A Cancer-associated Aurora A Mutant Is Mislocalized and Misregulated Due to Loss of Interaction with TPX2

Abstract

Mutations in protein kinases can drive cancer through alterations of the kinase activity or by uncoupling kinase activity from regulation. Changes to protein expression in Aurora A, a mitotic Ser/Thr kinase, are associated with the development of several human cancers, but the effects of somatic cancer-associated mutations have not been determined. In this study we show that Aurora A kinase activity is altered in different ways in three somatic cancer-associated mutations located within the catalytic domain; Aurora A(V174M) shows constitutively increased kinase activity, Aurora A(S155R) activity is decreased primarily due to misregulation, and Aurora A(S361*) activity is ablated due to loss of structural integrity. These alterations suggest vastly different mechanisms for the role of these three mutations in human cancer. We have further characterized the Aurora A(S155R) mutant protein, found that its reduced cellular activity and mislocalization are due to loss of interaction with TPX2, and deciphered the structural basis of the disruption at 2.5 Å resolution. Previous studies have shown that disruption of the Aurora A/TPX2 interaction results in defective spindles that generate chromosomal abnormalities. In a panel of 40 samples from microsatellite instability-positive colon cancer patients, we found one example in which the tumor contained only Aurora A(S155R), whereas the normal tissue contained only wild-type Aurora A. We propose that the S155R mutation is an example of a somatic mutation associated with this tumor type, albeit at modest frequency, that could promote aneuploidy through the loss of regulated interactions between Aurora A and its protein partners.Aurora A is a centrosome and mitotic spindle-associated, cell cycle-regulated serine/threonine kinase and is a key regulator of mitosis (1-5). The protein levels and the activity of Aurora A peak at G 2 and during mitosis, whereas its expression is low in resting cells (1, 6). The Aurora A gene is located on human chromosome 20q13, a region that has been found to be amplified in a variety of human tumors (7). Overexpression of Aurora A induces abnormal spindle formation, leading to prolonged mitosis and polyploidy (8). Aurora A was first described in human cancer cell lines but has subsequently been found to be overexpressed in a broad range of human tumors, including primary colorectal carcinoma, gliomas, breast, ovarian, and pancreatic cancers (1, 9 -12).The association of Aurora A with cancer has prompted many studies that have revealed details of the function and regulation of the kinase. Bona fide substrates that are regulated by Aurora A phosphorylation include TACC3, Plk1, Eg5, and p53 (13-22). The catalytic activity of Aurora A is activated by phosphorylation on Thr-288 in its activation loop and by interaction with partner proteins such as TPX2, Ajuba, and HEF1 (13, 23-27). Aurora A is deactivated by dephosphorylation of Thr-288 by protein phosphatase 1 (PP1), 4 which is prevented by TPX2 (24, 25, 27). Aurora A and TPX2 are both degraded by ...

TPX2andAURKApromote 20q amplicon-driven colorectal adenoma to carcinoma progression

“…These results are consistent with features expected from genes that drive gain of the amplicon. Somatic mutations in AURKA and TPX2 have been reported to occur at low frequency in CRC, which supports their putative oncogenic potential 21 23…”

On the molecular mechanisms of mitotic kinase activation

“…When bound to MLN8054, the DFG motif adopts a flipped DFG-up conformation distinct from DFG-out, in which Trp277 packs against Val174 and occupies a very similar position to that observed for Val600 in the BRAF structure, suggesting that this structure might represent an autoinhibitory state. Consistent with this idea, mutation of Val174 to methionine increases kinase activity and has been identified in melanoma as a potential cancer driver mutation [81]. …”

Aurora kinase inhibitors as anticancer molecules

“…The above results predict that MLN8054 can target VX-680/MK-0457 resistant cancer cells, and Aurora-A may develop mutations at sites different from those of Aurora-B to acquire resistance against the inhibitors. In fact, multiple cancer associated mutations in Aurora-A have been identified, of which, some alter the kinase activity and others create or abolish protein interaction (110–112). As mentioned earlier, I31F polymorphism in Aurora-A can activate NF-κB pathway through preferential UBE2N interaction to down-regulate I B (28,113).…”

“…The I31F and V57I haplotype is associated with increase in esophageal cancer risk (111). V174M mutant Aurora-A is constitutively active due to stabilization of the activation loop whereas S155R mutant of Aurora-A looses interaction with TPX2 resulting in reduced kinase activity as well as mis-localization on mitotic spindle (112). Moreover, overexpression of the kinase inactivated allele of Aurora-A does not interfere with cell proliferation unless its kinase activity is completely lost (114).…”