REGULATION OF HYPOXANTHINE PHOSPHORIBOSYLTRANSFERASE, GLYCERALDEHYDE-3-PHOSPHATE DEHYDROGENASE AND BETA-ACTIN MESSENGER-RNA EXPRESSION IN PORCINE IMMUNE CELLS AND TISSUES

Various ''housekeeping'' genes are often used as endogenous controls i n gene expression experiments. We have cloned from swine, three genes commonly used as endogenous controls in other species and have charact erized their relative levels of expression in various porcine tissues and their response to various cell activators. Glyceraldehyde-3-phosph ate dehydrogenase (GAPDH) and beta-actin were readily detected by nort hern hybridization in various tissues and in alveolar macrophages. The expression of hypoxanthine phosphoribosyltransferase (HPRT) was detec ted only by northern hybridization of poly-A(+) enriched RNA and by re verse transcriptase-polymerase chain reaction (RT-PCR), making it more suitable for highly sensitive detection methods. Expression of GAPDH varied less among tissues than did beta-actin, making it more useful c ontrol for comparisons of gene expression between tissues with norther n hybridizations. Various treatments of cultured alveolar macrophages differentially affected levels of beta-actin and GAPDH, while HPRT exp ression was unchanged in alveolar macrophages or spleen cells similarl y treated. Therefore, while HPRT can be used as the endogenous control with sensitive detection methods such as RT-PCR, less sensitive detec tion methods require a more abundant gene such as GAPDH.