Department of Microbiology and Immunology, University of Michigan Medical School, Ann Arbor, Michigan, USA.
J Virol. 2013 Jun;87(11):6441-54. doi: 10.1128/JVI.00040-13. Epub 2013 Mar 27.
In polarized T cells, HIV-1 Gag localizes to a rear-end protrusion known as the uropod in a multimerization-dependent manner. Gag-laden uropods participate in formation of virological synapses, intercellular contact structures that play a key role in cell-to-cell HIV-1 transmission. Our previous observations suggest that Gag associates with uropod-directed microdomains (UDMs) that eventually comigrate with Gag to the uropod over the cell surface. However, the nature of Gag multimerization required for this movement, the composition of the UDMs, and the molecular determinants for Gag association with these microdomains remain unknown. In this study, we found that Gag multimerization prior to budding but beyond dimerization is necessary for Gag localization to the uropods, indicating that uropod localization occurs early in the assembly process. We also found that prior to membrane curvature, Gag multimers associate with a specific subset of UDMs containing PSGL-1, CD43, and CD44 but not ICAM-1, ICAM-3, or CD59. Notably, upon association, Gag excludes ICAM-3 from this subset of UDMs, revealing an active and selective reorganization of these microdomains by Gag. This specific association between Gag and UDMs is dependent on the highly basic region (HBR) in the Gag matrix (MA) domain. The overall positive charge of the HBR was needed for the interaction with the specific UDM subset, while the exact HBR sequence was not, unlike that seen for MA binding to the plasma membrane phospholipid phosphatidylinositol-(4,5)-bisphosphate [PI(4,5)P2]. Taken together, these findings revealed that HIV-1 Gag associates with specific microdomains present in polarized T cells in an MA-dependent manner, which results in modification of the microdomain constituents.
在极化的 T 细胞中，HIV-1 Gag 以依赖多聚化的方式定位于后端突起，即所谓的尾足。载有 Gag 的尾足参与形成病毒学突触，这是细胞间接触结构，在细胞间 HIV-1 传播中起着关键作用。我们之前的观察结果表明，Gag 与尾足定向微区 (UDM) 相关联，这些微区最终与 Gag 在细胞表面共迁移到尾足。然而，这种运动所需的 Gag 多聚化的性质、UDM 的组成以及 Gag 与这些微区关联的分子决定因素仍不清楚。在这项研究中，我们发现，出芽前但超过二聚化的 Gag 多聚化对于 Gag 定位到尾足是必要的，这表明尾足定位发生在组装过程的早期。我们还发现，在膜曲率发生之前，Gag 多聚体与特定的 UDM 子集相关联，该子集包含 PSGL-1、CD43 和 CD44，但不包含 ICAM-1、ICAM-3 或 CD59。值得注意的是，在关联后，Gag 将 ICAM-3 排除在该 UDM 子集之外，揭示了 Gag 对这些微区的主动和选择性重组。Gag 与 UDM 之间的这种特定关联依赖于 Gag 基质 (MA) 域中的高度碱性区域 (HBR)。HBR 的总正电荷对于与特定 UDM 子集的相互作用是必需的，而不像 MA 与质膜磷脂磷脂酰肌醇-(4,5)-二磷酸 [PI(4,5)P2] 的结合那样需要精确的 HBR 序列。总的来说，这些发现表明，HIV-1 Gag 以 MA 依赖的方式与极化 T 细胞中存在的特定微区相关联，这导致微区成分的修饰。
