tissue samples were homogenized in lysis buffer (50 mM Tris-HCl, pH8.0, 150 mM NaCl, 1 mM EDTA, 1% (v/v) Triton X-100, and Complete Protease Inhibitor Cocktail [Roche]), and cell debris removed by centrifugation. Cleared supernatants were boiled in sodium dodecyl sulphate (SDS) sample buffer and run on an SDS polyacrylamide gel before transfer to a polyvinylidene difluoride (PVDF) membrane (Amersham) and subsequent immunoblotting with either purified rabbit anti-human ARL15 antibody (Proteintech Group) or anti-α-tubulin antibody (sc-8035; Santa Cruz Biotechnology). Full-length human wild type ARL15 cDNA was purchased from Open Biosystems and subcloned into pCDNA 3.1 (Invitrogen) using the XhoI and HindIII restriction sites. HEK293 cells (American Type Culture Collection [ATCC]) were transiently transfected using the CalPhos Mammalian Tranfection Kit (Clontech) according to the manufacturer's instructions.
