L-leucine increases [3H]-thymidine incorporation in chicken hepatocytes: Involvement of the PKC, PI3K/Akt, ERK1/2, and mTOR signaling pathways

• Published in: Journal of cellular biochemistry 2008-12, Vol.105 (6), p.1410-1419
• Main Authors: Lee, Min Young, Jo, Sung Duk, Lee, Jang Hern, Han, Ho Jae
• Format: Article
• Language: English
• Subjects: [3H]-thymidine incorporation; Animals; Cell Cycle; Cell Proliferation; chicken hepatocytes; Chickens; Hepatocytes - enzymology; Hepatocytes - metabolism; L-leucine; Leucine - pharmacology; MAP Kinase Signaling System; Mitogen-Activated Protein Kinase 1 - metabolism; Mitogen-Activated Protein Kinase 3 - metabolism; Phosphatidylinositol 3-Kinases - metabolism; Protein Kinase C - metabolism; Protein Kinases - metabolism; Proto-Oncogene Proteins c-akt - metabolism; Thymidine - metabolism; TOR Serine-Threonine Kinases
• ISSN: 0730-2312; 1097-4644
• DOI: 10.1002/jcb.21959

This study examined how L‐leucine affected DNA synthesis and cell cycle regulatory protein expression in cultured primary chicken hepatocytes. L‐Leucine promoted DNA synthesis in a dose‐ and time‐dependent manner, with concomitant increases in cyclin D1 and cyclin E expression. Phospholipase C (PLC) and protein kinase C (PKC) mediated the L‐leucine‐induced increases in [3H]‐thymidine incorporation and cyclin D1/CDK4 and cyclin E/CDK2 expression, as U73122 (a PLC inhibitor) or bisindolylmaleimide I (a PKC blocker) inhibited these effects. L‐Leucine also increased PKC phosphorylation and intracellular Ca2+ levels. L‐Leucine‐mediated increases in [3H]‐thymidine incorporation and cyclin/CDK expression were sensitive to LY 294002 (PI3K inhibitor), Akt inhibitor, PD 98059 (MEK inhibitor). It was also observed that L‐leucine‐induced increases of cyclin/CDK expression were inhibited by PI3K siRNA and ERK siRNA; L‐leucine increased extracellular signal‐regulated kinases 1/2 (ERK1/2) and Akt phosphorylation levels. Bisindolylmaleimide I attenuated L‐leucine‐induced phosphorylation of ERK1/2 but did not influence Akt phosphorylation, and PI3K siRNA and LY 294002 inhibited L‐leucine‐induced ERK1/2 phosphorylation, suggesting some cross‐talk between the PKC and ERK1/2 or PI3K/Akt and ERK1/2 pathways. L‐Leucine also increased the levels of phosphorylated molecular target of rapamycin (mTOR) and two of its targets, ribosomal protein S6 kinase (p70S6K), and 4E binding protein 1 (4E‐BP1); furthermore, rapamycin (an mTOR inhibitor) blocked all of the mitogenic effects of L‐leucine. In addition, Akt inhibitor blocked L‐leucine‐induced mTOR phosphorylation. In conclusion, L‐leucine stimulated DNA synthesis and promoted cell cycle progression in primary cultured chicken hepatocytes through PKC, ERK1/2, PI3K/Akt, and mTOR. J. Cell. Biochem. 105: 1410–1419, 2008. © 2008 Wiley‐Liss, Inc.