Rapid Maturation of Glycoprotein Hormone Free α-Subunit (GPHα) and GPHαα Homodimers

Abstract

The dynamics of glycoprotein hormone alpha-subunit (GPHalpha) maturation and GPHalpha alpha homodimer formation were studied in presence (JEG-3 choriocarcinoma cells) and absence (HeLa cells) of hCGbeta. In both cases, the major initially occurring GPHalpha variant in [35S]Met/Cys-labeled cells carried two N-glycans (M(r app) = 22 kDa). Moreover, a mono-N-glycosylated in vivo association-incompetent GPHalpha variant (M(r app) = 18 kDa) was observed. In JEG-3 cells the early 22-kDa GPHalpha either associated with hCGbeta, or showed self-association to yield GPHalpha alpha homodimers, or was later converted into heavily glycosylated large free GPHalpha (M(r app) = 24 kDa). Micro-preparative isolation of intracellular GPHalpha alpha homodimers of JEG-3 cells and their conversion by reduction revealed that they consisted of 22-kDa GPHalpha monomers and not of large free GPHalpha. In HeLa cells, the large free GPHalpha variant was not observed, whereas GPHalpha alpha homodimers were present. Intracellularly, early GPHalpha alpha homodimers (35 kDa) and late variants (JEG-3: 44 kDa, HeLa: 39 kDa) were found. Both cell types secreted 45 kDa GPHalpha alpha homodimers. Large free GPHalpha and GPHalpha alpha homodimers were more rapidly sialylated than hCG alphabeta-heterodimers indicating a sequestration mechanism in the secretory pathway. In GPHalpha alpha homo- as well as hCG alphabeta-heterodimers the subunit interaction site, located on loop 2 of GPHalpha (amino acids 33-42), became immunologically inaccessible indicating similar spatial orientation of GPHalpha in both types of dimers. The studies demonstrate the formation, in vivo dynamics of GPHalpha alpha homodimers, and the pathways of the cellular metabolism of variants of GPHalpha, monoglycosylated GPHalpha and large free GPHalpha.

Human Chorionic Gonadotropin Attracts Regulatory T Cells into the Fetal-Maternal Interface during Early Human Pregnancy

“…Both the hCG␣/␣ and hCG␤/␤ homodimer are biological stable and activate the LH/CG receptor specifically. Additionally, the homodimers can stimulate the signal transduction to the same extent as hCG heterodimers (73)(74)(75)(76). Thus, we assume that Treg migration observed to the ␣-or ␤-subunit alone might be a result of the homodimerization of these subunits followed by an activation of the LH/CG receptor on the Treg.…”

Unconventional Actions of Glycoprotein Hormone Subunits: A Comprehensive Review

“…The free α subunit differs from that associated in GPH heterodimers by presenting additional saccharide branching resulting in a slightly heavier molecular mass ( 104 , 105 ). Homodimers of the α subunit were characterized in both β subunit producing and non-producing cells ( 106 ). The α subunits are more tightly associated in the α homodimer than in the heterodimers ( 106 ).…”

“…Homodimers of the α subunit were characterized in both β subunit producing and non-producing cells ( 106 ). The α subunits are more tightly associated in the α homodimer than in the heterodimers ( 106 ). While the synthesis of large excess of α subunit allows the regulation of the heterodimer production to rely on the rate-limiting β subunit, the released free, heavily glycosylated α subunit may also serve other purposes and possibly, play a role by itself.…”

The Gonadotropin Hormones and Their Receptors∗

“…C, C-terminus; N, N-terminus. [38][39][40][41] In addition to the heterodimeric nature of the hormones, homodimers have also been found for LHβ, 42,43 and for the α subunit. [28][29][30][31][32] Moreover, it was posited that subunit association occurred before the seatbelt was latched by closure of Cys26 and Cys110.…”

“…exchange occurs during the maturation process and that subunit association occurred before completion of protein folding and disulfide formation. 40,41,44 Whether these homodimeric forms of the glycoprotein hormones have any associated bioactivity remains to be shown, although it has been reported that the free α subunit potentiates progesterone-mediated decidualization. In contrast to these reports, a different mechanism in which subunit assembly involved closure of the seatbelt latch, followed by threading of α loop 2, has been proposed.…”