The overall objective of the work proposed in this application and accomplished during the 39 years the grant has been active is to define and understand the mechanisms regulating the growth of gastrointestinal mucosa. This objective is unaltered in the current proposal, which focuses on elucidating the mechanism by which polyamines are required for apoptosis of intestinal epithelial cells. During the past period of support we determined that polyamines are essential to apoptosis in whole animals in the same manner as in cultured IEC-6 cells;that PP2A is inactivated following polyamine depletion and that the accounts for much of the antiapoptotic response;that Src-kinase is activated and plays an initial and central role in the antiapoptotic cascade;and that Src activates STAT3 as part of the response. During the current period of support we plan to determine the mechanism by which polyamine depletion inactivates JNK, which is the major proapoptotic mediator in intestinal epithelial cells. We have shown that ERK is constitutively activated by polyamine depletion due to the inactivation of PP2A and that JNK inactivation depends on ERK. We are hypothesizing that JNK inactivation is, therefore, dependent on PP2A inactivation. We will test this in the first specific aim and also examine the mechanisms involved in ERK activation.
In specific aim 2, we will determine the mechanism for the activation of Src in response to polyamine depletion and its relationship to integrin pS and the EGFR. We have already identified a polyamine binding site on Src. P53 is the major determinant of apoptosis and/or cell cycle arrest following DNA damage, and in the final specific aim, we will determine why DNA damage causes apoptosis in control intestinal epithelial cells, but cell cycle arrest in polyamine depleted cells.
This aim i s based on our findings that the levels of proteins whose transcription is regulated by p53 are all affected by polyamines. The finding that polyamines are required for apoptosis is new and of great potential clinical significance, since it suggests a way to protect normal cells during cancer therapy. Understanding the mechanisms involved in that protection will increase our knowledge of cancer, mucosal healing, inflammation and any other condition in which Gl mucosal growth is altered or involved.
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