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首页> 外文期刊>Molecular and Cellular Biology >Clonal variants of PC12 pheochromocytoma cells with defects in cAMP-dependent protein kinases induce ornithine decarboxylase in response to nerve growth factor but not to adenosine agonists.
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Clonal variants of PC12 pheochromocytoma cells with defects in cAMP-dependent protein kinases induce ornithine decarboxylase in response to nerve growth factor but not to adenosine agonists.

机译：在cAMP依赖的蛋白激酶中有缺陷的PC12嗜铬细胞瘤细胞的克隆变体诱导鸟氨酸脱羧酶响应神经生长因子而不是腺苷激动剂。

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We have isolated and partially characterized three mutants of the pheochromocytoma line PC12 by using dibutyryl cyclic AMP (cAMP) as a selective agent. Each of these variants, A126-1B2, A208-4, and A208-7, was resistant to both dibutyryl cAMP and cholera toxin when cell growth was measured. In comparison to wild-type PC12 cells, each of these mutants was deficient in the ability to induce ornithine decarboxylase (ODC) in response to agents that act via a cAMP-dependent pathway. In contrast, each of these mutants induced ODC in response to nerve growth factor. To understand the nature of the mutations, the cAMP-dependent protein kinases of the wild type and of each of these mutants were studied by measuring both histone kinase activity and 8-N3-[32P]cAMP labeling. Wild-type PC12 cells contained both cAMP-dependent protein kinase type I (cAMP-PKI) and cAMP-dependent protein kinase type II (cAMP-PKII). Regulatory subunits were detected in both soluble and particulate fractions. The mutant A126-1B2 contained near wild-type PC12 levels of cAMP-PKI but greatly reduced levels of cAMP-PKII. Furthermore, when compared with wild-type PC12 cells, this cell line had an altered distribution in ion-exchange chromatography of regulatory subunits of cAMP-PKI and cAMP-PKII. The mutant A208-4 demonstrated wild-type-level binding of 8-N3-[32P]cAMP to both type I and type II regulatory subunits, but only half the wild-type level of type II catalytic activity. The mutant A208-7 had type I and type II catalytic activities equivalent to those in wild-type cells. However, the regulatory subunit of cAMP-PKI occurring in A208-7 demonstrated decreased levels of binding 8-N3-[32P]cAMP in comparison with the wild type. Furthermore, all mutants were defective in their abilities to bind 8-N3-[32P]cAMP to the type II regulatory protein in the particulate fraction. Thus, cAMP-PK was altered in each of these mutants. We conclude that both cAMP-PKI and cAMP-PKII are apparently required to induce ODC in response to increases in cAMP. Finally, since all three mutants induced ODC in response to nerve growth factor, the nerve growth factor-dependent induction of OCD was not mediated by an increase in cAMP that led to an activation of cAMP-PK. These mutants will be useful in the elucidation of the many functions controlled by cAMP and nerve growth factor.
机译：我们已经通过使用二丁酰环AMP（cAMP）作为选择剂分离并部分表征了嗜铬细胞瘤品系PC12的三个突变体。当测量细胞生长时，这些变体A126-1B2，A208-4和A208-7中的每一个均对二丁酰cAMP和霍乱毒素具有抗性。与野生型PC12细胞相比，这些突变体中的每一个都缺乏对鸟氨酸脱羧酶（ODC）的反应能力，而这种作用是通过cAMP依赖性途径起作用的。相反，这些突变体中的每一个都响应神经生长因子而诱导ODC。为了了解突变的性质，通过测量组蛋白激酶活性和8-N3- [32P] cAMP标记，研究了野生型和每种突变体的cAMP依赖性蛋白激酶。野生型PC12细胞同时包含cAMP依赖性蛋白激酶I型（cAMP-PKI）和cAMP依赖性蛋白激酶II型（cAMP-PKII）。在可溶性和颗粒级分中均检测到调节亚基。突变体A126-1B2包含接近野生型PC12水平的cAMP-PKI，但大大降低了cAMP-PKII的水平。此外，当与野生型PC12细胞相比时，该细胞系在cAMP-PKI和cAMP-PKII调节亚基的离子交换层析中具有改变的分布。突变体A208-4展示了8-N3- [32P] cAMP与I型和II型调节亚基的野生型水平结合，但只有II型催化活性的野生型水平结合。突变体A208-7具有与野生型细胞相同的I型和II型催化活性。但是，与野生型相比，在A208-7中出现的cAMP-PKI调节亚基显示出降低的结合8-N3- [32P] cAMP水平。此外，所有突变体在将8-N3- [32P] cAMP与颗粒级分中的II型调节蛋白结合方面均存在缺陷。因此，在这些突变体的每一个中，cAMP-PK均发生了改变。我们得出的结论是，显然cAMP-PKI和cAMP-PKII都需要诱导ODC以响应cAMP的增加。最后，由于所有三个突变体均响应神经生长因子而诱导了ODC，因此，神经生长因子依赖性的OCD诱导并不由cAMP的增加介导，而cAMP的增加导致cAMP-PK的激活。这些突变体将有助于阐明由cAMP和神经生长因子控制的许多功能。

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