Humphries Michael J, Limesand Kirsten H, Schneider Jonathan C, Nakayama Keiichi I, Anderson Steven M, Reyland Mary E
Department of Craniofacial Biology and School of Dentistry, University of Colorado Health Sciences Center, Aurora, Colorado 80262, USA.
J Biol Chem. 2006 Apr 7;281(14):9728-37. doi: 10.1074/jbc.M507851200. Epub 2006 Feb 1.
Protein kinase C (PKC) delta is an essential regulator of mitochondrial dependent apoptosis in epithelial cells. We have used the PKCdelta(-/-) mouse to ask if loss of PKCdelta protects salivary glands against gamma-irradiation-induced apoptosis in vivo and to explore the mechanism underlying protection from apoptosis. We show that gamma-irradiation in vivo results in a robust induction of apoptosis in the parotid glands of wild type mice, whereas apoptosis is suppressed by greater than 60% in the parotid glands of PKCdelta(-/-) mice. Primary parotid cells from PKCdelta(-/-) mice are defective in mitochondrial dependent apoptosis as indicated by suppression of etoposide-induced cytochrome c release, poly(ADP-ribose) polymerase cleavage, and caspase-3 activation. Notably, apoptotic responsiveness can be restored by re-introduction of PKCdelta by adenoviral transduction. Etoposide and gamma-irradiation-induced activation of p53 is similar in primary parotid cells and parotid glands from PKCdelta(+/+) and PKCdelta(-/-) mice, indicating that PKCdelta functions downstream of the DNA damage response. In contrast, activation of the c-Jun amino-terminal kinase is reduced in primary parotid cells from PKCdelta(-/-) cells and in parotid C5 cells, which express a dominant inhibitory mutant of PKCdelta. Similarly, c-Jun amino-terminal kinase activation is suppressed in vivo in gamma-irradiated parotid glands from PKCdelta(-/-) mice. These studies indicate an essential role for PKCdelta downstream of the p53 response and upstream of the c-Jun amino-terminal kinase activation in DNA damage-induced apoptosis in vivo and in vitro.
蛋白激酶C（PKC）δ是上皮细胞中线粒体依赖性凋亡的关键调节因子。我们利用PKCδ基因敲除小鼠来探究PKCδ缺失是否能在体内保护唾液腺免受γ射线诱导的凋亡，并探索其抗凋亡的潜在机制。我们发现，体内γ射线照射可在野生型小鼠的腮腺中强烈诱导凋亡，而在PKCδ基因敲除小鼠的腮腺中，凋亡受到超过60%的抑制。PKCδ基因敲除小鼠的原代腮腺细胞在线粒体依赖性凋亡方面存在缺陷，这表现为依托泊苷诱导的细胞色素c释放、聚（ADP - 核糖）聚合酶裂解以及半胱天冬酶 - 3激活均受到抑制。值得注意的是，通过腺病毒转导重新引入PKCδ可恢复凋亡反应性。在PKCδ基因野生型和敲除型小鼠的原代腮腺细胞及腮腺中，依托泊苷和γ射线诱导的p53激活相似，这表明PKCδ在DNA损伤反应的下游发挥作用。相反，在PKCδ基因敲除的原代腮腺细胞以及表达PKCδ显性抑制突变体的腮腺C5细胞中，c - Jun氨基末端激酶的激活减少。同样，在PKCδ基因敲除小鼠经γ射线照射的腮腺中，体内c - Jun氨基末端激酶的激活也受到抑制。这些研究表明，PKCδ在体内外DNA损伤诱导的凋亡中，在p53反应的下游以及c - Jun氨基末端激酶激活的上游发挥着重要作用。
