise remarkably. It has the cause and effect relationship between the decreasing GR and the high cell factors blood syndrome, which is the effect of progressional magnification. Our analysis suggests the decreasing of GR is the main reason why the anti-inflammation mechanism has been weakened in the body, and is closely related to the Systemic Inflammatory Response Syndrome (SIRS). According to the changeable features of GR, controling the expression of GR and the functional state correspondingly will possibly reduce and prevent the over imflammation response and continuous damage of the whole body in the initial stages. Therefore, utilizing Glucocorticoid Receptor-Ligand Binding Domain (GR-LBD) as a bait, our task groups have screened 5 candidate proteins which interact with GR-LBD from a human marrow cDNA library by the yeast two-hybrid system. Then, using Glutathione S-Transferase (GST) Pull Down and Coimmunoprecipitation (Co-IP) analysis, it has been proved that interferon-inducible protein P56 can interact with GR-LBD in vitro or vivo. P56 can interact with GR by the way of ligand undependence, inhibits the GR-mediated transcriptional activity and is the corepressorof GR. The synthesis of P56 increases after the excess systemic inflammation response and continuous damage. The.additional P56 interacts with GR-LBD so that the quantity of GR is decreased and the activity of GR is weakened. Body is at the state of the high cell factors and the over systemic imflarnmation response occurs.Interferon-inducible protein P56 was discovered in the seventies last century. There has been little report on the interaction of P56 and GR, as a result its function and significance are unknown. In order to search the less fragment of P56 interacting with GR-LBD, GR is controlled with the fragment and the over systemic imflammation response is weakened. On the basis of our previous work, we have thoroughly analysed the interaction domains between P56 and GR with the yeast two-hybrid system. Finally, the specific interaction domain is found. The expressive plasmid of P56 is constructed and the recombined whole length P56 is induced and thus results hi preliminary expression. Their main results and conclusions read as follows:1 % Successful construction of the. yeast expressive plasmids for the fragments of varied deleted mutants of P56IFN was used to induce HT1080 cell, so as to abstract mRNA and reverse-transcribe cDNA of P56.The different primers of P56 were designed, the whole length and a series of deletion mutants of P56 were obtained through PCR amplification. The products of amplification were inserted into T-Vector. The positive clone was chosen and verifyied by restricted endonuclease. The fragments of whole lengtlrand a series of deletion mutants of P56 were each cloned into the yeast expression plasmid pGADT7 and named pGADT7-A P56. The sequential analysis revealed that all sequences were correct and the inserted reading frame was also correct.2 The identification of the special interacting domains between P56 and GR the recombined plasmids of a series of deletion mutants of P56 and pGBK7-GRLBD plasmid were transferred into the yeast cell in pairs. With...
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