Identification of GRO1 as a Critical Determinant for Mutant p53 Gain of Function

Abstract

Mutant p53 gain of function contributes to cancer progression, increased invasion and metastasis potentials, and resistance to anticancer therapy. The ability of mutant p53 to acquire its gain of function is shown to correlate with increased expression of progrowth genes, such as c-MYC, MDR1, and NF-B2. However, most of the published studies to identify mutant p53 target genes were performed in a cell system that artificially overexpresses mutant p53. Thus, it remains unclear whether such mutant p53 targets can be regulated by endogenous physiological levels of mutant p53. Here, we utilized SW480 and MIA-PaCa-2 cells, in which endogenous mutant p53 can be inducibly knocked down, to identify mutant p53 target genes that potentially mediate mutant p53 gain of function. We found that knockdown of mutant p53 inhibits GRO1 expression, whereas ectopic expression of mutant R175H in p53-null HCT116 cells increases GRO1 expression. In addition, we found that endogenous mutant p53 is capable of binding to and activating the GRO1 promoter. Interestingly, ectopic expression of GRO1 can rescue the proliferative defect in SW480 and MIAPaCa-2 cells induced by knockdown of mutant p53. Conversely, knockdown of endogenous GRO1 inhibits cell proliferation and thus abrogates mutant p53 gain of function in SW480 cells. Taken together, our findings define a novel mechanism by which mutant p53 acquires its gain of function via transactivating the GRO1 gene in cancer cells. Thus, targeting GRO1 for cancer therapy would be applicable to a large portion of human tumors with mutant p53, but the exploration of GRO1 as a potential target should take the mutation status of p53 into consideration.

Mutant p53 Protein Is Targeted by Arsenic for Degradation and Plays a Role in Arsenic-mediated Growth Suppression

“…To further examine the role of mutant p53 in promoting the survival and proliferation of arsenic-treated cells, a colony formation assay was performed with MIA PaCa-2 and SW480 cells, in which endogenous mutant p53 can be inducibly knocked down (14). Consistent with previous reports (14), cell proliferation was inhibited by knockdown of mutant p53 in both MIA PaCa-2 and SW480 cells (Fig. 8, A and B, left panels).…”

“…Inducible p53 knockdown SW480-p53-KD and MIA PaCa-2-p53-KD cell lines were cultured as described previously (14). To generate cell lines that inducibly express mutant p53(R175H) or p53(R273H), pcDNA4-FLAG-HA-p53(R175H) or pcDNA4-FLAG-HA-p53(R273H) was transfected into HCT116 (p53-null) cells, which express a tetracycline repressor by pcDNA6.…”

“…In our previous studies, we analyzed the requirement of endogenous mutant p53 in the physiological context for cell survival in the SW480 cell line, which contains mutant R273H/P309S, and the MIA PaCa-2 cell line, which contains mutant R248W. We found that mutant p53 is required for cell proliferation and resistance to DNA damage treatment in SW480 and MIA PaCa-2 cells (13,14). Thus, both cell lines along with the HaCaT cell line (containing mutant H179Y/R282W) were used to examine the effect of arsenic compounds on the expression of endogenous mutant p53.…”

Chemokines in cancer related inflammation

“…Mast cells have long been known to drive angiogenesis by sustaining new vessel formation and tumor growth [13]. Other examples are mutation of p53 in tumor cells, able to trans-activate CXCL1 [14] and of Notch1 in T cell-acute lymphocytic leukemia, up-regulating CCR7 expression [15].…”

Antitumor effect of novel small chemical inhibitors of Snail-p53 binding in K-Ras-mutated cancer cells

“…This result implies that mutant p53 may be eliminated by Snail despite of GN chemicals, because it cannot block the interaction between mutant p53 and Snail (Figure 5e). We also checked the effect of GN25 on other pancreatic cancer cell line, MIA-Paca 2, which also possesses wild/ mutant p53 alleles (Yan and Chen, 2009). In this cell line, GN25 suppressed the cell proliferation strongly (Supplementary Figure S5G).…”