Role for proteasome activator PA200 and postglutamyl proteasome activity in genomic stability

Proteasome activator PA200 enhances proteasome-mediated cleavage after acidic residues in vitro; however, its role within cells is not known. Here, we show that, in response to ionizing radiation, PA200 forms hybrid proteasomes with 19S caps and 20S core proteasomes that accumulate on chromatin, leading to an increase in proteolytic activity. Unlike many other proteins that respond to DNA damage, the response of PA200 appears to be independent of Ataxia Telangiectasia Mutated and p53, but dependent on DNA-dependent protein kinase activity. Nonetheless, PA200 is critical because PA200-knockdown cells show genomic instability and reduced survival after exposure to ionizing radiation. This phenotype is reproduced by specific inhibition of postglutamyl activity of proteasomes, but combined treatment with PA200 siRNA and postglutamyl inhibitor does not show additive effects on survival. Together, these data suggest a unique role for PA200 in genomic stability that is likely mediated through its ability to enhance postglutamyl cleavage by proteasomes

Distinct molecular mechanisms responsible for bortezomib-induced death of therapy-resistant versus -sensitive B-NHL cells

Resistance to currently available therapies is a major impediment to the successful treatment of hematological malignancies. Here, we used a model of therapy-resistant B-cell nonHodgkin lymphoma (B-NHL) developed in our laboratory along with primary B-NHL cells to study basic mechanisms of bortezomib activity. In resistant cells and a subset of primary B-NHLs, bortezomib treatment led to stabilization of Bak and subsequent Bak-dependent activation of apoptosis. In contrast to sensitive cells that die strictly by apoptosis, bortezomib was capable of killing resistant cells through activation of apoptosis or caspase-independent mechanism(s) when caspases were pharmacologically inhibited. Our data demonstrate that bortezomib is capable of killing B-NHL cells via multiple mechanisms, regardless of their basal apoptotic potential, and contributes to growing evidence that proteasome inhibitors can act via modulation of B-cell lymphoma 2 (Bcl-2) family proteins. The capacity of bortezomib to act independently of the intrinsic apoptotic threshold of a given B-NHL cell suggests that bortezomib-based therapies could potentially overcome resistance and result in relevant clinical activity in a relapsed/refractory setting