Distinctive roles of PHAP proteins and prothymosin-alpha in a death regulatory pathway

A small molecule, alpha-(trichloromethyl)-4-pyridineethanol (PETCM), was identified by high-throughput screening as an activator of Caspase-3 in extracts of a panel of Cancer cells. PETCM was used in combination with biochemical fractionation to identify a pathway that regulates mitochondria-initiated Caspase activation. This pathway consists of tumor suppressor putative HLA-DR-associated proteins (PHAP) and oncoprotein prothymosin-alpha (ProT). PHAP proteins promoted caspase-9 activation after apoptosome formation, whereas ProT negatively regulated caspase-9 activation by inhibiting apoptosome formation. PETCM relieved ProT inhibition and allowed apoptosome formation at a physiological concentration of deoxyadenosine triphosphate. Elimination of ProT expression by RNA interference sensitized cells to ultraviolet irradiation-induced Apoptosis and negated the requirement of PETCM for Caspase activation. Thus, this chemical-biological combinatory approach has revealed the regulatory roles of oncoprotein ProT and tumor suppressor PHAP in Apoptosis.