AQP4 and the Fate of Gliomas

Glioma is a relatively common type of tumor in the central nervous system and comprises tumors that differ in malignancy and cellular origin. Glioblastoma multiforme (GBM) is a glioma subtype originating in astrocytes. It is the most common type of brain cancer in adults and is characterized by diffuse and invasive growth, which in combination with high frequency of recurrence make the prognosis very poor (1). Most patients die within 8–18 months of diagnosis and the 2-year survival rate is about 3%. Thus, GBM represents a significant burden on the society and novel therapies are urgently needed.

The study by Simone and colleagues addresses the role of different isoforms of the water channel aquaporin-4 (AQP4) in the pathogenicity of gliomas (2). AQP4 is the predominant brain water channel protein. It is mainly expressed by astrocytes, where it is distributed in a highly polarized manner. The astrocytic foot processes abutting brain microvessels display about 10-fold higher AQP4 density than the astrocyte membrane domains facing the neuropil (3). The dystrophin-associated protein (DAP) complex is responsible for the polarized distribution of AQP4 as it connects intracellular actin cytoskeleton to extracellular matrix molecules present in the perivascular basal lamina. More specifically, the endfoot AQP4 molecules are tethered to the membrane by isoforms of syntrophin, which are PDZ domain–containing members of the DAP complex that interact with the intracellular tail of AQP4 (4).

AQP4 is expressed as two major isoforms that differ in regard to methionine (M) starting codon. The shorter and more abundant form is called M23 and the longer and less abundant form is called M1. The two AQP4 isoforms form heterotetramers, which in turn aggregate into higher order structures called orthogonal arrays of particles (OAP). The two AQP4 isoforms display the same water permeability, with the main established difference between them being their ability to aggregate into OAPs. The shorter M23 isoform forms large OAPs, whereas the longer M1 isoform forms very small or no OAPs (5). The functional significance of the OAPs has not yet been established, but in vitro data indicate that the M23 isoform may function as adhesion molecules, whereas the M1 isoform is mobile in the plasma membrane and is more involved in cell migration (6).

The role of AQP4 isoforms in glioblastoma biology has been addressed in earlier studies, which have shown that high-grade gliomas display higher expression of AQP4 than low-grade tumors (7). Furthermore, OAPs have been shown to be disintegrated or absent in glioblastomas, and more recently an inverse correlation between OAP prevalence and malignancy was demonstrated (8).

The study by Simone and colleagues provides a new perspective on the largely discussed role of AQP4 in brain tumors. They propose a role associated with AQP4 aggregation/disaggregation dynamics into the plasma membrane and their link with the actin cytoskeleton. Transfection of cell lines, commonly used as experimental models of glioma, with the two AQP4 isoforms revealed that M1-AQP4 contributes to the invasiveness of glioma cells, while aggregation in OAPs by M23-AQP4 is deleterious and promotes apoptosis.

The role of M1-AQP4 in migration and invasion has already been suggested, however, the novelty of the study by Simone and colleagues is that they show that the increased invasiveness was because of the increased activity of matrix metalloproteinase-9 (MMP9). High expression of MMP9 has previously been shown to be associated with glioma cell proliferation and patient survival rate (9). Future studies will need to explore the link between M1-AQP4 expression and increased MMP9 activity.

In contrast, glioma cell lines transfected with M23-AQP4, which promotes aggregation of AQP4 into OAPs, shrunk and had an altered morphology. The authors suggested that the shrinkage was associated with apoptosis. They linked apoptosis with cytoskeletal reorganization, as AQP4 aggregation into OAPs affected F-actin dynamics. In addition, they showed that the ratio between F-Actin and G-Actin was significantly increased in the M23-AQP4–transfected glioma cell lines, but not in cell lines transfected with M1-AQP4. A link between AQP4 and F-Actin dynamics has previously been shown by the same laboratory (10). However, this time they provide evidence showing that only the OAP-forming M23-AQP4 affects the cytoskeleton and not the tetramer-forming M1 isoform. Moreover, using point mutagenesis, they identified two prolines (P254 and P296) and the last six amino acids at the C-terminus as the key residues involved in the altered morphology observed in the M23-AQP4–transfected cells. On the basis of this, they postulate that these residues might be involved in the interaction between actin-cytoskeleton and M23-AQP4.

The differential effects of the aggregation state of AQP4 on invasiveness and induction of apoptosis were specific to the glioma cell lines as they were not observed in the non-glioma cell lines. However, these data need to be confirmed using models closer to the in vivo situation.

The authors suggest that because AQP4 aggregation into OAPs is “deleterious” for glioma cell survival, the glioma cell would react by altering the AQP4 aggregation state toward AQP4 tetrameric expression, therefore improving its invasiveness potential. From this perspective, it is not surprising that glioma cell biology is directed toward OAP disassembly into AQP4 tetramers, which allows invasiveness to be achieved while avoiding apoptosis.

In conclusion, the authors speculate that the aggregation state of AQP4 could be a key factor in determining whether glioma cells will die through activation of the apoptotic path or persist by enhancing their invasiveness potential. This study helps to understand the complex role of AQP4 in tumor biology, showing a new perspective in which the role of AQP4 in glioma is not necessarily associated with its water flux capacity and edema formation but with its mysterious role of aggregation into OAPs and its link with the actin cytoskeleton. Furthermore, this study strengthens the potential role of AQP4 as a target in the treatment of glioblastoma; glioblastoma cells could be possibly retransformed so they express the OAP-forming M23-AQP4 isoform that promotes adhesion and apoptosis. Knowledge about the mechanisms by which M23-AQP4 leads to apoptosis in gliomas might contribute to development of novel therapeutic strategies.

No potential conflicts of interest were disclosed.