Ehrlichia chaffeensis TRP32 Nucleomodulin Function and Localization Is Regulated by NEDD4L-Mediated Ubiquitination.
Ehrlichia chaffeensis TRP32 Nucleomodulin Function and Localization Is Regulated by NEDD4L-Mediated Ubiquitination.
复制标题
DOI：
10.3389/fcimb.2017.00534
复制
发表时间：
2017
影响因子：
5.7
通讯作者：
McBride JW
中科院分区：
文献类型：
--
作者：
Farris TR;Zhu B;Wang JY;McBride JW
恰菲埃里希体是一种专性细胞内细菌，它通过不同的效应子-宿主相互作用重新编程单核吞噬细胞，以调节各种宿主细胞过程。在之前的一项研究中，我们报道了查菲埃里希核调节蛋白 TRP32 在几个生物学相关类别中调节宿主基因的转录，包括细胞分化和增殖。在这项研究中，我们研究了泛素化对 TRP32 功能和宿主细胞内定位的影响。在感染期间和异位表达时，TRP32 在多个赖氨酸残基上被单泛素化和多泛素化。尽管缺乏典型的 PPxY 基序，TRP32 仍与人 HECT E3 泛素 (Ub) 连接酶 NEDD4L 相互作用并被其修饰。 TRP32 泛素化不是由 K48 连接的多聚泛素链实现的，也不是被蛋白酶体降解的；然而，TRP32 被细胞质和细胞核中检测到的 K63 连接的多聚泛素链修饰。 HECT 连接酶抑制剂 heclin 将异位表达的 TRP32 的核下定位从弥漫的核模式改变为花边、点状模式，其中 TRP32 分布在细胞核和核仁的周围。当 TRP32 赖氨酸无效 (K-null) 突变体异位表达时，它表现出与单一赖氨酸突变体（K63R、K93R 和 K123R）相似的表型。然而，与单个赖氨酸突变体或异位表达TRP32的heclin处理的细胞相比，K缺失突变体显示细胞质TRP32的量增加。这些定位的改变与 TRP32 转录抑制功能的变化相对应，在荧光素酶测定中，经 heclin 处理的单个赖氨酸突变体无法抑制 TRP32 靶基因的转录。
Ehrlichia chaffeensis is an obligately intracellular bacterium that reprograms the mononuclear phagocyte through diverse effector-host interactions to modulate various host cell processes. In a previous study, we reported that the E. chaffeensis nucleomodulin TRP32 regulates transcription of host genes in several biologically relevant categories, including cell differentiation and proliferation. In this study, we investigate the effect of ubiquitination on TRP32 function and localization within the host cell. TRP32 is both mono- and polyubiquitinated on multiple lysine residues during infection and when ectopically expressed. Despite lacking a canonical PPxY motif, TRP32 interacted with, and was modified by the human HECT E3 ubiquitin (Ub) ligase NEDD4L. TRP32 ubiquitination was not by K48-linked polyUb chains, nor was it degraded by the proteasome; however, TRP32 was modified by K63-linked polyUb chains detected both in the cytosol and nucleus. HECT ligase inhibitor, heclin, altered the subnuclear localization of ectopically expressed TRP32 from a diffuse nuclear pattern to a lacy, punctate pattern with TRP32 distributed around the periphery of the nucleus and nucleoli. When a TRP32 lysine null (K-null) mutant was ectopically expressed, it exhibited a similar phenotype as single lysine mutants (K63R, K93R, and K123R). However, the K-null mutant showed increased amounts of cytoplasmic TRP32 compared to single lysine mutants or heclin-treated cells ectopically expressing TRP32. These alterations in localization corresponded to changes in TRP32 transcriptional repressor function with heclin-treated and single lysine mutants unable to repress transcription of a TRP32 target genes in a luciferase assay.
影响因子：
3.7
作者：
O'Hara A;Howarth A;Varro A;Dimaline R
通讯作者：
Dimaline R