University of Lille, CNRS, UMR 8576 - UGSF - Unité de Glycobiologie Structurale et Fonctionnelle, F-59000 Lille, France.
Laboratoire Croissance, Réparation et Régénération Tissulaires (CRRET), CNRS ERL 9215, Université Paris-Est-Créteil, F-94000 Créteil, France.
Biochim Biophys Acta Gen Subj. 2018 Jul;1862(7):1644-1655. doi: 10.1016/j.bbagen.2018.04.013. Epub 2018 Apr 13.
Heparan sulfate (HS) 3-O-sulfation can be catalysed by seven 3-O-sulfotransferases (HS3STs) in humans, still it is the rarest modification in HS and its biological function is yet misunderstood. HS3ST2 and HS3ST3B exhibit the same activity in vitro. They are however differently expressed in macrophages depending on cell environment, which suggests that they may be involved in distinct cellular processes. Here, we hypothesized that both isozymes might also display distinct subcellular localizations.
The subcellular distribution of HS3ST2 and HS3ST3B was analysed by using overexpression systems in HeLa cells. The localization of endogenous HS3ST2 was confirmed by immunostaining in primary macrophages.
We found that HS3ST3B was only localized in the Golgi apparatus and no difference between full-length enzyme and truncated construct depleted of its catalytic domain was observed. In contrast, HS3ST2 was clearly visualized at the plasma membrane. Its truncated form remained in the Golgi apparatus, meaning that the catalytic domain might support correct addressing of HS3ST2 to cell surface. Moreover, we found a partial co-localization of HS3ST2 with syndecan-2 in HeLa cells and primary macrophages. Silencing the expression of this proteoglycan altered the localization of HS3ST2, which suggests that syndecan-2 is required to address the isozyme outside of the Golgi apparatus.
We demonstrated that HS3ST3B is a Golgi-resident isozyme, while HS3ST2 is addressed to the plasma membrane with syndecan-2.
The membrane localization of HS3ST2 suggests that this enzyme may participate in discrete processes that occur at the cell surface.
在人类中，肝素硫酸（HS）3-O-硫酸化可由七种 3-O-硫酸转移酶（HS3STs）催化，但其仍然是 HS 中最罕见的修饰，其生物学功能尚不清楚。HS3ST2 和 HS3ST3B 在体外具有相同的活性。然而，它们在巨噬细胞中的表达因细胞环境而异，这表明它们可能参与了不同的细胞过程。在这里，我们假设这两种同工酶也可能具有不同的亚细胞定位。
通过在 HeLa 细胞中使用过表达系统分析 HS3ST2 和 HS3ST3B 的亚细胞分布。通过在原代巨噬细胞中进行免疫染色，确认内源性 HS3ST2 的定位。
我们发现 HS3ST3B 仅定位于高尔基体，全长酶和缺失其催化结构域的截短构建体之间没有差异。相比之下，HS3ST2 清楚地定位于质膜上。其截短形式仍保留在高尔基体中，这意味着催化结构域可能支持 HS3ST2 正确定位于细胞表面。此外，我们在 HeLa 细胞和原代巨噬细胞中发现 HS3ST2 与 syndecan-2 部分共定位。沉默该蛋白聚糖的表达改变了 HS3ST2 的定位，这表明 syndecan-2 是将同工酶定位于高尔基体之外所必需的。
我们证明了 HS3ST3B 是一种高尔基体驻留同工酶，而 HS3ST2 与 syndecan-2 一起被定位于质膜。
HS3ST2 的膜定位表明该酶可能参与发生在细胞表面的离散过程。
