Intensive Study On The Phenomenon Of A Specific I?b? Cleavage In Acute Myeloid Leukemia

Acute myeloid leukemia(AML)is a kind of disease characterized by the clonal proliferation of myeloid precursor cells in hematopoietic system.It has been reported that the continuous activation of NF-?B signaling pathway,which is mainly regulated by I?B family proteins,is one of the important causes to maintain the proliferation of AML cells.The activation of the classical NF-?B signaling pathway is mainly achieved by the phosphorylation of I?B? and in ubiquitin proteasome degradation pathway.In cancer cells the abnormal degradation or cleavage of I?B? proteins could often promote or inhibit the entry of NF-?B into the nucleus,leading to the deregulation in activation of downstream genes.Recently,we found an unreported form of I?B? protein cleavage which was detected only in myeloid leukemia cell lines,but not in lymphoid neoplasms and non-hematological malignancies cell lines.When we mixed 293 T cells with AML cells NB4 lysate,we discovered that NB4 cells contained some kind of enzyme that could induce the specific I?B? cleavage.Furthermore,it was found that the phenomenon of I?B? cleavage could be inhibited by serine protease inhibitors.We found that one of the serine proteases PR3 which high expressed in AML cells NB4 with high enzyme activity may be involved in the specific I?B? cleavage.In this study,we observed the effect of PR3 on I?B? cleavage by treating Jurkat cell lysate with a certain amount of exogenous PR3.It was also shown that the wild type PR3 could induce I?B? cleavage in 293 T cells,but a PR3 with a key amino acid mutated in the enzyme active center couldn't do so.In addition,we transfected the PR3 interference plasmid with wild type PR3 plasmid in 293 T cells to observe I?B? cleavage.We found that I?B? cleavage could be inhibited by targeting inhibition of PR3 expression.From the results above,we concluded that PR3 was involved in the specific I?B? cleavage.Then we investigated the effects of PR3 and PR3 mediated I?B? cleavage on AML cells by utilizing Lent-viral vector system and RNA interference technology to inhibit PR3 expression in different types of AML cells(NB4,HL-60,U937).When the expression of PR3 in AML cells was disturbed,the intracellular I?B? cleavage could be inhibited.At the same time we could observe cell growth inhibition or increasing the sensitivity to ATRA and c AMP.Finally,we also detected the expressions of PR3 in primary bone marrow cells from AL patients.The expressions of PR3 in primary AML patients were relatively higher with a significant positive correlation between PR3 and AML prognosis.It seemed that the I?B?cleavage was different in primary AML and primary ALL patients,which may be related to different expressions of PR3 in them.All of these results implied PR3 and PR3 mediated specific I?B? cleavage might function as clinical biomarker for use in diagnosis and prognosis of AML.