cGMP-dependent protein kinase I interacts with TRIM39R, a novel Rpp21 domain-containing TRIM protein.
cGMP-dependent protein kinase I interacts with TRIM39R, a novel Rpp21 domain-containing TRIM protein.
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DOI：
10.1152/ajplung.00157.2007
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发表时间：
2007-10
期刊：
影响因子：
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通讯作者：
Jesse D. Roberts;J. Chiche;E. Kolpa;D. Bloch;K. Bloch
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文献类型：
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作者：
Jesse D. Roberts;J. Chiche;E. Kolpa;D. Bloch;K. Bloch
一氧化氮部分通过刺激 cGMP 依赖性蛋白激酶 I (PKGI) 来调节血管平滑肌细胞 (SMC) 细胞骨架动力学和表型。为了鉴定 PKGI 的分子靶标，使用编码 PKGI 催化区域的 cDNA 和人肺 cDNA 文库在酵母中进行了相互作用陷阱筛选。我们鉴定了一个编码假定的 PKGI 相互作用子的 cDNA，它是 TRIM39 的一个新变体，TRIM39 是真正有趣的新基因 (RING) 指蛋白家族的成员。虽然此 TRIM39 变体编码 TRIM39 的 NH(2) 末端环指 (RF)、B 盒和卷曲螺旋 (RBBC) 结构域，而不是完整的 COOH 末端 B30.2 结构域，但此 TRIM39 亚型包含 Rpp21（RNase P 的一个组成部分）的 COOH 末端部分。 RT-PCR 证明 TRIM39 变体（我们称为 TRIM39R）转录于人类胎儿肺和大鼠肺动脉 SMC。使用针对 TRIM39 和 TRIM39R 保守结构域生成的抗体进行的间接免疫荧光揭示了人急性单核细胞白血病 (THP-1) 和人表皮癌细胞系 (HEp-2) 细胞中斑点核内结构中的蛋白质。 PKGI 磷酸化 TRIM39 和 TRIM39R 保守区域中的典型 PKGI/PKA 磷酸化结构域。其他研究表明 PKGI 与酵母细胞中的 TRIM39 的两种亚型相互作用，并磷酸化人类细胞系中 TRIM39 的两种亚型。尽管已观察到 PKGI 与调节细胞骨架功能和基因表达的蛋白质相互作用，但这项研究首次表明 PKGI 与三联基序 (TRIM) 蛋白质相互作用，而三联基序 (TRIM) 蛋白质通常通过不同的分子途径来调节细胞稳态的重要方面。
Nitric oxide modulates vascular smooth muscle cell (SMC) cytoskeletal kinetics and phenotype, in part, by stimulating cGMP-dependent protein kinase I (PKGI). To identify molecular targets of PKGI, an interaction trap screen in yeast was performed using a cDNA encoding the catalytic region of PKGI and a human lung cDNA library. We identified a cDNA that encodes a putative PKGI-interactor that is a novel variant of TRIM39, a member of the really interesting new gene (RING) finger family of proteins. Although this TRIM39 variant encodes the NH(2)-terminal RING finger (RF), B-box, and coiled-coil (RBBC) domains of TRIM39, instead of a complete COOH-terminal B30.2 domain, this TRIM39 isoform contains the COOH-terminal portion of Rpp21, a component of RNase P. RT-PCR demonstrated that the TRIM39 variant, which we refer to as TRIM39R, is transcribed in the human fetal lung and in rat pulmonary artery SMC. Indirect immunofluorescence using an antibody generated against the conserved domains of TRIM39 and TRIM39R revealed the proteins in speckled intranuclear structures in human acute monocytic leukemia (THP-1) and human epidermal carcinoma line (HEp-2) cells. PKGI phosphorylated a typical PKGI/PKA phosphorylation domain in a conserved region of TRIM39 and TRIM39R. Additional studies demonstrated that PKGI interacts with both isoforms of TRIM39 in yeast cells and phosphorylates both isoforms of TRIM39 in human cell lines. Although PKGI has been observed to interact with proteins that regulate cytoskeletal function and gene expression, this investigation shows for the first time that PKGI interacts with tripartite motif (TRIM) proteins, which, through diverse molecular pathways, are often observed to regulate important aspects of cellular homeostasis.