Role of DAKAPs in Mitochondrial Function

关于线粒体的第二信使信号相对知之甚少。然而，近年来，已经在该细胞器中鉴定出许多底物以cAMP依赖性方式磷酸化。这些底物已在该细胞器中被鉴定为以cAMP依赖性方式磷酸化。这些底物包括电子传递链组分和促凋亡蛋白BAD，表明线粒体中cAMP依赖性蛋白激酶（PKA）的生理作用。在泰勒实验室进行的实验已经确定了两个新的AKAP（D= AKAP）定位于线粒体。D-AKAP 1在NH 2末端含有30个氨基酸，足以靶向线粒体外膜，而初步研究表明D-AKAP 2定位于基质。有趣的是，D-AKAP 2还含有两个推定的PGS结构域，即能够结合Ga亚基的位点，并导致推测D-AKAP 2可能是能够共定位多种信号传导分子的支架蛋白。我们推测PKA在改变特定线粒体功能中的作用是通过与这些新的锚定蛋白的结合来介导的。为了验证这一假设，本研究提出了确定PKA磷酸化的底物，作为与AKAP相关的结果。由于RI和RII亚型之间的生物化学差异，PKA对线粒体功能的相对影响可能是其亚型与锚定蛋白结合的功能。因此，本研究将寻求确定RI与RII亚型锚定在线粒体中的相对意义。最后，将进行实验以确定除了PKA之外，哪些蛋白质与线粒体中的D-AKAP 2结合。

Relatively little is known about second messenger signaling to mitochondria. In recent years though, a number of substrates have been identified in this organelle that are phosphorylated in a cAMP-dependent manner. These substrates have been identified in this organelle that are phosphorylated in a cAMP-dependent manner. These substrates including electron transport chain components and the proapoptotic protein, BAD, suggest a physiological role for cAMP-dependant protein kinase (PKA) in mitochondria. Experiments performed in the Taylor laboratory have identified two new AKAPs (D=AKAPs) are localized to the mitochondria. D-AKAP1 contains 30 amino acids at the NH2- terminus that are sufficient for targeting to the outer mitochondrial membrane, while preliminary studies suggest D-AKAP2 localizes to the matrix. Interestingly, D-AKAP2 also contains two putative PGS domains, sites capable of binding Galpha subunits and leading to speculation that D-AKAP2 may be a scaffolding protein capable of co- localizing multiple signaling molecules. We hypothesize that PKA's role in altering specific mitochondrial functions is mediated by its association with these novel anchoring proteins. To test this hypothesis, this research proposes to identify substrates phosphorylated by PKA as a consequence of this association with the AKAPs. Because of the biochemical differences between RI and RII isoforms, the relative effect of PKA on mitochondrial function may be a function of which isoform is bound to the anchoring protein. This research will thus seek to determine the relative significance of RI versus RII isoform anchoring in mitochondria. Finally, experiments will be performed to determine what proteins, in addition to PKA, are bound to D-AKAP2 in mitochondria.