Programa de Biologia Molecular e Biotecnologia, Instituto de Bioquímica Médica, Universidade Federal do Rio de Janeiro, Rio de Janeiro, Brasil.
PLoS One. 2012;7(7):e40192. doi: 10.1371/journal.pone.0040192. Epub 2012 Jul 3.
The mosquito Aedes aegypti can spread the dengue, chikungunya and yellow fever viruses. Thus, the search for key molecules involved in the mosquito survival represents today a promising vector control strategy. High Mobility Group Box (HMGB) proteins are essential nuclear factors that maintain the high-order structure of chromatin, keeping eukaryotic cells viable. Outside the nucleus, secreted HMGB proteins could alert the innate immune system to foreign antigens and trigger the initiation of host defenses. In this work, we cloned and functionally characterized the HMGB1 protein from Aedes aegypti (AaHMGB1). The AaHMGB1 protein typically consists of two HMG-box DNA binding domains and an acidic C-terminus. Interestingly, AaHMGB1 contains a unique alanine/glutamine-rich (AQ-rich) C-terminal region that seems to be exclusive of dipteran HMGB proteins. AaHMGB1 is localized to the cell nucleus, mainly associated with heterochromatin. Circular dichroism analyses of AaHMGB1 or the C-terminal truncated proteins revealed α-helical structures. We showed that AaHMGB1 can effectively bind and change the topology of DNA, and that the AQ-rich and the C-terminal acidic regions can modulate its ability to promote DNA supercoiling, as well as its preference to bind supercoiled DNA. AaHMGB1 is phosphorylated by PKA and PKC, but not by CK2. Importantly, phosphorylation of AaHMGB1 by PKA or PKC completely abolishes its DNA bending activity. Thus, our study shows that a functional HMGB1 protein occurs in Aedes aegypt and we provide the first description of a HMGB1 protein containing an AQ-rich regulatory C-terminus.
埃及伊蚊可以传播登革热、基孔肯雅热和黄热病病毒。因此,寻找与蚊子生存相关的关键分子代表了一种有前途的病媒控制策略。高迁移率族蛋白(HMGB)是维持染色质高级结构的必需核因子,使真核细胞保持活力。在核外,分泌的 HMGB 蛋白可以向先天免疫系统发出外来抗原的警报,并触发宿主防御的启动。在这项工作中,我们克隆并功能表征了埃及伊蚊(AaHMGB1)的 HMGB1 蛋白。AaHMGB1 蛋白通常由两个 HMG 盒 DNA 结合域和一个酸性 C 末端组成。有趣的是,AaHMGB1 包含一个独特的丙氨酸/谷氨酰胺丰富(AQ 丰富)的 C 末端区域,似乎是双翅目 HMGB 蛋白所独有的。AaHMGB1 定位于细胞核,主要与异染色质相关。AaHMGB1 或 C 末端截断蛋白的圆二色性分析显示出α-螺旋结构。我们表明 AaHMGB1 可以有效地结合并改变 DNA 的拓扑结构,并且 AQ 丰富区和 C 末端酸性区可以调节其促进 DNA 超螺旋化的能力,以及其对超螺旋 DNA 的偏好。AaHMGB1 可被 PKA 和 PKC 磷酸化,但不能被 CK2 磷酸化。重要的是,PKA 或 PKC 对 AaHMGB1 的磷酸化完全消除了其 DNA 弯曲活性。因此,我们的研究表明,埃及伊蚊中存在功能性的 HMGB1 蛋白,并首次描述了含有 AQ 丰富的调节 C 末端的 HMGB1 蛋白。