IDENTIFICATION OF THE PHOSPHATIDIC-ACID PHOSPHATASE TYPE 2A ISOZYME AS AN ANDROGEN-REGULATED GENE IN THE HUMAN PROSTATIC ADENOCARCINOMA CELL-LINE LNCAP

Differential display wars used to identify novel and androgen-regulate d genes in the human prostatic adenocarcinoma cell line LNCaP. A 322-b ase pair cDNA fragment that, was consistently induced by the synthetic androgen R1881 revealed 100% homology with the human phosphatidic aci d phosphatase type 2a isozyme very recently reported by Kai Et al; (PA P-2a; Kai., Wa., Wada, I., Imai, S.-i., Sakane, F., and Kanoh, H. (199 7) J. Biol. Chem. 272, 24572-24578). The fragment was used to clone th e corresponding cDNA from a human prostate library. The deduced amino acid sequence confirmed the identity with human PAP-2a. The inducibili ty of PAP-2a mRNA by androgens was confirmed by Northern blot hybridiz ation, The effect was time-and dose-dependent Math a maximal stimulati on (4-fold) after 24 h of treatment with 10(-9) M R1881. The steroid s pecificity of PAP-2a mRNA regulation was found to be in agreement with the aberrant ligand specificity of the mutated androgen receptor in L NCaP cells, supporting the involvement of the androgen receptor in the induction process. Furthermore, low basal levels of PAP-2a mRNA and a bsence of androgen inducibility were observed in the poorly differenti ated and androgen receptor-negative cell lines PC-3 and DU-145. Induct ion of PaP-2a mRNA was not affected by the protein synthesis inhibitor cycloheximide and was accompanied by a marked increase in PAP-2 activ ity as measured by the conversion of phosphatidic acid into diacylglyc erol in membrane fractions of LNCaP. Comparison of the expression of P AP-2a mRNA in 50 different human tissues revealed ubiquitous expressio n. The highest levels, however, were observed in the prostate, Since P AP-2 plays a pivotal role in the control of signal transduction by lip id mediators such as phosphatidate, lysophosphatidate, and ceramide-1- phosphate, the ability off androgens to stimulate the expression and a ctivity of this enzyme in prostatic cells may provide an important opp ortunity for cross-talk between signaling pathways involving lipid med iators and androgens.