Departments of Developmental Biology and Obstetrics and Gynecology, Washington University School of Medicine, St. Louis, Missouri, USA.
PLoS One. 2013 May 29;8(5):e65002. doi: 10.1371/journal.pone.0065002. Print 2013.
The coordinated secretion of LH and FSH are critical for reproductive functions. After translocation into the endoplasmic reticulum (ER), their biosynthetic routes diverge at a determinative step prior to sorting in the regulated (LH) and constitutive (FSH) secretion pathways. Recently, we identified a C-terminal heptapeptide sequence, present only in the LHβ subunit, as a critical signal for entry of the LH dimer into the regulated pathway. We showed that an LHβ mutant lacking the heptapeptide (LHβΔT) assembled more efficiently with the α subunit than wild-type LHβ subunit, and this LHΔT dimer was secreted constitutively. Thus, an association exists between the presence of the C-terminal heptapeptide and sorting of the LH heterodimer to the regulated pathway. To study how this delayed LHβ subunit assembly is related to the trafficking of LH, we exploited the single subunit transfection model in rat somatotrope-derived GH3 cells with the use of immunofluorescence confocal microscopy. The LHβ subunit showed a distinct immunofluorescent localization as compared to the FSHβ subunit and LHβ mutants. The wild-type LHβ subunit exhibited a perinuclear staining corresponding to the ER/nuclear envelope region. In contrast, the wild-type FSHβ subunit and the mutants LHβΔT and LHβL119A displayed no detectable perinuclear staining; only peripheral ER puncta were observed. Also, no perinuclear fluorescence was detected in cells expressing the LH heterodimer. We propose that the C-terminal heptapeptide is responsible for delayed heterodimer assembly within an ER sub-domain of the nuclear envelope, as an early partitioning event necessary for the entrance of LH into the regulated secretory pathway, whereas FSHβ does not traverse the nuclear envelope region. These data suggest that, at least for LH, the molecular decision to enter the regulated secretory pathway is a pre-Golgi event controlled by the novel C-terminal heptapeptide.
LH 和 FSH 的协调分泌对于生殖功能至关重要。在易位到内质网（ER）后，它们的生物合成途径在决定进入调节（LH）和组成型（FSH）分泌途径之前在一个决定性步骤处分叉。最近，我们确定了一个仅存在于 LHβ 亚基中的 C 端七肽序列，作为 LH 二聚体进入调节途径的关键信号。我们表明，缺乏七肽的 LHβ 突变体（LHβΔT）比野生型 LHβ 亚基更有效地与α 亚基组装，并且这种 LHΔT 二聚体被组成型分泌。因此，C 端七肽的存在与 LH 异二聚体到调节途径的分拣之间存在关联。为了研究这种延迟的 LHβ 亚基组装与 LH 的运输之间的关系，我们利用免疫荧光共聚焦显微镜在大鼠生长激素细胞 GH3 中转染单个亚基模型进行了研究。与 FSHβ 亚基和 LHβ 突变体相比，LHβ 亚基表现出明显不同的免疫荧光定位。野生型 LHβ 亚基表现出核周染色，对应于内质网/核膜区域。相比之下，野生型 FSHβ 亚基和突变体 LHβΔT 和 LHβL119A 则没有可检测到的核周染色；仅观察到周围内质网点状结构。此外，表达 LH 异二聚体的细胞中也没有检测到核周荧光。我们提出，C 端七肽负责在核膜的内质网亚区中延迟异二聚体组装，作为 LH 进入调节分泌途径的早期分隔事件，而 FSHβ 不穿过核膜区域。这些数据表明，至少对于 LH 而言，进入调节分泌途径的分子决定是由新型 C 端七肽控制的高尔基体前事件。
