Kim Kami
Departments of Medicine, Pathology and Microbiology & Immunology, Albert Einstein College of Medicine, Bronx, New York, USA
mBio. 2017 Jul 25;8(4):e01076-17. doi: 10.1128/mBio.01076-17.
The cyclic GMP-dependent protein kinase (PKG) of apicomplexan parasites is essential for secretion of micronemes and host cell invasion and egress. Both kinase specificity and localization can determine which substrates are phosphorylated. The functions of plasma membrane and cytosolic PKG isoforms of were unknown because of difficulties precisely manipulating expression of essential genes. Brown et al. (K. M. Brown, S. Long, and L. D. Sibley, mBio 8:e00375-17, https://doi.org/10.1128/mBio.00375-17) adapted the auxin-inducible degron (AID) system for conditional expression of proteins. AID, in combination with clustered regularly interspaced short palindromic repeat (CRISPR)-Cas9 gene editing, facilitated creation of a panel of PKG mutants to demonstrate that the membrane association via acylation of PKG is critical for its essential functions in tachyzoites. The cytosolic form of PKG is not sufficient for viability and is dispensable. These studies illuminate a critical role for targeting of kinase complexes for parasite viability. The AID system enables rapid, conditional regulation of protein expression that expands the molecular toolbox of .
顶复门寄生虫的环磷酸鸟苷依赖性蛋白激酶(PKG)对于微线体的分泌以及宿主细胞的入侵和逸出至关重要。激酶特异性和定位均可决定哪些底物会被磷酸化。由于难以精确操控必需基因的表达,疟原虫质膜和胞质PKG亚型的功能尚不清楚。布朗等人(K.M.布朗、S.朗和L.D.西布利,《mBio》8:e00375 - 17,https://doi.org/10.1128/mBio.00375 - 17)采用生长素诱导降解子(AID)系统实现了蛋白质的条件性表达。AID与成簇规律间隔短回文重复序列(CRISPR)-Cas9基因编辑相结合,有助于创建一组PKG突变体,以证明通过PKG酰化作用实现的膜结合对于其在速殖子中的基本功能至关重要。胞质形式的PKG不足以维持生存,且并非必需。这些研究揭示了激酶复合物靶向作用对寄生虫生存能力的关键作用。AID系统能够快速、条件性地调节蛋白质表达,从而扩展了疟原虫的分子工具库。